Lafora disease, seizures and sugars
D.M. AnDrADe1, J. Turnbull2, b.A. MinAssiAn2, 3
1 Division of Neurology, Krembil Neuroscience Centre, University of Toronto, Toronto Western Hospital, Toronto, Canada;
2 Program in Genetics and Genomic Biology, 3 Division of Neurology, Department of Paediatrics, The University of Toronto,
The Hospital for Sick Children, 555 University Ave. Toronto, Canada
Acta Myologica • 2007; XXVI; p. 83-86
Address for correspondence: Berge A Minassian, Division of Neurology, Department of Paediatrics, The Hospital for Sick Children, The
University of Toronto, 555 University Ave, Toronto, Ontario M5G 1X8, Canada. Fax +1 416 8136334. E-mail: email@example.com
Lafora disease (LD) is the most severe form of Progressive Myo-
clonus Epilepsy with teenage onset. It has an autosomal reces-
sive mode of inheritance and is almost universally fatal by the
second or third decade of life. To date, there is no prevention
or cure. In the last decade, with the identification of the genes
responsible for this disease, much knowledge has been gained
with the potential for the future development of effective treat-
ment. This review will briefly address clinical issues and will
focus on the molecular aspects of the disease.
Key words: Lafora disease, progressive myoclonus epilepsy, la-
Onset is usually between the ages of 12 and 17, al-
though it may occur as early as 6 or as late as 22. The
first symptom clearly recognized by the family is usually
a generalized tonic-clonic seizure. However, any of the
other symptoms or seizures that are seen in LD can occur
precede the major convulsion and sometimes go unrec-
ognized for a period of time. Other seizure types include
myoclonic seizures, occipital seizures, atypical absence,
atonic and complex partial seizures. The seizures may
initially respond to anticonvulsants, but they become in-
creasingly resistant to medication (1). Occipital seizures
are frequent and can manifest as transient blindness or
simple or complex visual hallucinations, although the lat-
ter may not be epileptic, and be a psychosis instead (2).
Myoclonus can be fragmentary, symmetric or massive,
and is usually the primary reason for early wheelchair
dependency. Other symptoms that commonly accompany
this syndrome include emotional disturbance, depression
and confusion early, later evolving into dementia. Nurs-
ing and feeding care can alter life span, but it still usu-
ally ranges between 17 and 30 years. Recent observations
report a slightly longer life span in patients with EPM2B
mutations, compared to those with EMP2A (see below).
Status epilepticus and aspiration pneumonia are the most
common causes of death (1, 3).
EEG abnormalities can be seen prior to the clinical
symptoms (1, 3). The background activity slows and the
normal alpha-rhythm as well as sleep features are lost
with disease progression. Photoconvulsive responses are
common. Interictal epileptiform abnormalities are seen
either with generalized, focal (especially in the occipital
regions) or multifocal distribution. Impaired cortical in-
hibitory mechanisms leading to hyperexcitability are also
represented by giant somatosensory and visual-evoked
potentials. Progressive prolongation of the central laten-
cies and of brainstem auditory responses are seen during
disease evolution (4).
Lafora bodies (LBs) are carbohydrate storage prod-
ucts that characterize LD and underlie the epileptic dis-
order. They are composed of polyglucosans, which are
abnormally formed glycogen molecules resembling
starch. The polyglucosans in LD consist of long chains of
glucose units that are infrequently branched. This makes
them insoluble, leading to their accumulation and forma-
tion of the LBs (3, 5). LBs stain strongly with periodic
acid-Schiff due to their polysaccharide composition, and
they are resistant to amylase digestion owing to dense
packing (6). Ultrastructural analysis suggests a physical
association between newly formed polyglucosans and en-
doplasmic reticulum (ER) or ER ribosomes (7).
LBs are found in brain, skin, liver, cardiac and skel-
etal muscle. However, despite this distribution, patients
usually do not have extra-neurological manifestations.
In skin, LBs are seen in either eccrine sweat gland duct
cells or in apocrine sweat gland myoepithelial cells.
Skin biopsy can be used for diagnosis if genetic test-
ing is not possible (8). In the central nervous system,
LBs are found in the perikarya or dendrites, but not in
axons. Perikaryal LBs can grow very large, outgrowing
the neuronal body and destroying the cell. However, the
total amount of LBs in dendrites exceed the perikaryal
accumulation (9). Very large numbers of small dendritic
D.M. Andrade et al
estingly HIRIP5, like laforin, is ubiquitously expressed
in subregions of the brain, but predominantly in the cer-
ebellum and hippocampus. This protein also co-local-
izes with laforin at the subcellular level. Finally, laforin
was able to dephosphorylate HIRIP5 on both tyrosine
and serine/threonine residues, suggesting that HIRIP5
is a substrate for laforin (24). A third protein shown to
interact with laforin, called PTG, is a regulatory subunit
of protein phosphatese-1 (PP1) that enhances glycogen
accumulation (21). It was shown that the G240S mis-
sense mutation identified in some LD patients disrupts
the interaction between laforin and PTG (while glyco-
gen binding and phosphatase activity remain preserved).
This observation suggests that PTG is critical for laforin
function and that laforin is part of a complex of pro-
teins associated with glycogen and may have a role in
regulating its metabolism. Studies using a mammalian
two-hybrid system demonstrated that laforin interacts
with glycogen synthase kinase-3 (GSK3). Furthermore,
laforin reduces GSK3 Ser 9 phosphorylation (25, 26).
GSK3 is a potent glycogen synthase (GS) inhibitor. The
relationship between GSK3, GS, laforin and LBs is dis-
EPM2B gene was identified through genome-wide
linkage scan followed by haplotype analysis and homozy-
gosity mapping performed in a cluster of French-Canadi-
an families from Quebec (11, 27). To date, 40 mutations
have been found in the EMP2B gene, including insertion,
missense and nonsense changes, frameshifts and deletions
in both compound heterozygous as well as homozygous
states. The EMP2B gene product encodes a 395 amino
acid protein named malin which contains a zinc finger of
the RING type at the N-terminus and six NHL-repeat mo-
tifs at the C-terminus. NHL motifs are likely involved in
protein-protein interactions, while the RING-finger motif
of malin is typical of E3 ubiquitin ligases. Sub-celllular
localization studies showed that MYC-tagged malin, sim-
ilarly to laforin, also localizes to the cytoplasm at the ER
and the nucleus (16, 17, 28).
The E3 ubiquitin ligase activity of malin was con-
firmed in vitro (25, 29). At least two mutations associated
with LD (Cys26Ser and Phe33Ser) result in inactivation
of malin’s ubiquitinase function (13, 25). Ubiquitination
can serve several purposes including targeting the ubiq-
uitinated protein for destruction or actively regulating its
function (30, 31). Recent studies demonstrated that la-
forin and malin interact and that this interaction occurs
at the central regions of both proteins (25, 29). There is
data suggesting that malin ubiquitinates laforin, targeting
laforin for destruction, but this is presently difficult to
understand, as destruction of laforin by malin would be
expected to result in Lafora disease (29). Finally, it was
demonstrated through co-immunoprecipitation studies
that malin and glycogen synthase (GS) interact, although
the result of such interaction is not known.
LBs in an exceedingly high number of dendrites may
play a role in the epileptic diathesis.
So far, two genes have been identified as causative of
LD, namely EMP2A and EPM2B (also known as NHL-
RC1) (10, 11). The proportion of LD patients with muta-
tions in one or the other gene varies according to the pop-
ulation studied. For instance, one Italian study showed
that EMP2A is mutated in 22% and EPM2B in 72% of
the patients (12). In our families, EPM2A and EMP2B
are mutated in 45% and 43%, respectively. Some biopsy
proven LD families do not have mutations in the coding
regions of those genes. Linkage and haplotype analysis
also excluded linkage to either of the two known genes,
suggesting the existence of a third LD locus (13).
Genotype-phenotype correlations are a challenge at
this point. However, some studies have suggested that EP-
M2B patients have a slower disease progression (12, 14).
Another correlation was suggested associating mutations
in the first exon of EMP2A to an early onset of cognitive
EMP2A gene is located on chromosome 6q24. It
consists of four exons coding for a 331 amino acid pro-
tein called laforin (10). Laforin has two isoforms, A and
B which localize to the ER and to the nucleus, respec-
tively (16, 17). The isoforms differ in their C-termini, and
mutations in the unique isoform A’s C-terminus suggests
that this is the disease-relevant isoform (17). To date, 40
different mutations and four polymorphisms were identi-
fied in this gene (18). These include missense and non-
sense mutations, frameshifts and deletions located in the
coding region of the gene.
Laforin is a unique protein in that it contains a carbo-
hydrate-binding domain (CBD) of the CBM20 type (19)
in its N-terminus and a dual-specificity protein tyrosine
phosphatase (DSP) domain in its C-terminus (6, 20).
Given the accumulation of polyglycosans in LD and the
presence of a CBD, laforin is thought to play an important
role in glycogen metabolism (either its synthesis or deg-
radation) (6). Importantly, self-dimerization appears to be
necessary for laforin to be functional in vivo (21, 22).
Co-immunoprecipitation studies suggest that full-
length laforin binds an uncharacterized protein termed
EMP2AIP1 (for EPM2A interacting protein). This pro-
tein does not appear to be responsible for LD in those
LD families with normal EPM2A and EPM2B genes
(23). HIRIP5 is another protein shown to interact with
laforin. This protein contains a NifU-like domain and
a putative MurD ligase domain. However the role of
those domains in HIRIP5 function is not yet clear. Inter-
Lafora disease, seizures and sugars
Animal models of Lafora Disease
Animal models of Lafora Disease known to date in-
clude a naturally occurring dog, one transgenic mouse
and one knockout mouse.
The canine model was observed in approximately 5%
of Miniature Wirehaired Dachshunds (MWHDs) in Eng-
land. The identified mutation was a dodecamer expansion
in the EPM2B gene (32). These animals have a phenotype
very similar to the human form of LD, except for the late
age of onset (age 6 in dogs, which is equivalent to adult-
hood in humans).
The transgenic model was created through overexpres-
sion of laforin carrying a phosphatase inactivating point
mutation (Cys26-6Ser). This dominant-negative model
was used to trap the unknown laforin substrate and produce
LD pathology. The transgenic LD mouse provided valu-
able information regarding the laforin protein localization.
In brain, this protein localizes to the neuronal soma and
dendrites. It was also demonstrated that laforin localizes
to the ER, but not to the ribosomes, as initially thought.
Importantly, this model allowed the characterization of la-
forin binding to different polysaccharides, and the prefer-
ential binding to polyglucosans over glycogen (33).
The knockout model was created through deletion
of the EPM2A exon containing the PTP domain. EPM2A
null mice had pathological evidence of LD as early as 2
months of age. Phenotypically, they had normal growth
and development until the age of 4 months, when be-
havioral changes started to occur. Myoclonic seizures,
ataxia and electroencephalographic changes were seen at
9 months. These animals showed a peculiar neurodegen-
eration mechanism involving organellar disintegration. In
addition, it was observed that neurodegeneration and the
onset of LB inclusions occurred prior to any behavioral
Pathophysiology of LBs
To date it is not clear why LBs are formed in the ab-
sence of laforin or malin. Evidence to date indicates that
both proteins are involved in the metabolism of glycogen.
Further supporting this hypothesis is the observation that
higher levels of cytosolic glycogen are correlated with
higher levels of laforin, while absence of glycogen is cor-
related with a 60% reduction of laforin. Mouse models
which over-express glycogen synthase and have massive
over-accumulation of glycogen with lafora-like bodies
show a 7-fold elevation of muscle laforin (35). However,
laforin binds preferentially to the polyglucosans forming
the LBs than to glycogen (33). This observation suggests
that polyglucosans do normally form in the cell, likely as
a by-product of glycogen metabolism, and laforin’s (and
possibly malin’s) proper function is important in prevent-
ing the accumulation of the toxic polyglucosans.
Crafting of properly branched and soluble glycogen
requires a number of coordinated enzymatic activities.
The complex between these enzymes and glycogen has
been termed the glycogenosome. A main component of
this complex is glycogen synthase (GS), which elongates
glycogen strands by adding glucose units. Glycogen
branching enzyme (BE) then moves the extended oli-
gosaccharides to branch points, maintaining the globular-
ity and solubility of glycogen. An abnormally high GS
to BE ratio results in inadequately branched polysaccha-
rides, namely polyglucosans (36).
Laforin may downregulate GS via PTG-PP1 and via
GSK3. PTG serves to target PP1 to the glycogenosome,
where PP1 activates GS by dephosphorylation (37). La-
forin binds PTG at PTG’s binding site with GS (21).
Laforin would therefore downregulate GS by physically
outcompeting PTG-PP1 off of GS. GSK3 is the main
inhibitor of GS, through phosphorylation of five phos-
phoregulatory sites on GS (37). Laforin activates GSK3
through dephosphorylation of GSK3 (25, 26). Laforin-ac-
tivated GSK3 would inactivate GS. In sum, absence of
laforin would lead to excess GS activity, GS/BE imbal-
ance, formation of insoluble polyglucosans, and their ac-
cumulation into LBs.
The concept of malin and laforin agonistically act-
ing to decrease GS activity in order to promote the right
GS/BE balance is in contrast with the observation that
malin polyubiquitinates laforin, targeting it for destruc-
tion (29). A possible explanation follows: LBs are much
more phosphorylated than glycogen, and are in fact more
similar to amylopectin than to glycogen. Laforin is able to
dephosphorylate amylopectin (38). Therefore, it is possi-
ble that laforin could also dephosphorylate LBs, and that
the high phosphate content in LBs, compared to normal
glycogen, may be a direct consequence of the mutated
laforin. Interestingly, glycogen binding appears to inhibit
laforin activity (39). Laforin inhibition may be a feedback
mechanism to preserve a certain degree of phosphoryla-
tion of the glycogen molecule. The role of glycogen de-
phosphorylation is not clear, but it may be correlated to
the maintenance of a properly branched polysaccharide.
If laforin activity needs to be kept in check (by glycogen
inhibition) to avoid over dephosphorylation of glycogen,
it is possible that a mutated malin would lead to lack of
ubiquitination and destruction of laforin. Could excess
laforin cause such an imbalance of glycogen dephospho-
rylation to lead to the formation of LBs? Finally, poly-
glucosans are even more potent inhibitors of laforin DSP
activity than normal glycogen. In that case, the initial
formation of polyglucosans (either because of mutated
laforin, malin or another yet unknown protein) would be
aggravated by the further inhibition of any residual la-
Much information has been gained in LD, but knowl-
edge remains very tentative. Clearly more data are need-
D.M. Andrade et al Download full-text
ed to understand the mechanisms causing LD, and maybe
then to find a way to make this disease go away.
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