Paternal dominance of trans-eQTL influences gene expression patterns in maize hybrids.
ABSTRACT Heterosis refers to the superior performance of hybrid progeny relative to their inbred parents, but the mechanisms responsible are unknown. Hybrids between the maize inbred lines B73 and Mo17 exhibit heterosis regardless of cross direction. These reciprocal hybrids differ from each other phenotypically, and 30 to 50% of their genes are differentially expressed. We identified approximately 4000 expression quantitative trait loci (eQTL) that allowed us to identify markers linked to variation in expression. We found that over three-quarters of these eQTL act in trans (78%) and that 86% of these differentially regulate transcript accumulation in a manner consistent with gene expression in the hybrid being regulated exclusively by the paternally transmitted allele. This result suggests that widespread imprinting contributes to the regulation of gene expression in maize hybrids.
- SourceAvailable from: Nancy Terrier[Show abstract] [Hide abstract]
ABSTRACT: Flavonoids are secondary metabolites with multiple functions. In grape (Vitis vinifera), the most abundant flavonoids are proanthocyanidins (PAs), major quality determinants for fruit and wine. However, knowledge about the regulation of PA composition is sparse. Thus, we aimed to identify novel genomic regions involved in this mechanism. Expression quantitative trait locus (eQTL) mapping was performed on the transcript abun-dance of five downstream PA synthesis genes (dihydroflavonol reductase (VvDFR), leuco-anthocyanidin dioxygenase (VvLDOX), leucoanthocyanidin reductase (VvLAR1), VvLAR2 and anthocyanidin reductase (VvANR)) measured by real-time quantitative PCR on a pseudo F1 population in two growing seasons. Twenty-one eQTLs were identified; 17 of them did not overlap with known candidate tran-scription factors or cis-regulatory sequences. These novel loci and the presence of digenic epistasis support the previous hypothesis of a polygenic regulatory mechanism for PA biosyn-thesis. In a genomic region co-locating eQTLs for VvDFR, VvLDOX and VvLAR1, gene annotation and a transcriptomic survey suggested that VvMYBC2-L1, a gene coding for an R2R3-MYB protein, is involved in regulating PA synthesis. Phylogenetic analysis showed its high similarity to characterized negative MYB factors. Its spatiotemporal expression profile in grape coin-cided with PA synthesis. Its functional characterization via overexpression in grapevine hairy roots demonstrated its ability to reduce the amount of PA and to down-regulate expression of PA genes.New Phytologist 02/2014; 201(3). · 6.74 Impact Factor
- [Show abstract] [Hide abstract]
ABSTRACT: Sprague and Tatum (1942) introduced the concepts of general combining ability (GCA) and specific combining ability (SCA) to evaluate the breeding parents and F1 hybrid performance, respectively. Since then, the GCA was widely used in cross breeding for elite parent selection. However, the molecular basis of GCA remains to unknown. We studied the transcriptomes of three varieties and three F1 hybrids using RNA-Sequencing. Transcriptome sequence analysis revealed that the transcriptome profiles of the F1s were similar to the positive GCA-effect parent. Moreover, the expression levels of most differentially expressed genes (DEGs) were equal to the parent with a positive GCA effect. Analysis of the gene expression patterns of gibberellic acid (GA) and flowering time pathways that determine plant height and flowering time in rice validated the preferential transcriptome expression of the parents with positive GCA effect. Furthermore, H3K36me3 modification bias in the Pseudo-Response Regulators (PRR) gene family was observed in the positive GCA effect parents and demonstrated that the phenotype and transcriptome bias in the positive GCA effect parents have been epigenetically regulated by either global modification or specific signaling pathways in rice. The results revealed that the transcriptome profiles and DEGs in the F1s were highly related to phenotype bias to the positive GCA-effect parent. The transcriptome bias toward high GCA parents in F1 hybrids attributed to H3K36me3 modification both on global modification level and specific signaling pathways. Our results indicated the transcriptome profile and epigenetic modification level bias to high GCA parents could be the molecular basis of GCA.BMC Genomics 04/2014; 15(1):297. · 4.40 Impact Factor
- International Journal of Molecular Sciences 07/2014; · 2.46 Impact Factor
, 1118 (2009);
et al.Ruth A. Swanson-Wagner,
Expression Patterns in Maize Hybrids
Paternal Dominance of Trans-eQTL Influences Gene
www.sciencemag.org (this information is current as of November 20, 2009 ):
The following resources related to this article are available online at
version of this article at:
including high-resolution figures, can be found in the online
Updated information and services,
can be found at:
Supporting Online Material
, 13 of which can be accessed for free:
cites 30 articles
1 articles hosted by HighWire Press; see:
This article has been
This article appears in the following
in whole or in part can be found at:
permission to reproduce
of this article or about obtaining
Information about obtaining
registered trademark of AAAS.
is aScience2009 by the American Association for the Advancement of Science; all rights reserved. The title
CopyrightAmerican Association for the Advancement of Science, 1200 New York Avenue NW, Washington, DC 20005.
(print ISSN 0036-8075; online ISSN 1095-9203) is published weekly, except the last week in December, by theScience
on November 20, 2009
Paternal Dominance of Trans-eQTL
Influences Gene Expression Patterns in
Ruth A. Swanson-Wagner, Rhonda DeCook,* Yi Jia, Tim Bancroft, Tieming Ji, Xuefeng Zhao,
Dan Nettleton, Patrick S. Schnable†
Heterosis refers to the superior performance of hybrid progeny relative to their inbred parents, but the
mechanisms responsible are unknown. Hybrids between the maize inbred lines B73 and Mo17 exhibit
30 to 50% of their genes are differentially expressed. We identified ~4000 expression quantitative trait
loci (eQTL) that allowed us to identify markers linked to variation in expression. We found that over
three-quarters of these eQTL act in trans (78%) and that 86% of these differentially regulate transcript
accumulation in a manner consistent with gene expression in the hybrid being regulated exclusively by
the paternally transmitted allele. This result suggests that widespread imprinting contributes to the
regulation of gene expression in maize hybrids.
underlying molecular mechanisms have not been
deciphered (2). There are widespread differences
in gene expression in the maize Mo17 × B73
hybrid relative to its inbred parents, B73 and
Mo17 (3–5). Because maize is monoecious, with
a physical separation between the male and
female flowers on a single plant, any given plant
can be used as both male and female parents of
and Mo17 × B73 hybrids are both highly het-
erotic but, despite having identical nuclear ge-
in development (table S1). Reciprocal effects on
phenotypes have been documented in plants (6)
but have not been widely investigated at the mo-
lecular level (7, 8).
Genome-wide transcript accumulation in the
B73 × Mo17 and Mo17 × B73 hybrids was
measured with a cDNA microarray (9), and the
analysis identified 1515 (~11%) significantly
differentially expressed genes with a 5% false
discovery rate (FDR) cutoff (table S2). Similar
sequencing experiment (tables S2 and S3). Al-
(9) that >50% (N = 7325/14,118) of the genes on
the array and ~33% (N = 871/2640) of the highly
expressed genes in the RNA sequencing exper-
iment were differentially expressed. Smaller pro-
portions of genes showing reciprocal expression
effects have been previously reported in several
Expression QTL (eQTL) experiments (14)
are typically conducted with recombinant inbred
eterosis, the enhanced agronomic per-
parents (1), is widely exploited, but the
lines (RILs) or other inbred genotypes and thus
are unable to determine the effects of heterozy-
gosity on gene expression patterns (10). We
examined eQTL in a set of 29 IBM (intermated
the maize B73 and Mo17 lines and hybrids
RIL) and Mo17 (Mo17 × RIL). These hybrids
provided a contrast between gene expression
regulation at heterozygous and homozygous
genotypes across all loci that are polymorphic
between B73 and Mo17 (Fig. 1A).
Separate eQTL analyses were conducted
within each cross type (B73 × RIL, Mo17 ×
RIL, and RIL) by scanning the genome with
ing the genetic map positions of differentially
regulated genes relative to the genetic positions
of the regulating eQTL. We defined an eQTL as
acting in cis if a regulated gene and its corre-
sponding eQTL were within 5 cM of each other.
Trans-eQTL were defined as those located on dif-
P values and estimated FDRs (9) resulted
in 1334 to 1904 significant eQTL associations
within each cross type (table S4). About 25% of
the significant genes were regulated in multiple
cross types. In the majority of such cases, the same
genomic region regulated the gene among multi-
ple cross types (table S5 and figs. S2 and S3).
Only 10% of the detected eQTL acted in cis,
which on average showed larger effects than the
previous reports for maize (4, 14) and other
species (15). Trans-eQTL with large effects may
be rare because they can regulate many genes or
regulated, could be detrimental (16, 17). Many
cis-regulated genes detected within the RIL cross
type were previously (5) identified as differen-
tially expressed between the B73 and Mo17
inbred lines (79/114) and exhibited consistent
directions of effect in the current study. Because
the RILs used for this study were mosaics of the
ity of these cis-eQTL across genetic backgrounds.
Nearly 80% of the eQTL acted in trans (table
S4 and fig. S3), consistent with reports from
other species (15, 18–23) and of trans-regulation
of diverse biological processes in maize (24–28).
Previous studies may have overestimated the num-
ber or proportion of cis-regulated genes (3, 29)
due to ascertainment bias because they only an-
alyzed genes containing single nucleotide poly-
morphisms between B73 and Mo17 and/or had
limited statistical power to detect the more subtle
effects of trans-eQTL (15).
Clustering of expression patterns of all genes
regulated by eQTL distinguished cis- and trans-
regulated genes (Fig. 1B). Most (93%) genes
with additive gene action are cis-regulated and
have similar amounts of expression in the two
heterozygotes (Mo17 × RILBB and B73 ×
RILMM; clusters 1 and 2, Fig. 1, A and B). In-
stead of showing additive gene action, most
(86%) trans-eQTL exhibit a mode of gene action
we term “paternal dominance” that is consistent
with imprinting, wherein expression values in
those heterozygotes with a RILBBas the paternal
parent matched expression values in lines that
were homozygous for the B allele of the trans-
eQTL; similarly, heterozygotes with a RILMMas
the paternal parent had expression values that
matched the expression values in lines that were
homozygous for the M allele of the trans-eQTL
(i.e., MB was not equal to BM; clusters 3 and 4,
Fig. 1B). Because our experimental design held
the maternally contributed allele in hybrids con-
stant,we could detect variation in paternally con-
tributed alleles (Fig. 1A) while controlling for
maternal or cytoplasmic effects. Hence, these re-
sults suggest that gene expression in the hybrids
is regulated exclusively by the paternally trans-
mitted allele of these trans-eQTL.
An eQTL marker (interval 377) associated
with the differential regulation of more than 20
genes was back-crossed into the Mo17 inbred
genetic background for multiple generations (fig.
S5). Seedlings of the BM and MM genotypes
generated at this interval in genetic backgrounds
that were otherwise 87.5% homozygous for the
expression that were consistent with patterns
observed in the eQTL analysis. Thus, this trans-
eQTL is stable across generations and technolo-
gies (table S6).
Because no direct comparisons were per-
formed between B73 × RILMMand Mo17 ×
RILBB, we validated the unusual paternal dom-
inance patterns of gene expression for five genes
regulated in trans by interval 377 by using a com-
bination of quantitative reverse transcription poly-
merase chain reaction and Sequenom (Sequenom,
assays (table S6).
It is not possible to conclude how widely
distributed paternal dominance is across species
because previous studies have most commonly
Iowa State University, Ames, IA 50011, USA.
*Present address: Department of Statistics and Actuarial
Science, University of Iowa, Iowa City, IA 52242, USA.
†To whom correspondence should be addressed. E-mail:
20 NOVEMBER 2009VOL 326
on November 20, 2009
experiments cannot define the mode of gene
action and cannot detect genomic imprinting
because the effects of heterozygosity and recip-
rocally generated genotypes are not typically
investigated. If we had only examined one of the
two hybrid cross types, the gene action of the
paternally dominant trans-eQTL would have
incorrectly been classified as Mendelian domi-
nant. Lastly, sufficient statistical power is needed
to detect the modest effects of trans-eQTL.
We hypothesize that at least some paternally
dominant trans-eQTL are small RNAs, because
small RNAs regulate gene expression in trans
(30, 31) and can be subject to parent-specific
genomic imprinting (32, 33). Because there are
many paternally dominant trans-eQTL in maize,
and many of these regulate multiple genes, their
effects are broadly propagated throughout the
transcriptome. Paternal dominance may, there-
fore, contribute to the observed phenotypic
differences between reciprocal hybrids.
References and Notes
1. G. H. Shull, Am. Breeders Assoc. 4, 296 (1908).
2. P. S. Schnable, R. A. Swanson-Wagner, in Handbook of
Maize: Its Biology, J. L. Bennetzen, S. C. Hake, Eds.
(Springer, New York, 2009), pp. 457–467.
3. M. Guo et al., Plant Mol. Biol. 66, 551 (2008).
4. R. M. Stupar, N. M. Springer, Genetics 173, 2199 (2006).
5. R. A. Swanson-Wagner et al., Proc. Natl. Acad. Sci. U.S.A.
103, 6805 (2006).
6. M. Gonzalo, T. J. Vyn, J. B. Holland, L. M. McIntyre,
Heredity 99, 14 (2007).
7. R. M. Stupar, P. J. Hermanson, N. M. Springer, Plant
Physiol. 145, 411 (2007).
8. N. Hoecker, B. Keller, H. P. Piepho, F. Hochholdinger,
Theor. Appl. Genet. 112, 421 (2006).
9. Materials and methods are available as supporting
material on Science Online.
10. G. Gibson et al., Genetics 167, 1791 (2004).
11. M. Vuylsteke, F. van Eeuwijk, P. Van Hummelen,
M. Kuiper, M. Zabeau, Genetics 171, 1267 (2005).
12. X. Wang et al., PLoS One 3, e3839 (2008).
13. P. J. Wittkopp, B. K. Haerum, A. G. Clark, Genetics 173,
14. E. E. Schadt et al., Nature 422, 297 (2003).
15. D. Kliebenstein, Annu. Rev. Plant Biol. 60, 93 (2009).
16. J. M. Aury et al., Nature 444, 171 (2006).
17. B. G. Hansen, B. A. Halkier, D. J. Kliebenstein, Trends
Plant Sci. 13, 72 (2008).
18. R. B. Brem, G. Yvert, R. Clinton, L. Kruglyak, Science 296,
752 (2002); published online 28 March 2002 (10.1126/
19. N. Hubner et al., Nat. Genet. 37, 243 (2005).
20. S. A. Monks et al., Am. J. Hum. Genet. 75, 1094 (2004).
21. M. Morley et al., Nature 430, 743 (2004).
22. M. A. West et al., Genetics 175, 1441 (2007).
23. G. Yvert et al., Nat. Genet. 35, 57 (2003).
24. X. Niu, T. Helentjaris, N. J. Bate, Plant Cell 14, 2565
25. F. T. Nogueira, A. K. Sarkar, D. H. Chitwood,
M. C. Timmermans, Cold Spring Harbor Symp. Quant.
Biol. 71, 157 (2006).
26. N. Satoh-Nagasawa, N. Nagasawa, S. Malcomber,
H. Sakai, D. Jackson, Nature 441, 227 (2006).
27. C. Shi et al., BMC Genomics 8, 22 (2007).
28. M. D. Yandeau-Nelson, B. J. Nikolau, P. S. Schnable,
Genetics 174, 101 (2006).
29. N. M. Springer, R. M. Stupar, Plant Cell 19, 2391 (2007).
30. G. Chuck, H. Candela, S. Hake, Curr. Opin. Plant Biol. 12,
31. J. A. Goodrich, J. F. Kugel, Crit. Rev. Biochem. Mol. Biol.
44, 3 (2009).
32. J. Brennecke et al., Science 322, 1387 (2008).
33. R. K. Slotkin et al., Cell 136, 461 (2009).
Cis n= 60
Trans n= 218
Other n= 30
Unmapped n= 105Total N= 413
Cis n= 228
Trans n= 136
Other n= 93
Unmapped n= 161Total N= 618
Cis n= 4
Trans n= 1,028
Other n= 113
Unmapped n= 259Total N= 1,404
Cis n= 17
Trans n= 1,138
Other n= 132
Unmapped n= 275Total N= 1,562
Cluster 1Cluster 2
Cluster 3Cluster 4
Fig. 1. Cis-andtrans-regulatedgenesexhibitdifferentpatternsofexpression.(A)BecausetheinbredIBMRILs
are derived from a cross between B73 and Mo17, a given IBM RIL is homozygous for either the B (B73) or M
(Mo17) allele of a given marker (denoted by a gray dashed line). We designate such RILs as RILBBand RILMM,
respectively. When crossed onto B73 (BB), RILBBand RILMMRILs will generate hybrids that are homozygous and
RILs will generate hybrids that are heterozygous and homozygous for the marker in question,respectively. Thus,
paternal parent is underlined. (B) Standardized expression values (y axis) were clustered to group genes with
cross type in which the expression was measured is indicated at the top of each plot (B73 × RIL, green; RIL,
orange; and Mo17 × RIL, purple), and the allele inherited from the paternal genome is underlined in thex-axis
of an eQTL locus show higher expression than lines homozygous for the M allele or vice versa. Clusters 3 and 4
both show paternal dominance but have opposite modes of gene action as described for clusters 1 and 2.
VOL 32620 NOVEMBER 2009
on November 20, 2009
34. We thank S. Hargreaves, M. Smith, M. Wilkening,
T. Kemmerer, P. Lu, C.-T. Yeh, and L. Coffey for sample
and data processing and the maize genome sequence
project (NSF DBI-0527192) for sharing genome sequences
before publication. Funding provided by Iowa State
University’s Plant Sciences Institute. Microarray and
sequencing data are available at the National Center for
BiotechnologyInformationGEOdatabase inseries GSE16136.
Supporting Online Material
Materials and Methods
Figs. S1 to S6
Tables S1 to S9
26 June 2009; accepted 15 October 2009
Symbiotic Nitrogen Fixation in the
Fungus Gardens of Leaf-Cutter Ants
Adrián A. Pinto-Tomás,1,2,3Mark A. Anderson,4Garret Suen,1,5David M. Stevenson,6
Fiona S. T. Chu,4W. Wallace Cleland,4Paul J. Weimer,6Cameron R. Currie1,5*
Bacteria-mediated acquisition of atmospheric N2serves as a critical source of nitrogen in terrestrial
ecosystems. Here we reveal that symbiotic nitrogen fixation facilitates the cultivation of specialized
fungal crops by leaf-cutter ants. By using acetylene reduction and stable isotope experiments,
we demonstrated that N2fixation occurred in the fungus gardens of eight leaf-cutter ant species
and, further, that this fixed nitrogen was incorporated into ant biomass. Symbiotic N2-fixing
bacteria were consistently isolated from the fungus gardens of 80 leaf-cutter ant colonies collected
in Argentina, Costa Rica, and Panama. The discovery of N2fixation within the leaf-cutter ant−microbe
symbiosis reveals a previously unrecognized nitrogen source in neotropical ecosystems.
mass and 86% of the arthropod biomass in trop-
ical forest canopies, and, in the Amazon forest,
they represent four times more biomass than do
all land vertebrates combined (1–3). Among
ants, the leaf cutters (tribe Attini: genera Atta
and Acromyrmex) play an important role as one
of the most dominant herbivores in New World
tropical ecosystems, stimulating new plant growth
and facilitating nutrient cycling (4). Mature Atta
colonies are among the largest of any social
insect, consisting of up to 8 million workers and
occupying an underground volume of more than
20 m3(Fig. 1, A and B) (5). These “superorga-
nisms” harvest more than 240 kg dry weight of
leaf material per year (4), which they use to cul-
tivate a fungus for food (6). This ability to grow
a specialized fungal crop using freshly cut plant
material is a key factor in the ecological success
of leaf-cutter ants (7). In addition to their rela-
tionship with fungal mutualists (family Lepiota-
ceae), the ants engage in a second mutualism
with Actinobacteria (genus Pseudonocardia),
which produce antibiotics to help defend the
nts play a critical role in shaping terres-
trial ecosystems. They make up at least
one-third of the global insect fauna bio-
fungus garden from parasites (8, 9). We explored
the possibility that leaf-cutter ants engage in mu-
tualistic associations with N2-fixing symbionts
to supplement the nitrogen budget of their fun-
Nitrogen is expected to be a growth-limiting
resource in leaf-cutter ant agriculture: The pri-
mary nutrient input into their colonies is fresh
leaves, which have a much lower nitrogen-to-
carbon (N:C) ratio than is required by insects
(10, 11). In contrast to this expectation, several
field studies have shown that the exhausted leaf
substrate removed from the bottom of fungus
gardens by ant workers contains higher pro-
portions of N than either freshly harvested leaf
material or surrounding leaf litter does, indi-
cating that N enrichment occurs as the plant
substrate is processed by the colony (4, 12, 13).
Although these findings suggest the presence of
N2-fixing symbionts, potential additional sources
are mineralized N from the soil and compen-
satory feeding by the ants (13, 14). We analyzed
the N content of laboratory-maintained colonies
of Atta cephalotes in which we prevented N in-
put from these alternate sources (15). We found
an increase in N content as leaf substrate passes
through the system: N content was lowest in fresh
leaf cuttings, significantly higher in the fungus
1550 Linden Drive, Madison, WI 53706, USA.2Departamento
de Bioquímica, Facultad de Medicina, Universidad de Costa
de Investigacionesen EstructurasMicroscópicas,Universidad de
Costa Rica, San Pedro de Montes de Oca, San José, Costa Rica.
4Institute for Enzyme Research, Department of Biochemistry,
University of Wisconsin–Madison, 1710 University Avenue,
Madison, WI 53726, USA.5U.S. Department of Energy Great
Lakes Bioenergy Research Center, University of Wisconsin–
Madison, 1550 Linden Drive, Madison, WI 53706, USA.6U.S.
Department of Agriculture–Agricultural Research Service, U.S.
Dairy Forage Research Center, 1925 Linden Drive West,
Madison, WI 53706, USA.
*To whom correspondence should be addressed. E-mail:
Fig. 1. Evidence for N2fixation in the fungus gardens of leaf-cutter ants. (A and B) The agricultural
system of leaf-cutter ants is extremely efficient, allowing colonies to grow from a single fungus garden
chamber [(A) incipient At. cephalotes colony with queen (black arrow) on top of fungus garden; scale
bar = 1 cm] to a massive underground operation with hundreds of chambers, intricate tunnel systems,
and millions of workers [(B) partially excavated nest of a mature Atta colony]. (C) Nitrogen content of
the different components of five At. cephalotes colonies. (D) N2-fixation activity measured by acetylene
reduction for different components of 10 Atta spp. colonies. All results are shown as means T SEM.
Means labeled with different letters (a to e) are statistically different (P < 0.05). [Photo credits: (A)
Graham D. Anderson, (B) M. Moffett/Minden Pictures]
20 NOVEMBER 2009VOL 326
on November 20, 2009