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Muscle development takes place by a series of regulated steps, beginning with the specification of the muscle forming germ layer, the mesoderm. From the mesoderm originate different muscles, each one distinct in its shape, size, attachment and innervation. Muscles achieve their unique identities by utilising genetic information that is common to the development of all muscles as well as specialised information that determines their distinct properties. For this, both mesoderm autonomous information and inductive signals imparted by neighbouring tissues like epidermis and nervous system are required. To understand the genetic, the cellular and the molecular mechanisms that mediate these complex interactions, such that a functional muscle pattern emerges, is a challenge in developmental biology. Studies in different animal systems, both vertebrate and invertebrate, have provided useful insights into myogenesis. The fruitfly, Drosophila melanogaster, being highly amenable to genetic dissection, has proved to be a useful tool for such studies and many significant aspects of myogenesis have been described in the fly embryo. Our laboratory has used the adult flight muscles of Drosophila to understand the mechanisms that govern events of myogenesis such as cell fate specification, mesoderm diversification and muscle patterning. Here we review current developments in this area. Cell Fate Specification and Differentiation in the Embryo Early Mesodermal Subdivision Very early in embryonic development, cells that will form mesoderm and its derivatives express twist (twi) (Thisse et al. 1988), invaginate into the embryo, spread as an epithelial sheet closely apposed to the external epidermis and divide (Leptin & Grunewald 1990). This contact is important because secreted products of patterning genes like decapentaplegic (dpp) signal from the epidermis to pattern the mesoderm below. dpp expressed in a dorsal band of ectodermal cells maintains expression of the mesodermal marker tinman (tin) and represses ventrally expressed genes such as pox meso (Staehling-Hampton et al. 1994, Frasch 1995). The mesoderm also undergoes partitioning along the anterior-posterior axis by action of segmentation genes even skipped (eve) and sloppy paired (slp) (Azpiazu et al. 1996, Riechmann et al. 1997). Consequently, a refinement of Twi expression occurs into a modulated pattern in different mesodermal anlages. Wingless (Wg) amplifies the distinctions between cells of the eve and slp domains by maintaining high levels of Twi in the slp domain. These cells form somatic muscles (Baylies et al. 1995). Cells of the eve domain become cardiac muscles and visceral muscles (Azpiazu & Frasch 1993, Bate & Rushton 1993, Yin et al. 1997, Riechmann et al. 1997).
a, b Gene regulation by combinatorial logic. Specification of Eve expressing progenitor cell which occurs through convergence of multiple signals at a single enhancer of the gene is shown in the schematic in a and b. Wg and Dpp signal along the dorsal epidermis to act on the underlying mesodermal cells that express Twi and Tin. This defines a cluster of ‘competent’ cells expressing the transcription factor L’sc. Signalling through the Wg and Dpp pathways, along with the RTK pathways involving DER and Htl eventually produce just one Eve expressing cell [a]. Proteins downstream of Wg, Dpp and RTK signalling pathways combine with the intrinsic factors Twi and Tin and converge at the Eve-enhancer [b] to regulate its mesoderm specific expression and specification of the progenitor (Ghazi and VijayRaghavan, 2000). [c]. Specification of adult muscle founders: Adult muscle founders form as siblings of embryonic muscle founders, as a consequence of asymmetric cell division of progenitors. In some instances, as in the case of the Eve-expressing progenitor in the thoracic segments shown here, when the progenitor divides, one of the progeny retains expression of Eve [red nucleus] and the other loses it [white nucleus]. This is determined by the distribution of the cytoplasmic protein Nb [green crescent] between the two progeny. The descendent that gets Nb undergoes inactivation of N activity by Nb, continues maintaining Eve expression, and becomes an embryonic muscle founder. Its sibling which does not get Nb continues to have active N signalling, loses Eve and becomes the founder of an adult muscle.
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Proc. Indian natn Sci Acad. B69 No.5 pp 691-702 (2003)
*Corresponding author: E-mail: vijay@ncbs.res.in; Tel: (080) 3636420-432 Ext 3003; Fax: 3636675, 3636662
Muscle Development in Drosophila
ARJUMAND GHAZI and K VIJAYRAGHAVAN*
National Centre for Biological Sciences, TIFR Centre, UAS-GKVK Campus, Bangalore 560065
(Received on 3 September 2001; Accepted after revision on 3 April 2002)
Muscle development takes place by a series of regulated steps, beginning with the specification of the
muscle forming germ layer, the mesoderm. From the mesoderm originate different muscles, each one
distinct in its shape, size, attachment and innervation. Muscles achieve their unique identities by
utilising genetic information that is common to the development of all muscles as well as specialised
information that determines their distinct properties. For this, both mesoderm autonomous information
and inductive signals imparted by neighbouring tissues like epidermis and nervous system are required.
To understand the genetic, the cellular and the molecular mechanisms that mediate these complex
interactions, such that a functional muscle pattern emerges, is a challenge in developmental biology.
Studies in different animal systems, both vertebrate and invertebrate, have provided useful insights
into myogenesis. The fruitfly, Drosophila melanogaster, being highly amenable to genetic dissection,
has proved to be a useful tool for such studies and many significant aspects of myogenesis have been
described in the fly embryo. Our laboratory has used the adult flight muscles of Drosophila to
understand the mechanisms that govern events of myogenesis such as cell fate specification, mesoderm
diversification and muscle patterning. Here we review current developments in this area.
Key Words: Drosophila, Muscle development, Genetic information, Muscle pattern, Fruit fly embryo
Cell Fate Specification and Differentiation in
the Embryo
Early Mesodermal Subdivision
Very early in embryonic development, cells that will
form mesoderm and its derivatives express twist
(twi) (Thisse et al. 1988), invaginate into the embryo,
spread as an epithelial sheet closely apposed to the
external epidermis and divide (Leptin & Grunewald
1990). This contact is important because secreted
products of patterning genes like decapentaplegic
(dpp) signal from the epidermis to pattern the
mesoderm below. dpp expressed in a dorsal band of
ectodermal cells maintains expression of the
mesodermal marker tinman (tin) and represses
ventrally expressed genes such as pox meso
(Staehling-Hampton et al. 1994, Frasch 1995). The
mesoderm also undergoes partitioning along the
anterior-posterior axis by action of segmentation
genes even skipped (eve) and sloppy paired (slp)
(Azpiazu et al. 1996, Riechmann et al. 1997).
Consequently, a refinement of Twi expression occurs
into a modulated pattern in different mesodermal
anlages. Wingless (Wg) amplifies the distinctions
between cells of the eve and slp domains by
maintaining high levels of Twi in the slp domain.
These cells form somatic muscles (Baylies et al. 1995).
Cells of the eve domain become cardiac muscles and
visceral muscles (Azpiazu & Frasch 1993, Bate &
Rushton 1993, Yin et al. 1997, Riechmann et al. 1997).
Early Muscle Patterning: Founder and Feeders
Midway through embryogenesis, mesodermal cells
destined to form somatic muscles lose Twi
expression, fuse and differentiate to form muscle
fibres (Bate 1990, Bate et al. 1991). Unlike
vertebrates where aggregates of muscle fibres
constitute a single muscle, in the Drosophila
embryo, each muscle is a single, multinucleate fibre,
unique in its position, size, sites of attachment and
692 Arjumand Ghazi and K Vijayraghavan
patterns of innervation (Bate 1993, Bernstein et al.
1993). The development of each muscle fibre in the
embryo is seeded by a specialised myoblast called
the Founder Cell. The founder cell fuses with
neighbouring fusion competent myoblasts, the
Feeder Cells, entraining them to its pattern of
gene expression and forming a syncytial muscle
(Bate 1990, 1993). Founders, thus, are privileged
cells that are self sufficient in their access to genetic
information required to complete myogenesis and
form a specific muscle, as against the naive
feeders that cannot form muscles independent of
the founders. In the absence of fusion, founders
differentiate miniature, mononucleate muscles at
proper positions, normal in all aspects of
myogenesis except their size. Feeders in such a
situation fail to form muscle and remain
undifferentiated (Rushton et al. 1995).
Founder Cell Specification
Founder cells are characterised by the expression of
specific muscle identity genes (Frasch 1999),
encoding transcription factors, including S59,
apterous (ap), Kruppel (Kr), vestigial (vg), ladybird
(lb) etc., (Dohrmann et al. 1990, Bourgouin et al.
1992, Ruiz-Gomez et al. 1997, Jagla et al. 1998). The
overlapping expression of these genes in different
sets of muscle, and functional analysis in genetic
experiments, has led to the hypothesis that
individual muscles are specified by defined
combinatorial codes of identity genes. Mutations in
muscle identity genes have been shown to result in
loss of, as well as, transformation in identities of
specific muscles (Ruiz-Gomez & Bate 1997).
Recently, loss of function of nautilus (nau)- also
called slouch and S59- which is the Drosophila
homologue of a very important family of vertebrate
genes controlling multiple stages of myogenesis, has
shown its requirement of a small subset of
embryonic muscles (Balagopalan et al. 2001).
Founder cells are division products of Progenitor
cells which are chosen from equivalent groups of
mesodermal cells (Carmena et al. 1995, Ruiz-Gomez
& Bate 1997) and express the proneural gene lethal
of scute (lsc) (Carmena et al. 1995). This selection is
mediated by intrinsic mesoderm encoded
information as well as cues arising from regions of
the epidermis overlying that part of the mesoderm.
This has been clearly illustrated in the case of
progenitors expressing Eve. Wg and Dpp from the
epidermis activate receptor tyrosine kinase (RTK)
pathways {the Drosophila Egf receptor (DER) and
the FGF receptor encoded by heartless (htl)}, and
thus Ras signalling, in the equivalence group.
Consequently, a single cell gets selected, from this
Equivalent Cluster of Eve expressing cells, to
continue expressing Eve. This becomes a
Progenitor. The remaining cells lose Eve and adopt
feeder fate (Carmena et al. 1998). This convergence
of multiple signals in regulation of eve expression
and determination of progenitor identity occurs at
a single enhancer in the eve promoter region. It has
binding sites for, and responds to, the extrinsic Wg,
Dpp and RTK signals, as well as tissue specific
proteins Tin and Twi, suggesting that fates in the
mesoderm are determined by a combination of
extrinsic cues and the developmental histories of
cells (figure 1a, b, Halfon et al. 2000). Once the
proneural cluster is formed, the singling out of a
single progenitor is also dependent on the Notch
(N) mediated process of Lateral Inhibition (Corbin
et al. 1991, Bate et al. 1993, Baker & Schubiger 1996).
Once the identity of a progenitor is determined, its
asymmetric division contributes to the
diversification of individual muscle fates. An
important consequence of this asymmetric division
appears to be the differential maintenance of
expression of muscle identity genes (Kr, for
instance), in only one of the two descendent
founders (Ruiz-Gomez et al. 1997). The generation
of this distinction depends on the cytoplasmic
protein Numb (Nb) and determines founder fate
(Ruiz-Gomez et al. 1997, Carmena et al. 1998). Nb is
retained in one of the progenitor descendants,
causes downregulation of N (Frise et al. 1996) and
maintains expression of the muscle identity gene to
form a founder for a specific muscle. It is lost from
the other progeny, which allows continued N activity
and subsequent loss of expression of the muscle
identity gene of the progenitor, generating a founder
for another muscle (figure 1c). Mechanisms
controlling asymmetric segregation of Nb are not yet
known, however, the product of the inscutable (insc)
gene appears to be one of the key components.
Altogether, the observed involvement of lineage
genes such as insc, nb, and N may occur during the
asymmetric division of all muscle progenitors.
693
Muscle Development in Drosophila
Asymmetry of Muscle Fusion
With founder cell specification, Drosophila
myoblasts become segregated into two types of
cells, founders and feeders. Founders are
competent only to fuse with feeders and vice
versa. The two cell types cannot fuse with
members of their own class. Identification of two
new genes, dumb-founded (duf) and sticks-and-
stones (sns), (Ruiz-Gomez et al. 2000, Bour et al.
2000) suggest that an asymmetric distribution of
cell-cell interaction molecules might be implicated
in this asymmetry generation. Both Duf and Sns
are novel members of the Immunoglobulin
superfamily and seem to act at the earliest steps of
muscle fusion. Duf is expressed in the founders
but not in the feeders, whereas Sns is specifically
expressed in the feeders. Both are crucial for
fusion. By contrast, molecules described earlier
like Drosophila Rac1 (Drac1), Myoblast city (Mbl/
Dock180) and Blown fuse (Blow) are intracellular
and not known to be asymmetrically expressed in
the two myoblast populations (Luo et al. 1994,
Doberstein et al. 1997, Erickson et al. 1997).
However, how muscle identity genes influence the
extent of muscle fusion is not clear.
Founder cells have also been shown to be
sources of cue(s) required to trigger defasciculation
and targeted growth of motor axons that innervate
their unique muscles. A single founder myoblast is
found to trigger the defasciculation of an entire
nerve branch, suggesting that the muscle field is
structured into sets of muscles each expressing a
common defasciculation cue for a given nerve
branch (Landgraf et al. 1999).
Adult Flight Muscle Specification and
Differentiation
Specification of Adult Muscle Precursors
During the asymmetric division of progenitors, not
all descendent cells get specified to form founders
of larval muscles. In some regions, like the
abdomen and thoracic segments, one of the
progeny gets allocated to form precursors of adult
muscles. The progenitor descendent that retains
Nb, and the muscle identity gene expression, gets
committed to become an embryonic founder. Its
sibling which does not inherit Nb and which has
active N signalling continues to express Twi and
postpones differentiation to become an adult muscle
Figure 1a, b Gene regulation by combinatorial logic.
Specification of Eve expressing progenitor cell which
occurs through convergence of multiple signals at a
single enhancer of the gene is shown in the schematic
in a and b. Wg and Dpp signal along the dorsal
epidermis to act on the underlying mesodermal cells
that express Twi and Tin. This defines a cluster of
competent cells expressing the transcription factor
Lsc. Signalling through the Wg and Dpp pathways,
along with the RTK pathways involving DER and Htl
eventually produce just one Eve expressing cell [a].
Proteins downstream of Wg, Dpp and RTK signalling
pathways combine with the intrinsic factors Twi and
Tin and converge at the Eve- enhancer [b] to regulate
its mesoderm specific expression and specification of
the progenitor (Ghazi and VijayRaghavan, 2000). [c].
Specification of adult muscle founders: Adult muscle
founders form as siblings of embryonic muscle
founders, as a consequence of asymmetric cell division
of progenitors. In some instances, as in the case of
the Eve- expressing progenitor in the thoracic
segments shown here, when the progenitor divides,
one of the progeny retains expression of Eve [red
nucleus] and the other loses it [white nucleus]. This
is determined by the distribution of the cytoplasmic
protein Nb [green crescent] between the two progeny.
The descendent that gets Nb undergoes inactivation
of N activity by Nb, continues maintaining Eve
expression, and becomes an embryonic muscle
founder. Its sibling which does not get Nb continues
to have active N signalling, loses Eve and becomes
the founder of an adult muscle.
694 Arjumand Ghazi and K Vijayraghavan
Figure 2 Schematic representation of thoracic flight
myogenesis:
a] Myoblasts [blue dots] on the wing disc are associated
with the presumptive notum and proliferate during
larval life. sr expression [red regions indicated by
arrows] on the presumptive notum marks the future
epidermal attachment sites of thoracic muscles. IFM
development is represented in B, C and D. In these
panels anterior is to the top and dorsal midline to the
right; b] Myoblasts migrate on the everting disc
epithelium [schematic represents 7-12h APF] onto three
larval muscles [green fibres] that escape histolysis and
serve as templates that attach to sr expressing
epidermal attachment sites [red spots]; c] As myoblasts
fuse to the templates, they begin to split longitudinally
between 14-18h APF. By 19h APF, six dorsal
longitudinal muscles [DLMs] are in place; d] shows
IFMs in an adult heminotum with their attachment
sites. Six DLMs [dark green] run antero-posteriorly
and attach to sr expressing attachment domains [red].
DVMs run dorsoventrally [light green; e] Direct flight
muscles [DFMs] are also derived from wing disc
associated myoblasts. Muscles 49 and 51-55 followed
in our study are shown in blue. In panels C, D and E,
the expression of ap, has been indicated with bright
orange arrows. Wing disc shows low levels of ap
expression through out the presumptive notum region.
In all panels except a, wing bud [b, c] and adult wing
nub [d, e] are indicated by asterisks. Schematic from
Ghazi et al. 2000.
precursor (Ruiz-Gomez & Bate 1997). Not only are
adult precursors produced as siblings of embryonic
founders, they are also produced at precisely
analogous geographical locations where ultimately
they organise an adult pattern in the pupa.
Wing Disc Associated Myoblasts
By the end of embryogenesis, Twi expression
persists in a handful of mesodermal cells, that are
precursors of adult muscles and divide actively to
produce pools of myoblasts in thoracic segments
and adhere to the wing imaginal disc (Bate et al.
1991). Cell fate decisions in the wing disc distinguish
the future wing from the body wall (notum). This
decision depends on the antagonistic interactions of
Wg and DER. Wg specifies the wing primordium
and DER signalling, stimulated by its ligand Vein
(Vn), directs cells to become notum (Wang et al.
2000, Baonza et al. 2000). Adult muscle precursors
associated with the wing disc are located in the
presumptive notum region and continue to remain
restricted to this location till metamorphosis
(figure 2a). Besides Twi, several other genes have
been shown to be expressed in these myoblasts.
These include cut (ct), vg, scalloped (sd), htl and the
Drosophila homologue of the vertebrate myocyte
enhancer factor (Dmef- 2) (Campbell et al. 1991,
1992, Blochlinger et al. 1993, Emori & Saigo 1993,
Williams et al. 1993, 1994, Ng et al. 1996, Couso et al.
1995, Ranganayakulu et al. 1995).
The entire population of wing disc myoblasts
was believed to be a homogenous pool of cells,
which during pupation adopted diverse muscle
fates, in response to unknown autonomous or
extrinsic cues. Similarly, presence of adult
counterparts of embryonic muscle founders was
speculated upon but not confirmed. Recent reports
substantiate the presence of adult muscle founders
and dispel the notion of the wing disc harbouring an
identical pool of myoblasts. One of the members of
the Enhancer of split Complex {E(spl-C)} (nuclear
effectors of N signalling), E(Spl-C)m6, is found to
accumulate in a small subset of wing disc myoblasts
and the expression relies on N activity (Lai et al.
2000). This suggests that distinctions exist between
myoblasts on the disc itself. There has also been
evidence for the presence of a morphologically
distinct class of disc associated myoblasts that
prefigure the formation of some of the adult flight
695
Muscle Development in Drosophila
1991, Barthmaier & Fyrberg 1995). The DLMs and
the DVMs have different developmental histories:
DLMs develop using persistent larval muscles as
scaffolds (Shatoury 1956, Crossley 1978, Fernandes
et al. 1991) and DVMs develop by de novo fusion of
myoblasts (Fernandes et al. 1991). During early
pupation, as the wing disc evaginates, the disc
myoblasts migrate on to the developing dorsal
mesothorax at sites of muscle formation (Bate et al.
1991, Fernandes & VijayRaghavan 1993). At this
time, when larval muscles undergo histolysis, three
muscles -the dorsal oblique 1,2 and 3- persist.
Myoblasts swarm over these templates, fuse with
them and result in their splitting into six fibres to form
the final pattern of six DLMs observed in the adult.
Groups of the same myoblasts organise themselves at
positions where DVMs develop and undergo fusion
to form adult DVMs (Fernandes et al. 1991). Figure 2
represents the events of flight myogenesis.
Splitting of larval templates to form six fibres is
an important event in IFM development and
imaginal myoblasts are directly involved in the
process. If wing discs are depleted of their myoblasts
splitting fails and muscles degenerate (Roy &
VijayRaghavan 1998). The larval muscles themselves
are required for regulating the proper number of
DLM fibres: on template ablation, DLM development
proceeds normally but the number of fibres varies
considerably (Fernandes & Keshishian 1996). Flight
myogenesis also progresses in close synchrony with
the development of innervation. Laser ablation of
their motor nerves does not substantially affect DLM
development but denervation disrupts DVM
development (Fernandes & Keshishian 1998). This
also indicates inherent differences in the way DLMs
and DVMs are patterned, with the DVMs relying on
neural cues and DLMs on persistent larval templates.
The specification of DFM fate is found to depend on
the gene apterous (ap) which gets expressed in groups
of myoblasts that go onto form the DFM (Ghazi et al.
2000), whereas myoblasts that do not express ap give
rise to the IFMs.
Flight Muscle Differentiation
As fusion of myoblasts to larval templates
proceeds, expression of Twi from these cells
declines and expression of differentiation markers,
like Erectwing (Ewg), commences in developing
muscles (Roy & VijayRaghavan 1998). This
muscles too (Rivlin et al. 2000). These reports
indicate that larval stages are not quiescent carriers
of adult muscle precursors and that besides
proliferation, active signalling and cell fate
determination are undergone by these cells, before
metamorphosis commences. The epidermal cells
that serve as attachment sites for flight muscles are
specified at the third larval instar stage itself
(described ahead). The reason for this early
segregation is not clear but the fact that myoblasts
remain in close association with these cells during
their residence on the prospective notum suggests
that important patterning information could be
exchanged between attachment sites and myoblasts.
Besides wing disc myoblasts, there are groups of
myoblasts that remain associated with peripheral
nerves that innervate larval thoracic muscles (Bate
et al. 1991). Whether these cells represent a special
class of myoblasts that are different from the disc
myoblasts and whether they contribute to definite
sets of muscles is unclear.
Thoracic Flight Muscles
Two kinds of flight muscles are present in the adult
notum. These are the fibrillar indirect flight muscles
(IFMs; figure 2d) and the tubular direct flight
muscles (DFMs; figure 2e). Both the IFMs and the
DFMs derive from the wing disc myoblasts, whereas
myoblasts associated with the mesothoracic leg discs
contribute to a large muscle called the tergal
depressor of trochanter (TDT) or jump muscle
(Crossley 1978, Fernandes et al. 1991). Despite a
common origin the IFMs and the DFMs differ from
each other in their morphology, physiology and
molecular markers displayed. But unlike the IFMs,
not much is known about the development of the
DFMs, because of their small size, and lack of markers
that specifically label their developmental stages.
Development of the two subsets of IFMs, the
dorsal longitudinal muscles (DLMs) and the
dorsoventral muscles (DVMs) has been described
extensively (Fernandes et al. 1991, Fernandes &
VijayRaghavan 1993, Anant et al. 1998, Roy et al. 1997,
Roy & VijayRaghavan 1998, 1999). The availability of
several reporter genes and antibody probes that
label different stages of development of these
muscles, their innervation, their differentiation and
their attachment, has provided insights into the
mechanism of their development (Fernandes et al.
696 Arjumand Ghazi and K Vijayraghavan
downregulation of Twi is an essential step for
normal differentiation, which also requires
N. Overexpression of an activated form of N can
result in an abnormal persistence in Twi expression
in nuclei of differentiating fibres suggesting that
N is an important signal for maintenance of Twi
expression in adult precursors which in turn
influences differentiation (Anant et al. 1998).
Patterning Mechanisms: Attachment of Muscles
Normal motor function requires that muscles form,
and maintain, stable muscle attachments at correct
skeletal locations. While little is known about
mechanisms of muscle attachment in vertebrates,
genetic studies in the fruitfly are beginning to reveal
its cellular and molecular basis. Unlike vertebrates,
where muscles attach to cartilage or bone with the
help of tendons, invertebrate muscles attach to
epidermal Tendon Cells (TCs). The ectoderm is
thought to provide positional information for correct
migration and arrangement of different types of
myotubes. Early experiments in the mealworm
Tenebrio had shown that rotation of pieces of the
ectoderm can induce changes in somatic muscle
patterning (Williams & Caveney 1980a,b, Williams
et al. 1984). Tissue culture experiments in the
Drosophila embryo illustrated that the cells along the
segment borders, to which muscles attach, provide
guidance cues that influence muscle migration and
attachment (Volk & VijayRaghavan 1994).
TCs differentiate from early Tendon Precursor
Cells (TPCs) (also called Epidermal Muscle
Attachment {EMA} cells). Following fusion, each
developing fibre extends its leading edges (filopodia)
in both directions towards the TPCs (Bate 1990).
Establishment of an attachment between the
approaching myotube and a TPC is followed by
arrest of myotube extension. Only myotube bound
cells differentiate into the fusiform TCs (Becker et al.
1997). Forces exerted by the muscles are transmitted
to the cuticle through a series of muscle and TC
specializations at the MyoTendinous Junction
(MTJ), where muscle and tendon cell membranes
interdigitate extensively, each secured by specialized
junctions to the intervening ExtraCellular Matrix
(ECM). Differentiation of the MTJs requires a
molecular conversation between muscle and TCs,
interruption of which prevents effective muscle
attachment (Becker et al. 1997).
Determination of Tendon Cell Fate
Pools of epidermal cells acquire the competence to
attach muscles and become TPCs by expression of
the gene stripe (sr). sr encodes a DNA- binding
protein with triple Zn finger domains. It is a
Drosophila member of the early growth response
(egr) family of transcription factors and is
homologous to vertebrate Egr-1 and Egr-2
proteins (Volk & VijayRaghavan 1994, Lee et al.
1995, Frommer et al. 1996). sr is crucial and
sufficient for induction of an array of tendon
specific genes, including its own expression. In sr
mutant embryos, expression of most tendon
specific genes is drastically diminished and
muscles ignore their normal attachment sites, insert
at alternate positions and often degenerate.
Conversely, generation of ectopic TCs by ectopic Sr
expression leads to attraction of myotubes towards
these new target cells. TPCs thus provide essential
attractive cues that direct muscle extension and also
arrest further filopodia formation (Volk &
VijayRaghavan 1994, Frommer et al. 1996, Becker
et al. 1997, Vorbruggen & Jackle 1997).
The differentiation of TCs is biphasic. The
initial phase is muscle independent, characterised
by expression of Sr at low levels in large groups of
cells and by expression of markers like Groovin,
Alien etc (Goubeaud et al. 1996, Becker et al. 1997,
Strumpf & Volk 1998). These are postulated to
require low levels of Sr for their induction. The
second phase is muscle dependent, triggered by
musculo- epithelial contact and marked by high
levels of Sr in muscle attached TCs. Delilah (Del)
and β-1-Tubulin are the markers of this late stage
and probably require high levels of Sr for
induction (Armand et al. 1994, Buttgereit 1996).
Initial determination of TPC identity is induced by
positional cues that pattern the entire embryonic
ectoderm. Embryos mutant for the segment
polarity genes like patched, naked, lines, wg etc.,
show impaired patterns of sr expression (Volk &
VijayRaghavan 1994). At the segment borders, sr
expressing TPCs are specified at precise locations
as a result of interaction of repressive wg and
inductive hh signals. A single enhancer in the sr
promoter region has binding sites for, and
responds to, both signals to determine the sr
expression domain (Piepenburg et al. 2000).
697
Muscle Development in Drosophila
Molecular Regulation of TC Differentiation
While TPCs are defined autonomously in the
ectoderm, their terminal differentiation into TCs is
induced by muscle attachment. Transmission of this
signal from muscle to the epidermis involves the DER
pathway and its ligand Vn. vn mRNA is present at
high levels in all muscles before they adhere to the
epidermis, but the protein is highly concentrated
specifically at MTJs. This localized accumulation of
Vn is critical for triggering the maturation of the
single muscle bound TC and strongly activates the
DER pathway in the TPC. Activation of Ras signalling
follows this, followed by expression of TC specific
differentiation markers like β-1-Tubulin and Del. Vn
is, thus, a muscle derived signal that activate DER in
TCs for their differentiation (Schnepp et al. 1996,
Yarnitzky et al. 1997, 1998). Clues to the restricted
localization of Vn to the MTJs emerge from studies
with the gene kakapo (kak; earlier called Groovin).
Kak is an intracellular protein expressed mainly along
the TPC plasma membrane, presumably associated
with various cytoskeletal elements within the TC and
may directly regulate the extracellular localization of
Vn. The precise mechanism by which this is brought
about is not clear (Strumpf & Volk 1998, Gregory &
Brown 1998).
The cellular interactions between muscle and
tendon cell that allow the terminal differentiation of
the latter involve modulating Sr levels before and
after muscle binding. The mechanism that regulates
the transition from TPCs to TCs by effecting Sr
postranscriptionally has been shown to rely on the
gene held out wings (how). (Zaffran et al. 1997,
Baehrecke 1997). The two protein variants encoded
by how are differentially distributed in the TPCs and
TCs. How Long {How(L)} is nucleus restricted and
How Short {How(S)} is present both in the cytoplasm
as well as the nucleus of the tendon cells. Sr is a
target of How which binds sr mRNA at the 3 UTR.
At the initial stage How(L) is the predominant form
expressed so that Sr is maintained at low levels and
the cell is maintained in a partially differentiated
state. Activation of the DER pathway in the TC, on
muscle binding, causes increase in levels of How(S)
which presumably competes with How(L) for
binding sr mRNA, thereby leading to increased Sr
nuclear export. The resulting increase in Sr protein
levels lead to a terminal differentiation of TC
(figure 3; Nabel-Rosen et al. 1999). TPCs that do
not undergo muscle attachment lose sr expression
eventually. Continued muscle contact is crucial for
maintenance of high levels of Sr in the TCs and a loss
of muscle is known to cause an eradication of Sr from
the TC (Becker et al. 1997).
Ultrastructure of Myotendinous Junctions
Once in contact, muscle and tendon cells must
synthesize and localize the structural components of
the MTJ such that the junction can withstand
mechanical stress. In the mature MTJ of the
Drosophila embryo, membranes of the muscle and
tendon cells are highly interdigitated to increase
surface area and linked indirectly via
hemidesmosomal or Hemi Adherens Junctions
(HAJs) that anchor each cell to a common specialized
ECM (Tepass & Hartenstein 1994). The HAJs found
along the basal surface of the contacted epidermal cell
have a layer of electron dense material on the inside
Figure 3 Differentiation of tendon precursor cells
into tendon cells. Tendon precursor cells [orange]
exhibit the nuclear How[L] form [red squares] of
how, which binds sr mRNA and prevents its
translation. On contact with a myofibre [pink], DER
signalling [blue] pathway gets activated in the
precursor. This is dependent on the accumulation of
the DER ligand Vn [orange] at the muscle-epidermal
junction. As a consequence of DER activation the
levels of the nuclear cum cytoplasmic form of how,
How[S] [red triangles] increases in the cell. How[S]
competes with How[L] in binding sr mRNA and
results in its increased export from the nucleus
[yellow]. This causes an increased translation of Sr and
results in differentiation of the precursor into a tendon
cell. This schematic is modified from Volk 1999.
698 Arjumand Ghazi and K Vijayraghavan
into which cytoskeletal elements insert: actin
filaments of the contractile apparatus in the muscle
and microtubules in the epidermis. The muscle
membrane is linked to the microfilaments via
modified terminal Z bands. The basal membrane of
the TC is linked via numerous microtubules to
specialized anchors (tonofibrillae) embedded in the
cuticle, an apical secretion of the same cell (Caveney
1969, Prokop et al. 1998). The ECM present between
the epidermal and mesodermal components of the
MTJ displays a large number of proteins, which have
been reported in the Drosophila embryo. The most
well characterised of these are the Position Specific
(PS) Integrins. PS integrins are dimeric- they share a
common β-subunit and differ in the α-subunits. The
β-subunit is encoded by myospheroid (mys),
mutations in which cause a muscle detachment
phenotype in embryos (Newman & Wright 1981).
α-PS2 is encoded by the gene inflated (if) and
mutations do cause muscle detachment phenotypes,
though not as strong as in mys (Brown 1994). The
gene encoding α-PS1 is multiple edematous wing
(mew) (Brower et al. 1995a,b). PS1 (αPS1βPS) is
expressed at the basal surface of the TCs and PS2
(αPS2βPS) localizes to the ends of muscles where
they attach to these cells (reviewed in Brown 1993).
In the embryo, PS integrins are required in both
layers- muscles and epidermis- to help DER
mediated regulation of TC differentiation (Martin-
Bermudo 2000). Besides integrins, there are other
molecules like Tenascin A, Tiggrin, Laminin, Slit etc.
(Baumgartner & Chiquet-Ehrisman 1993, Fogerty
et al. 1994, Rothberg et al. 1990, Kidd et al. 1999).
Attachment of Adult Flight Muscles
During pupation, as the adult epidermis arising from
the evaginating imaginal discs gradually replaces the
larval epidermis, the larval connections are replaced
with new attachments to the adult cuticle. In the
adult, like in the embryo, these attachment sites are
specified by the gene sr (Fernandes et al. 1996). On
the presumptive notum region of the wing disc, sr is
expressed in a discrete set of domains, in the
epidermal cells. There are four regions in the
anterior notum and one thin line in the posterior
notum (Figure 4a), which prefigure attachment sites
of flight muscles. sr is the earliest marker known to
specify these domains, at the disc stage itself. During
pupation, these regions form the tendon cells of flight
muscles (figure 4b). The development of these
attachment sites with respect to the developing
muscles has been described, and correlation of
different IFM fibres to specific subsets of sr domains
has been mapped in detail (Fernandes et al. 1996). The
anterior clusters serve as insertion points for the
DLMs, the DVMs and the TDT, whereas the
posterior region is adhered to by the posterior DLM
ends and the ventral end of DVM III. Ventral
attachments of DVM I and II and TDT arise from the
leg discs (figure 4b). It is conceivable that matching of
muscle fibres and specific sr subsets occurs and sr
may act in conjunction with other molecules to ensure
that correct muscle pattern develops. Mutations at
the sr locus cause defects in IFMs, especially in the
DLMs. There is DLM loss characterised by normal
early steps of myogenesis followed by detachment of
the fibres, which leads to curling up and eventual
degeneration (Costello & Wyman 1986).
Information on the function of sr in adult muscle
attachment comes from expression pattern analysis,
partial loss of function mutants and from data from
the embryo, but the details of its action are not clear.
Figure 4 Flight Muscle Attachment Sites. a, Schematic
representation of a wing imaginal disc with the sr-
expressing attachment sites shown in red. The four
domains a-d give rise to the anterior dorsal
attachment sites of the IFMs while the thin posterior
stripe [arrow] gives rise to the posterior attachment
sites; b, Schematic of an adult heminotum that forms
from the presumptive notum region of a wing disc.
DLMs are shown as dark green fibres, indicated by
yellow asterisks. DVMs are shown in lighter green
and indicated by red asterisks. The attachment sites
of these muscles, derived from the sr- expressing cells
of the wing disc, are shown in red. Different domains
[a-d] that correlate with the attachment of different
muscle subsets are indicated in the figure. Posterior
attachment site is marked by a blue arrow. The brown
arrow marks the epidermal extensions, apodemes,
which connect the muscle to the tendon cells. In a,
anterior is to the left. In b, anterior is to the top.
699
Muscle Development in Drosophila
Besides sr, a few other molecules are known to
show remarkable expressions in adult TCs.
Integrins display a dynamic temporal expression
profile during flight muscle attachment. They are
not expressed when muscle fibres first make their
appearance (12-20h APF) but following muscle-
epidermis contact are detected at the attachment
sites. PS1 is at the muscle ends and also in the fibres
that connect the developing muscles to their
attachment sites, while PS2 is restricted to ends of
larval muscles (Fernandes et al. 1996). A few
mutants that show attachment specific defects have
been reported. These include the Broad Complex
(BR-C) transcriptions factors that are induced by
the hormone 20-hydroxyecdysone (20E).
Mutations of the reduced bristles on palpus (rbp)
complementation group, which corresponds to the
BRC-Z1, the isoform expressed in pupal tendon
cells, reduce or eliminate DVMs selectively and
disrupt muscle attachments. Mosaic analyses have
revealed that rbp+ function is required in dorsal TCs
for normal DVM attachment. Presumably BR-C Z1,
under control of 20E regulates target genes whose
products control specific features of TC maturation
(Sandstorm et al. 1997, Sandstorm & Restifo 1999).
A Type I Ser/Thr Protein Phosphatase 1(PP1) that is
encoded by the gene flapwing (flw) also functions in
maintenance of IFM attachments (Raghavan et al.
2000). Ultrastructure of adult flight muscle MTJs
has been described and resembles the embryonic
description. A recent study however, has described
the presence of an additional sheath of overlapping
flattened cytoplasmic extensions surrounding the
TC processes (Sandstorm & Restifo 1999).
While much is now known about the
mechanisms governing different aspects of muscle
development, both in the embryo, and in the adult
Drosophila, there is much more that needs to be
deciphered. This includes the genetic and molecular
aspects of muscle size control, muscle diversity etc.
The importance of signalling networks interacting
with each other in controlling myogenic targets and
transcriptional heirarchies are beginning to be
unravelled now. Greater insights into the
complexities of these interactions and feedback
networks, along with discovery of novel factors
whose functions remain to be examined will help
elucidate the details of myogenesis. The adult flight
muscle system continues to remain an exciting
model to ask such questions of wide biological
significance and will surely contribute in providing
answers to many of them.
References
Anant S, Roy S and VijayRaghavan K 1998 Twist and
Notch negatively regulate adult muscle differentiation
in Drosophila; Development 125 1361-1369
Armand P, Knapp A C, Hirsch A J, Wieschaus E F and
Cole M D (1994). A novel basic helix-loop-helix
protein is expressed in muscle attachment sites
of the Drosophila epidermis; Mol. Cell Biol. 14
4145-4154
Azpiazu N and Frasch M 1993 tinman and bagpipe:
two homeo box genes that determine cell fates in
the dorsal mesoderm of Drosophila; Genes Dev. 7
1325-1340
_____, Lawrence P A, Vincent J P and Frasch M 1996
Segmentation and specification of the Drosophila
mesoderm; Genes Dev. 10 3183-3194
Baehrecke E H 1997 who encodes a KH RNA binding
protein that functions in muscle development;
Development 124 1323-1332
Balagopalan L, Keller C and Abmayr S 2001 Loss of
function mutations reveal that the Drosophila
nautilus gene is not essential for embryonic
myogenesis; Dev. Biol. 231 374-382
Baker R and Schubiger G 1996 Autonomous and
nonautonomous Notch functions for embryonic
muscle and epidermis development in Drosophila;
Development 122 617-626
Baonza A, Roch F and Martin-Blanco E 2000 DER
signaling restricts the boundaries of the wing field
during Drosophila development; Proc. Natl. Acad.
Sci. USA 97 7331-7335
Barthmaier P and Fyrberg E 1995 Monitoring
development and pathology of Drosophila indirect
flight muscles using green fluorescent protein; Dev.
Biol. 169 770-774
Bate M 1990 The embryonic development of larval muscles
in Drosophila; Development 110 791-804
Bate M, Rushton E and Currie D A 1991 Cells with
persistent Twist expression are the embryonic
precursors of adult muscles in Drosophila;
Development 113 79-89
______ and Rushton E 1993 Myogenesis and muscle
patterning in Drosophila; C R Acad. Sci. III 316
1047-1061
_____ 1993 The mesoderm and its derivatives, in The
Development of Drosophila melanogaster. Vol. 2 eds
M Bate and A Martinez-Arias, pp.1013-1090 (New
York: CSHL press)
700 Arjumand Ghazi and K Vijayraghavan
Bate M, Rushton E and Frasch M 1993 A dual requirement
for neurogenic genes in Drosophila myogenesis; Dev.
Suppl. 1993 149-161
Baumgartner S and Chiquet-Ehrismann R 1993 Tena, a
Drosophila gene related to tenascin, shows selective
transcript localization; Mech. Dev 40 165-176
Baylies M K, Martinez Arias A and Bate M 1995 wingless
is required for the formation of a subset of muscle
founder cells during Drosophila embryogenesis;
Development 121 3829-3837
Becker S, Pasca G, Strumpf D, Min L and Volk T 1997
Reciprocal signaling between Drosophila epidermal
muscle attachment cells and their corresponding
muscles; Development 124 2615-2622
Bernstein S I, ODonnell P T and Cripps R M 1993 Molecular
genetic analysis of muscle development, structure, and
function in Drosophila; Int. Rev. Cytol. 143 63-152
Blochlinger K, Jan L Y and Jan Y N 1993 Postembryonic
patterns of expression of cut, a locus regulating
sensory organ identity in Drosophila; Development
117 441-450
Bour B A, Chakravarti M, West J M and Abmayr S M
2000 Drosophila SNS, a member of the
immunoglobulin superfamily that is essential for
myoblast fusion; Genes Dev. 14 1498-1511
Bourgouin C, Lundgren S E and Thomas J B 1992
Apterous is a Drosophila LIM domain gene required
for the development of a subset of embryonic muscles;
Neuron 9 549-561
Brower D L, Bunch T A, Mukai L, Adamson T E, Wehrli
M, Lam S, Friedlander E, Roote C E and Zusman S
1995a Nonequivalent requirements for PS1 and PS2
integrin at cell attachments in Drosophila: genetic
analysis of the alpha PS1 integrin subunit;
Development 121 1311-1320
_____, Brabant M C and Bunch T A 1995b Role of the PS
integrins in Drosophila development. Immunol; Cell
Biol. 73 558-64
Brown N H 1993 Integrins hold Drosophila together;
Bioessays 15(6)383-390 Review
______ 1994 Null mutations in the alpha PS2 and beta PS
integrin subunit genes have distinct phenotypes;
Development 120 1221-1231
Buttgereit D 1996 Transcription of the beta 1 tubulin (beta
Tub56D) gene in apodemes is strictly dependent on
muscle insertion during embryogenesis in Drosophila
melanogaster; Eur. J. Cell Biol. 71 183-191
Campbell S D, Duttaroy A, Katzen A L and Chovnick A
1991 Cloning and characterization of the scalloped region
of Drosophila melanogaster; Genetics 127 367-380
Carmena A, Bate M and Jimenez F 1995 lethal of scute, a
proneural gene, participates in the specification of
muscle progenitors during Drosophila embryogenesis;
Genes Dev. 9 2373-2383
Carmena A, Gisselbrecht S, Harrison J, Jimenez F and
Michelson A M 1998 Combinatorial signaling codes
for the progressive determination of cell fates in
the Drosophila embryonic mesoderm; Genes Dev.
12 3910-3922
Caveney S 1969 Muscle attachment related to cuticle
architecture in Apterygota; J. Cell Sci. 4 541-559
Corbin V, Michelson A M, Abmayr S M, Neel V, Alcamo
E, Maniatis T and Young M W 1991 A role for the
Drosophila neurogenic genes in mesoderm
differentiation; Cell 67 311-323
Costello W J and Wyman R J 1986 Development of an
indirect flight muscle in a muscle-specific mutant of
Drosophila melanogaster; Dev. Biol. 118 247-258
Couso J P, Knust E and Martinez Arias A 1995 Serrate
and wingless cooperate to induce vestigial gene
expression and wing formation in Drosophila; Curr.
Biol. 5 1437-1448
Crossley A C 1978 The morphology and development of
the Drosophila muscular system; in Genetics and
Biology of Drosophila. Vol. 2b eds M Ashburner and
T R F Wright pp. 499-560 (New York: Acad. press)
Doberstein S K, Fetter R D, Mehta A Y and Goodman C
S 1997 Genetic analysis of myoblast fusion: blown
fuse is required for progression beyond the prefusion
complex; J. Cell Biol. 136 1249-1261
Dohrmann C, Azpiazu N and Frasch M 1990 A new
Drosophila homeo box gene is expressed in
mesodermal precursor cells of distinct muscles during
embryogenesis; Genes Dev. 4 2098-2111
Emori Y and Saigo K 1993 Distinct expression of two
Drosophila homologs of fibroblast growth factor
receptors in imaginal discs; FEBS Lett. 332 111-114
Erickson M R, Galletta B J and Abmayr S M 1997 Drosophila
myoblast city encodes a conserved protein that is
essential for myoblast fusion, dorsal closure, and
cytoskeletal organization; J. Cell Biol. 138 589-603
Fernandes J J, Bate M and Vijayraghavan K 1991
Development of the indirect flight muscles of
Drosophila; Development 113 67-77
_____ and VijayRaghavan K 1993 Development of the
indirect flight muscle innervation in Drosophila
melanogaster; Development 118 215-227
_____ and Keshishian H 1996 Patterning the dorsal
longitudinal flight muscles (DLM) of Drosophila:
insights from the ablation of larval scaffolds;
Development 122 3755-3763
______ and _____ 1998 Nerve-muscle interactions during
flight muscle development in Drosophila;
Development 125 1769-1779
_____, Celniker S E and VijayRaghavan K 1996
Development of the indirect flight muscle attachment
sites in Drosophila: role of the PS integrins and the
stripe gene; Dev. Biol. 176 166-184
701
Muscle Development in Drosophila
Fogerty F J, Fessler L I, Bunch T A, Yaron Y, Parker C
G, Nelson R E, Brower D L, Gullberg D and Fessler
J H 1994 Tiggrin, a novel Drosophila extracellular
matrix protein that functions as a ligand for
Drosophila alpha PS2 beta PS integrins; Development
120 1747-1758
Frasch M 1995 Induction of visceral and cardiac mesoderm
by ectodermal Dpp in the early Drosophila embryo;
Nature 374 464-467
_____1999 Controls in patterning and diversification
of somatic muscles during Drosophila
embryogenesis; Curr. Opin. Genet. Dev. 9
522-529
Frise E, Knoblich J A, Younger-Shepherd S, Jan L Y and
Jan Y N 1996 The Drosophila Numb protein inhibits
signaling of the Notch receptor during cell-cell
interaction in sensory organ lineage; Proc. Natl. Acad.
Sci. U S A 93 11925-11932
Frommer G, Vorbruggen G, Pasca G, Jackle H and Volk T
1996 Epidermal egr-like zinc finger protein of
Drosophila participates in myotube guidance; Embo.
J. 15 1642-1649
Ghazi A and VijayRaghavan K 2000 Control by
combinatorial codes; Nature 408(6811) 419-420
_____, Anant S and VijayRaghavan K 2000 Apterous
mediates development of direct flight muscles
autonomously and indirect flight muscles through
epidermal cues; Development 127 5309-5318
Goubeaud A, Knirr S, Renkawitz-Pohl R and Paululat
A 1996 The Drosophila gene alien is expressed in
the muscle attachment sites during embryogenesis
and encodes a protein highly conserved between
plants, Drosophila and vertebrates; Mech. Dev. 57
59-68
Gregory S L and Brown N H 1998 kakapo, a gene required
for adhesion between and within cell layers in
Drosophila, encodes a large cytoskeletal linker protein
related to plectin and dystrophin; J. Cell Biol. 143
1271-1282
Halfon M S, Carmena A, Gisselbrecht S, Sackerson C
M, Jimenez F, Baylies M K and Michelson A M
2000 Ras pathway specificity is determined by
the integration of multiple signal-activated and
tissue-restricted transcription factors; Cell 103
63-74
Jagla T, Bellard F, Lutz Y, Dretzen G, Bellard M and Jagla
K 1998 ladybird determines cell fate decisions during
diversification of Drosophila somatic muscles;
Development 125 3699-3708
Kidd T, Bland K S and Goodman C S 1999 Slit is the
midline repellent for the robo receptor in Drosophila;
Cell 96 785-794
Lai E C, Bodner R and Posakony J W 2000 The enhancer
of split complex of Drosophila includes four Notch-
regulated members of the bearded gene family;
Development 127 3441-3455
Landgraf M, Baylies M and Bate M 1999 Muscle founder
cells regulate defasciculation and targeting of motor
axons in the Drosophila embryo; Curr. Biol. 9 589-592
Lee J C, VijayRaghavan K, Celniker S E and Tanouye M A
1995 Identification of a Drosophila muscle development
gene with structural homology to mammalian early
growth response transcription factors; Proc. Natl.
Acad. Sci. U S A 92 10344-10348
Leptin M and Grunewald B 1990 Cell shape changes during
gastrulation in Drosophila; Development 110(1) 73-84
Luo L, Liao Y J, Jan L Y and Jan Y N 1994 Distinct
morphogenetic functions of similar small GTPases:
Drosophila Drac1 is involved in axonal outgrowth
and myoblast fusion; Genes Dev. 8 1787-1802
Martin-Bermudo M D 2000 Integrins modulate the Egfr
signaling pathway to regulate tendon cell
differentiation in the Drosophila embryo; Development
127 2607-2615
Nabel-Rosen H, Dorevitch N, Reuveny A and Volk T 1999
The balance between two isoforms of the Drosophila
RNA-binding protein How controls tendon cell
differentiation; Mol. Cell 4 573-584
Newman S M and Wright T R 1981 A histological and
ultrastructural analysis of developmental defects
produced by the mutation, lethal(1)myospheroid, in
Drosophila melanogaster; Dev. Biol. 86 393-402
Ng M, Diaz-Benjumea F J, Vincent J P, Wu J and Cohen
S M 1996 Specification of the wing by localized
expression of wingless protein; Nature 381 316-318
Piepenburg O, Vorbruggen G and Jackle H 2000 Drosophila
segment borders result from unilateral repression of
Hedgehog activity by Wingless signalling; Mol. Cell 6
203-209
Prokop A, Martin-Bermudo M D, Bate M and Brown N H
1998 Absence of PS integrins or laminin A affects
extracellular adhesion, but not intracellular assembly,
of hemiadherens and neuromuscular junctions in
Drosophila embryos; Dev. Biol. 196 58-76
Raghavan S, Williams I, Aslam H, Thomas D, Szoor B,
Morgan G, Gross S, Turner J, Fernandes J,
VijayRaghavan K and Alphey L 2000 Protein
phosphatase 1beta is required for the maintenance of
muscle attachments; Curr. Biol. 10 269-272
Ranganayakulu G, Zhao B, Dokidis A, Molkentin J D,
Olson E N and Schulz R A 1995 A series of mutations
in the D-MEF2 transcription factor reveal multiple
functions in larval and adult myogenesis in
Drosophila; Dev. Biol. 171 169-81
Riechmann V, Irion U, Wilson R, Grosskortenhaus R
and Leptin M 1997 Control of cell fates and
segmentation in the Drosophila mesoderm;
Development 124 2915-2922
Rivlin P K, Schneiderman A M and Booker R 2000 Imaginal
pioneers prefigure the formation of adult thoracic
muscles in Drosophila melanogaster; Dev. Biol. 222
450-459
702 Arjumand Ghazi and K Vijayraghavan
Rothberg J M, Jacobs J R, Goodman C S and Artavanis-
Tsakonas S 1990 Slit: an extracellular protein necessary
for development of midline glia and commissural
axon pathways contains both EGF and LRR domains;
Genes Dev. 4 2169-2187
Roy S, Shashidhara L S and VijayRaghavan K 1997 Muscles
in the Drosophila second thoracic segment are
patterned independently of autonomous homeotic
gene function; Curr. Biol. 7 222-227
______ and ______ 1997 Homeotic genes and the regulation
of myoblast migration, fusion, and fibre-specific gene
expression during adult myogenesis in Drosophila;
Development 124 3333-3341
______ and ______ 1998 Patterning muscles using organizers:
larval muscle templates and adult myoblasts actively
interact to pattern the dorsal longitudinal flight muscles
of Drosophila; J. Cell Biol. 141 1135-1145
______ and ______ 1999 Muscle pattern diversification in
Drosophila: the story of imaginal myogenesis;
Bioessays 21 486-498
Ruiz-Gomez M and Bate M 1997 Segregation of myogenic
lineages in Drosophila requires numb; Development
124 4857-4866
______, Romani S, Hartmann C, Jackle H and Bate M 1997
Specific muscle identities are regulated by Kruppel
during Drosophila embryogenesis; Development 124
3407-3414
______, Coutts N, Price A, Taylor M V and Bate M 2000
Drosophila dumbfounded: a myoblast attractant
essential for fusion; Cell 102 189-198
Rushton E., Drysdale R, Abmayr S. M, Michelson A M,
and Bate M 1995 Mutations in a novel gene, myoblast
city, provide evidence in support of the founder cell
hypothesis for Drosophila muscle development;
Development 121 1979-1988
Sandstrom D J, Bayer C A, Fristrom J W and Restifo L L
1997 Broad-complex transcription factors regulate
thoracic muscle attachment in Drosophila; Dev. Biol.
181 168-185
______ and Restifo L L 1999 Epidermal tendon cells require
Broad Complex function for correct attachment of
the indirect flight muscles in Drosophila melanogaster;
J. Cell Sci. 112 4051-4065
Schnepp B, Grumbling G, Donaldson T and Simcox A 1996
Vein is a novel component in the Drosophila epidermal
growth factor receptor pathway with similarity to the
neuregulins; Genes Dev. 10 2302-2313
Shatoury H H E 1956 Developmental interactions in the
development of the imaginal muscles of Drosophila;
J. Embryol. Exp. Morph. 4 228-239
Staehling-Hampton K, Hoffmann F M, Baylies M K,
Rushton E and Bate M 1994 dpp induces
mesodermal gene expression in Drosophila; Nature
372 783-786
Strumpf D and Volk T 1998 Kakapo, a novel cytoskeletal-
associated protein is essential for the restricted
localization of the neuregulin-like factor, vein, at the
muscle-tendon junction site; J. Cell Biol. 143 1259-1270
Tepass U and Hartenstein V 1994 The development of
cellular junctions in the Drosophila embryo; Dev. Biol.
161 563-596
Thisse B, Stoetzel C, Gorostiza-Thisse C and Perrin-Schmitt
F 1988 Sequence of the twist gene and nuclear
localization of its protein in endomesodermal cells of
early Drosophila embryos; EMBO J. 7 2175-2183
Volk T and VijayRaghavan K 1994 A central role for
epidermal segment border cells in the induction of
muscle patterning in the Drosophila embryo;
Development 120 59-70
Vorbruggen G and Jackle H 1997 Epidermal muscle
attachment site specific target gene expression and
interference with myotube guidance in response to
ectopic stripe expression in the developing Drosophila
epidermis; Proc. Natl. Acad. Sci. USA 94 8606-8611
Wang S H, Simcox A and Campbell G 2000 Dual role
for Drosophila epidermal growth factor receptor
signaling in early wing disc development; Genes
Dev. 14 2271-2276
Williams G J and Caveney S 1980a A gradient of
morphogenetic information involved in muscle
patterning; J. Embryol. Exp. Morphol. 58 35-61
______ and Caveney S 1980b Changing muscle patterns
in a segmental epidermal field; J. Embryol. Exp.
Morphol. 58 13-33
Williams J, Littlefield C L and Nothiger R 1984 Regulative
interactions between cells of wing discs from different
dipteran species; Dev. Biol. 105 227-233
Williams J A, Paddock S W and Carroll S B 1993 Pattern
formation in a secondary field: a hierarchy of
regulatory genes subdivides the developing
Drosophila wing disc into discrete subregions;
Development 117 571-584
______, Paddock S W, Vorwerk K and Carroll S B 1994
Organization of wing formation and induction of
a wing-patterning gene at the dorsal/ventral
compartment boundary; Nature 368(6469) 299-305
Yarnitzky T, Min L and Volk T 1997 The Drosophila
neuregulin homolog Vein mediates inductive
interactions between myotubes and their epidermal
attachment cells; Genes Dev. 11 2691-2700
Yin Z, Xu X L and Frasch M 1997 Regulation of the twist
target gene tinman by modular cis-regulatory elements
during early mesoderm development; Development
124 4971-4982
Zaffran S, Astier M, Gratecos D and Semeriva M 1997 The
held out wings (how) Drosophila gene encodes a
putative RNA-binding protein involved in the control
of muscular and cardiac activity; Development 124
2087-2098
... Secreted signalling molecules from ectoderm are instrumental in directing the mesodermal cells to adopt restrictive fates as cardiac, visceral and somatic muscle lineages. This topic has been reviewed extensively [41,47]. Mesodermal cells are exposed to external signalling factors, wingless (Wg), Decapentaplegic (Dpp) or Hedgehog (Hh) and become compartmentalized along antero-posterior (A-P) and dorsoventral (D-V) axis in each hemisegment of an embryo [48]. ...
... Remaining mesodermal cells with low levels of l'sc become fusion competent myoblasts (FCMs), earlier known as feeder cells [49]. Muscle progenitors divide asymmetrically via differential localization of Numb (Nb) and Inscutable (insc) proteins to form either two FC or an adult muscle precursor (AMP) cell and a FC [47,50]. Thirty FCs are seeded in each abdominal hemisegment, with each specifying an individual muscle fibre. ...
... Studies related to the AMPs that are present in the gaps between the larval muscles and peripheral nerves associated with them [47,75] have shed light on their niche that maintains them in a quiescent state [74]. Aradhya et al. [76] have demonstrated a niche of these quiescent AMPs during embryonic stages maintains filopodia-based contact with larval muscles, and this contact is needed for their future reactivation through initiation of insulin pathway and activation of N in AMPs. ...
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Movement assisted by muscles forms the basis of various behavioural traits seen in Drosophila. Myogenesis involves developmental processes like cellular specification, differentiation, migration, fusion, adherence to tendons and neuronal innervation in a series of coordinated event well defined in body space and time. Gene regulatory networks are switched on–off, fine tuning at the right developmental stage to assist each cellular event. Drosophila is a holometabolous organism that undergoes myogenesis waves at two developmental stages, and is ideal for comparative analysis of the role of genes and genetic pathways conserved across phyla. In this review we have summarized myogenic events from the embryo to adult focussing on the somatic muscle development during the early embryonic stage and then on indirect flight muscles (IFM) formation required for adult life, emphasizing on recent trends of analysing muscle mutants and advances in Drosophila muscle biology.
... Thus, in the thorax, the largest muscles of the adult Drosophila, the DLM indirect flight muscles, are generated through the use of one set of larval muscles as a persistent scaffold-like template that is transformed into functional adult muscle by numerous swarming and fusing myoblasts derived from the high Twist AMPs. In the abdomen, the interstices of larval muscles and peripheral nerves associated with larval muscles appear to act as position-specific niches for AMPs until adult abdominal muscle development is started during the pupal stage [6,[27][28][29]. ...
... The genes involved in the various stages of the fusion are mentioned in black letters. Similarly, the distances between fusing myoblast membrane and myotube membrane from electron microscopic studies are mentioned in black letters [27]. not involve Notch mediated lateral inhibition. ...
... The epidermal cells that serve as attachment sites for the DLMs are located in the notal region of the wing disc epithelium directly adjacent to the site of adult-specific myoblast proliferation. These prospective muscle attachment cells and their location in the disc epidermis are specified by expression of the stripe gene [27,41]. (The actual physical attachment between the epidermal tendon cell and muscle cells at the myotendinous junction is mediated by interactions between shared and cell type-specific integrin adhesion molecules during pupal stages.) ...
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Myogenesis is a highly orchestrated, complex developmental process by which cell lineages that are mesodermal in origin generate differentiated multinucleate muscle cells as a final product. Considerable insight into the process of myogenesis has been obtained for the embryonic development of the larval muscles of Drosophila. More recently, the postembryonic development of the muscles of the adult fly has become a focus of experimental investigation of myogenesis since specific flight muscles of the fly manifest remarkable similarities to vertebrate muscles in their development and organization. In this review, we catalog some of the milestones in the study of myogenesis in the large adult-specific flight muscles of Drosophila. The identification of mesoderm-derived muscle stem cell lineages, the characterization of the symmetric and asymmetric divisions through which they produce adult-specific myoblasts, the multifaceted processes of myoblast fusion, and the unexpected discovery of quiescent satellite cells that can be activated by injury are discussed. Moreover, the finding that all of these processes incorporate a plethora of signaling interactions with other myogenic cells and with niche-like neighboring tissue is considered. Finally, we briefly point out possible future developments in the area of Drosophila myogenesis that may lead to of new avenues of genetic research into the roles of muscle stem cells in development, disease and aging.
... During the embryonic development a single founder cell is determined for each fiber of a prospective flight muscle (Dutta et al., 2004). These cells later recruit other myoblasts, the feeder cells, which merge and thus form the definite fiber (Ghazi and Vijayraghavan, 2003). The evagination of the imaginal disc takes place during the metamorphosis. ...
... This suggests that connections between fibers can be formed, but does not explain why fibers with an unmodified course can be partly fused, with partially or completely unrecognizable borders. However, wingless is also involved in the cell determination in the mesoderm and regulates whether myoblasts develop as somatic or visceral muscles, together with other factors (Ghazi and Vijayraghavan, 2003;Carmena and Baylies, 2006). It is conceivable that this can result in modified fusion processes of myoblasts differing from the normal pattern of myogenesis. ...
Article
Genetically modified organisms are crucial for our understanding of gene regulatory networks, physiological processes and ontogeny. With modern molecular genetic techniques allowing the rapid generation of different Drosophila melanogaster mutants, efficient in-depth morphological investigations become an important issue. Anatomical studies can elucidate the role of certain genes in developmental processes and point out which parts of gene regulatory networks are involved in evolutionary changes of morphological structures. The wingless mutation wg(1) of Drosophila melanogaster was discovered more than forty years ago. While early studies addressed the external phenotype of these mutants, the documentation of the internal organization was largely restricted to the prominent indirect flight muscles. We used SEM micrographs, histological serial sections, μ-computed tomography, CLSM and 3D reconstructions to study and document the thoracic skeletomuscular system of the wild type and mutant. A recently introduced nomenclature for the musculature of neopteran insects was applied to facilitate comparisons with closely or more distantly related taxa. The mutation is phenotypically mainly characterized by the absence of one or both wings and halteres. The wing is partly or entirely replaced by duplications of mesonotal structures, whereas the haltere and its associated muscles are completely absent on body sides showing the reduction. Both the direct and indirect mesothoracic flight muscles are affected by loss and reorientation of bundles or fibers. Our observations lead to the conclusion that the wingless mutation causes a homeotic transformation in the imaginal discs of wings and halteres with a direct effect on the development of skeletal structures and an indirect effect on the associated muscular system.
... Consistent with our results, such abdominal responses are activated by full-field rotation and to a higher (lesser) degree in response to FTB (BTF) motion (Götz et al., 1979). These movements result from the coordinated activation of associated muscles by descending inputs to T3 and the abdominal ganglion (Milde and Strausfeld, 1990; Hartenstein, 2006) and as similar to our observations for head yaw (Figure 5B), are largely correlated with movements of the wings (Götz et al., 1979; Zanker et al., 1991). ...
... In addition, an array of homo and heterolateral descending neurons (DNs) with dendrites in deutocerebral, as well as proto and tritocerebral clusters, supply the motor systems of the thoracic ganglion and presumably trigger robust compensatory steering behaviors (Strausfeld and Bassemir, 1985a,b; Strausfeld and Gronenberg, 1990). Although it is not known whether the muscles controlling head movements in Drosophila are similar among all flies (Hartenstein, 2006), in larger flies, oblique horizontal muscles spanning the cervical connective are activated by the neck motor neurons (NMNs) of the cervical nerve arising in the subesophageal ganglion (SOG) and are potentially involved in yaw movements of the head (Strausfeld et al., 1987). Half-field FTB motion generates hemispheric differences in the depolarization or hyperpolarization of neurons residing in the left and right lobula plate (Figure 6, red vs. blue), providing sufficient signaling to modulate the stroke amplitudes of the left and right wings and yaw movements of the head (Figures 3G,H). ...
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In response to imposed course deviations, the optomotor reactions of animals reduce motion blur and facilitate the maintenance of stable body posture. In flies, many anatomical and electrophysiological studies suggest that disparate motion cues stimulating the left and right eyes are not processed in isolation but rather are integrated in the brain to produce a cohesive panoramic percept. To investigate the strength of such inter-ocular interactions and their role in compensatory sensory-motor transformations, we utilize a virtual reality flight simulator to record wing and head optomotor reactions by tethered flying flies in response to imposed binocular rotation and monocular front-to-back and back-to-front motion. Within a narrow range of stimulus parameters that generates large contrast insensitive optomotor responses to binocular rotation, we find that responses to monocular front-to-back motion are larger than those to panoramic rotation, but are contrast sensitive. Conversely, responses to monocular back-to-front motion are slower than those to rotation and peak at the lowest tested contrast. Together our results suggest that optomotor responses to binocular rotation result from the influence of non-additive contralateral inhibitory as well as excitatory circuit interactions that serve to confer contrast insensitivity to flight behaviors influenced by rotatory optic flow.
... Adult muscle progenitors [15] fuse to three remnant syncytia of larval body wall muscles on either side of the midline, called templates. By 20 h apf (after puparium formation), each template splits into two syncytia, giving rise to six fibres in each adult hemithorax [16]. In figure 2c-e, we show three animals where templates failed to split during pupariation. ...
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Indirect flight muscles (IFMs) in adult Drosophila provide the key power stroke for wing beating. They also serve as a valuable model for studying muscle development. An age-dependent decline in Drosophila free flight has been documented, but its relation to gross muscle structure has not yet been explored satisfactorily. Such analyses are impeded by conventional histological preparations and imaging techniques that limit exact morphometry of flight muscles. In this study, we employ microCT scanning on a tissue preparation that retains muscle morphology under homeostatic conditions. Focusing on a subset of IFMs called the dorsal longitudinal muscles (DLMs), we find that DLM volumes increase with age, partially due to the increased separation between myofibrillar fascicles, in a sex-dependent manner. We have uncovered and quantified asymmetry in the size of these muscles on either side of the longitudinal midline. Measurements of this resolution and scale make substantive studies that test the connection between form and function possible. We also demonstrate the application of this method to other insect species making it a valuable tool for histological analysis of insect biodiversity.
... In Drosophila melanogaster, the organization and metabolism of skeletal muscle fibers is similar to that in mammals (Taylor, 2006; Piccirillo et al., 2013). However, the muscles undergo more dramatic age-related deterioration, presumably due to the lack of satellite stem cells and limited capacity for muscle repair in this organism. ...
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A characteristic feature of aged humans and other mammals is the debilitating, progressive loss of skeletal muscle function and mass that is known as sarcopenia. Age-related muscle dysfunction occurs to an even greater extent during the relatively short lifespan of the fruit fly Drosophila melanogaster. Studies in model organisms indicate that sarcopenia is driven by a combination of muscle tissue extrinsic and intrinsic factors, and that it fundamentally differs from the rapid atrophy of muscles observed following disuse and fasting. Extrinsic changes in innervation, stem cell function and endocrine regulation of muscle homeostasis contribute to muscle aging. In addition, organelle dysfunction and compromised protein homeostasis are among the primary intrinsic causes. Some of these age-related changes can in turn contribute to the induction of compensatory stress responses that have a protective role during muscle aging. In this Review, we outline how studies in Drosophila and mammalian model organisms can each provide distinct advantages to facilitate the understanding of this complex multifactorial condition and how they can be used to identify suitable therapies.
... Unlike the simple bipolar dendrite in the larva, the adult form of the cell has a densely fasciculated dendritic bundle that fills the space between two of the dorsal longitudinal muscle fibers that extend from midsegment to the posterior segment border (Figure 1C-C''). These dorsal abdominal muscles are retractors of the abdominal tergites [25], and a single dbd serves as an internal stretch receptor in the set of 15 to 25 muscles per hemi-segment. The larval muscles, with which dbd makes contact, degenerate early in metamorphosis and are replaced by adult-specific muscles. ...
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Various members of the family of BTB/POZ zinc-finger transcription factors influence patterns of dendritic branching. One such member, Broad, is notable because its BrZ3 isoform is widely expressed in Drosophila in immature neurons around the time of arbor outgrowth. We used the metamorphic remodeling of an identified sensory neuron, the dorsal bipolar dendrite sensory neuron (dbd), to examine the effects of BrZ3 expression on the extent and pattern of dendrite growth during metamorphosis. Using live imaging of dbd in Drosophila pupae, we followed its normal development during metamorphosis and the effect of ectopic expression of BrZ3 on this development. After migration of its cell body, dbd extends a growth-cone that grows between two muscle bands followed by branching and turning back on itself to form a compact dendritic bundle. The ectopic expression of the BrZ3 isoform, using the GAL4/UAS system, caused dbd's dendritic tree to transform from its normal, compact, fasciculated form into a comb-like arbor that spread over on the body wall. Time-lapse analysis revealed that the expression of BrZ3 caused the premature extension of the primary dendrite onto immature myoblasts, ectopic growth past the muscle target region, and subsequent elaboration onto the epidermis. To control the timing of expression of BrZ3, we used a temperature-sensitive GAL80 mutant. When BrZ3 expression was delayed until after the extension of the primary dendrite, then a normal arbor was formed. By contrast, when BrZ3 expression was confined to only the early outgrowth phase, then ectopic arbors were subsequently formed and maintained on the epidermis despite the subsequent absence of BrZ3. The adult arbor of dbd is a highly branched arbor whose branches self-fasciculate to form a compact dendritic bundle. The ectopic expression of BrZ3 in this cell causes a premature extension of its growth-cone, resulting in dendrites that extend beyond their normal muscle substrate and onto the epidermis, where they form a comb-shaped, ectopic arbor. Our quantitative data suggest that new ectopic arbor represents an 'unpacking' of the normally fasciculated arbor onto the epidermis. These data suggest that the nature of their local environment can change dendrite behavior from self-adhesion to self-avoidance.
... D. melanogaster adults have muscles of all three kinds that are divided into somatic gonad muscles, visceral muscles, cardiac muscles and somatic muscles. In contrast, C. elegans hermaphrodites' muscles belong to two categories: striated somatic and nonstriated muscles (Altun & Hall, 2008b;Hartenstein, 2006;Taylor, 1995). ...
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Protein quality control exerts an essential role in the development and maintenance of living cells. An alteration in the balance of protein homeostasis can lead to the manifestation of different pathologies. Components of a certain class of protein ubiquitin ligases (E3/E4) have been recently identified as pivotal proteins linking the proteasome degradation pathway to protein quality control. In particular, the highly conserved E3/E4 ligases CHN-1/CHIP was shown to be part of a multiprotein complex in muscles, interacting with molecular chaperons and other E3/E4 ubiquitin ligases. This protein complex is required for proper myosin assembly and is therefore crucial for myogenesis. In this study, the Duchenne Muscular Dystrophy (DMD) model of C. elegans was further characterised and a relevant contribution of CHN-1/CHIP in the manifestation of DMD was revealed. Detailed analysis of the C. elegans DMD model evidenced necrotic degradation of muscle cells and a shorter lifespan compared to wild type. These features recapitulate the mammalian DMD model and the human muscular pathology. The data presented in this study show that chn-1 mutation specifically suppresses the age related muscle degeneration of DMD worms. Blockage of the proteasome with the chemical compound MG132 phenocopied the motility amelioration triggered by the chn-1 mutation. This proves that the proteasome exerts a major role in the progression of muscle loss. Analysis of transgenic lines overexpressing chn-1 exclusively in muscles or in neurons provided a direct link between CHN-1 muscular activity and muscle wasting. Moreover, the results show that a role in muscle degeneration of the CHN-1/CHIP interacting partner PDR-1/Parkin cannot be excluded. To identify new genetic factors involved in muscle wasting, two independent RNAi screenings with selected candidate genes were performed. The RNAi experiments identified the two genes F33H1.4 and vps-11 as possible suppressors of muscle degeneration. Detailed RNAi feeding experiments with F33H1.4 confirmed its suppression activity and identified this gene as a bona fide suppressor of muscle wasting in the C. elegans DMD model. Additionally, an interaction screening based on the split ubiquitin technology (a modified method of the yeast two hybrid system) was performed to find new partners and substrates of CHN-1/CHIP. Further biochemical characterisation of promising candidates led to the identification of the proteasomal component C56G2.7/Rpn13 as interactor partner of CHN-1/CHIP. Altogether, these data disclose a critical role for CHN-1/CHIP in the control of muscle wasting and degeneration, designating CHN-1/CHIP as a potential drug target for the treatment of DMD. In addition, the study provides new candidate genes possibly involved in the process of muscle wasting and expands the CHN-1/CHIP interactome.
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Mutations in the LMNA gene cause a collection of diseases known as laminopathies, including muscular dystrophies, lipodystrophies, and early-onset aging syndromes. The LMNA gene encodes A-type lamins, lamins A/C, intermediate filaments that form a meshwork underlying the inner nuclear membrane. Lamins have a conserved domain structure consisting of a head, coiled-coil rod, and C-terminal tail domain possessing an Ig-like fold. This study identified differences between two mutant lamins that cause distinct clinical diseases. One of the LMNA mutations encodes lamin A/C p.R527P and the other codes lamin A/C p.R482W, which are typically associated with muscular dystrophy and lipodystrophy, respectively. To determine how these mutations differentially affect muscle, we generated the equivalent mutations in the Drosophila Lamin C (LamC) gene, an orthologue of human LMNA. The muscle-specific expression of the R527P equivalent showed cytoplasmic aggregation of LamC, a reduced larval muscle size, decreased larval motility, and cardiac defects resulting in a reduced adult lifespan. By contrast, the muscle-specific expression of the R482W equivalent caused an abnormal nuclear shape without a change in larval muscle size, larval motility, and adult lifespan compared to controls. Collectively, these studies identified fundamental differences in the properties of mutant lamins that cause clinically distinct phenotypes, providing insights into disease mechanisms.
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Rab11, an evolutionary conserved, ubiquitously expressed subfamily of small monomeric GTPase has been known to regulate diverse cellular and developmental events, by regulating the exocytotic and transcytotic events inside the cell. Our studies show that Rab11 regulates Drosophila adult myogenesis by controlling proliferation and differentiation of the Adult muscle precursors (AMPs). Blocking Rab11 in the AMPs, which fuse to form the Indirect Flight Muscles (IFMs) of fly, renders flies completely flightless and non-viable. The indirect flight musculature, comprising of the differentially patterned dorsal longitudinal muscles (DLMs) and dorsal ventral muscles (DVMs), is affected to different extents. Abrogating or knocking down normal Rab11 function results in severely disrupted IFMs. DLMs forming from larval templates are reduced in number along with a significant reduction in their fibre size. The de novo developing DVMs are frequently absent. The DLMs in Rab11 hypomorphs are highly reduced, showing as a small constricted mass in one half of the thorax. Further, Rab11 function is essential for growth of these muscles during later half of adult myogenesis, as down regulation of Rab11 in IFMs results in degenerated muscles and broken fibres. Finally, we show that loss of Rab11 activity in the AMPs result in acquisition of migratory characteristic of myoblast as they show cellular protrusion at their polar ends accompanied with loss of cell-cell contacts. Our data provide the first evidence of a trafficking protein playing an indispensable role in regulating early stages of adult muscle development.
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The mechanism by which patterns are produced appears to be repeated in each segment of an animal, and it has been proposed that it may even have been conserved in evolution so that different species would have the same system of positional information. This idea has been tested by mixing cells of a defined fragment of the wing disc of Drosophila melanogaster with wing disc fragments of five other dipteran species to assay the ability of these disc fragments to stimulate intercalary regeneration of the D. melanogaster cells. The genetically marked (y; mwh) D. melanogaster fragment was mechanically mixed with wing discs or wing disc fragments of four drosophilids (D. melanogaster as a control, D. virilis, D. hydei, Zaprionus vittiger), of Musca domestica, and of Piophila casei. The mixed aggregates were cultured in vivo for 7 days, then metamorphosed in D. melanogaster larval hosts. The D. melanogaster fragments were only stimulated to regenerate when combined with complementary fragments from D. melanogaster or D. virilis wing discs. In the combination between D. melanogaster and D. hydei, the tissue formed integrated mosaic patterns, but no regeneration ensued. The one positive result (D. melanogaster mixed with D. virilis) shows that positional cues can be exchanged and correctly interpreted between cells of different species. The negative results do not prove that the mechanism for establishing patterns is different in the tested species, but may be due to incompatibilities that are not related to pattern formation.
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The results of grafts performed in the larva of the beetle Tenebrio molitor reveal that the position at which certain adult muscle attachments form is specified by a regulative, rather than a mosaic, mechanism. The results of grafts involving the rotation of squares of integument through 180 degrees or 90 degrees, or the antero-posterior transposition of two adjacent rectangles of integument, show that the site of muscle attachments in the antero-posterior axis of an abdominal sternite is specified by an epidermal segmental gradient of positional information. This gradient is presumed to be identical to the gradient which specifies cuticular patterns in this insect. There is a good correlation between the effect of grafts on adult muscle morphology and cuticular patterns.
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The spatial rearrangements that take place during metamorphosis in the abdominal sternites and associated retractor muscles of the beetle Tenebrio molitor are described. This paper provides the descriptive background needed to consider whether a morphogenetic gradient is involved in specifying the position at which adult muscle attachments develop. (Experimental work in support of this gradient hypothesis is published in a companion paper.) The ventral abdominal retractor muscles are extensively remodelled at metamorphosis so that the adult muscles differ considerably in appearance from the larval muscles from which they are derived. In particular, there is a change in both the absolute and relative positions of the sites of muscle attachment. Rearrangement of muscles takes place during both the prepupal period and the pupal stage. It is achieved by means of two separate and temporally distinct mechanisms. Epidermal remodelling in the pre-pupal period results in the movement of attached retractor muscles (epidermokinetic muscle movement). In the pupal stage, however, the muscles move over the basal surface of the epidermis (myokinetic muscle movement). Myokinetic movements may be brought about by extension of myoblast processes from the metamorphosing muscles. These findings are considered in terms of Poyarkoff's theory that the pupa serves as an integumental mould, approximating the shape of the adult, within which certain adult muscles develop.