- Research article
- Open Access
Conservation of the glucan phosphatase laforin is linked to rates of molecular evolution and the glucan metabolism of the organism
© Gentry and Pace; licensee BioMed Central Ltd. 2009
- Received: 20 February 2009
- Accepted: 22 June 2009
- Published: 22 June 2009
Lafora disease (LD) is a fatal autosomal recessive neurodegenerative disease. A hallmark of LD is cytoplasmic accumulation of insoluble glucans, called Lafora bodies (LBs). Mutations in the gene encoding the phosphatase laforin account for ~50% of LD cases, and this gene is conserved in all vertebrates. We recently demonstrated that laforin is the founding member of a unique class of phosphatases that dephosphorylate glucans.
Herein, we identify laforin orthologs in a protist and two invertebrate genomes, and report that laforin is absent in the vast majority of protozoan genomes and it is lacking in all other invertebrate genomes sequenced to date. We biochemically characterized recombinant proteins from the sea anemone Nematostella vectensis and the amphioxus Branchiostoma floridae to demonstrate that they are laforin orthologs. We demonstrate that the laforin gene has a unique evolutionary lineage; it is conserved in all vertebrates, a subclass of protists that metabolize insoluble glucans resembling LBs, and two invertebrates. We analyzed the intron-exon boundaries of the laforin genes in each organism and determine, based on recently published reports describing rates of molecular evolution in Branchiostoma and Nematostella, that the conservation of laforin is linked to the molecular rate of evolution and the glucan metabolism of an organism.
Our results alter the existing view of glucan phosphorylation/dephosphorylation and strongly suggest that glucan phosphorylation is a multi-Kingdom regulatory mechanism, encompassing at least some invertebrates. These results establish boundaries concerning which organisms contain laforin. Laforin is conserved in all vertebrates, it has been lost in the vast majority of lower organisms, and yet it is an ancient gene that is conserved in a subset of protists and invertebrates that have undergone slower rates of molecular evolution and/or metabolize a carbohydrate similar to LBs. Thus, the laforin gene holds a unique place in evolutionary biology and has yielded insights into glucan metabolism and the molecular etiology of Lafora disease.
- Carbohydrate Binding Module
- Intron Loss
- Dual Specificity Phosphatase
- Lafora Disease
Lafora disease (LD; OMIM 254780) is an autosomal recessive neurodegenerative disorder. It is one of five major progressive myoclonus epilepsies (PMEs) . LD commonly presents as a single seizure in the second decade of the patient's life, followed by progressive central nervous system degeneration, intellectual decline, and death within ten years of the first seizure [2–4]. LD is unique among the PMEs because of the patient's rapid neurological deterioration and the accumulation of insoluble glucans/carbohydrates called Lafora bodies (LB) [5, 6].
While animals normally store glucans as soluble glycogen, LBs are accumulations of poorly branched, hyperphosphorylated, insoluble glucans and are not glycogen. Forty years ago, Sakai and co-workers biochemically characterized LBs and found that they more closely resemble plant starch than glycogen [6–8]. Although LBs are found in the cytoplasm of most cells, cell death only occurs in neurons . LD patients exhibit increased neuronal cell death, number of seizures, and LB accumulation as they age; thus, it is hypothesized that LBs trigger these symptoms and ultimately the death of the patient .
We recently demonstrated that laforin is not restricted to vertebrate genomes, as originally thought , but that laforin orthologs are present in five protists (i.e. single-cell eukaryotes) . Each of these five protists undergoes hibernation during their life cycle and when they hibernate they generate an insoluble glucan as an energy source [16–18]. We recognized that the glucan produced by these protists are biochemically similar to Lafora bodies and proposed that laforin functions to convert insoluble glucans into energy, and in vertebrates it functions to inhibit insoluble glucan accumulation (i.e. LBs) .
To test this hypothesis, we utilized a recently characterized gene in Arabidopsis called s tarch ex cess 4 (SEX4) [19, 20]. SEX4 encodes a protein with similar domains as laforin but in the opposite orientation (Fig. 1B). Disruption of SEX4 leads to increased accumulation of insoluble glucans (i.e. starch), a cellular phenotype reminiscent of LD patients . We found that when laforin was targeted to the chloroplast of sex4-deficient plants that it rescued the sex4 mutant phenotype. Demonstrating that laforin and SEX4 are functional equivalents and that a laforin-like activity is required to regulate the metabolism of insoluble glucans in multiple Kingdoms .
We also demonstrated the nature of this activity; we showed that laforin and SEX4 dephosphorylate the glucan itself [15, 21, 22]. Thus, we proposed that when laforin is absent, phosphates accumulate in glycogen precursors, branching is inhibited, and Lafora bodies form. This prediction was supported by work published 40 years ago showing LBs from human patients are poorly branched and contain 4–5 fold more phosphate than glycogen, and further corroborated by data from the Roach lab confirming these results in a LD mouse model [23, 24]. Therefore, laforin regulates an overlooked aspect of glycogen metabolism in vertebrates by removing phosphate from glycans during glycogen synthesis.
Herein, we establish definitive boundaries concerning the evolutionary conservation of laforin by probing more than 210 eukaryotic genomes. We utilized criteria that we previously defined to correctly predict a laforin ortholog in the genome of the protozoan Neospora caninum. In addition, we extend our previous results by uncovering putative laforin orthologs in two invertebrates, Nematostella vectensis and Branchiostoma floridae. We cloned the respective genes and biochemically verified that they are laforin orthologs. Furthermore, we present evidence and hypothesize why laforin is conserved in these two invertebrates and is absent in all others sequenced to date. Cumulatively, these results demonstrate that the glucan phosphatase laforin is conserved in a subset of eukaryotic organisms from an array of evolutionary niches, vertebrates, invertebrates, and protists, and highlights the fundamental importance of glucan phosphorylation/dephosphorylation.
Carbohydrate binding modules (CBMs) are domains typically found in glucosylhydrolases and glucotransferases in bacterial, fungal, or plant genomes [25–27]. The vast majority of enzymes containing CBMs utilize the domain to bind a specific glucan and enzymatically act on the sugar, as in the case of α-amylase . Accordingly, we recently demonstrated, and others confirmed, that laforin and SEX4 bind and dephosphorylate glucans, glycogen and starch, respectively [15, 21, 23, 24]. While laforin and SEX4 bind similar types of glucans, they utilize evolutionarily distinct CBMs . CBMs are classified into fifty-three evolutionarily distinct families, based on primary sequence, secondary and tertiary predictions, and crystal structures . Laforin contains an amino-terminal CBM20 and SEX4 a carboxy-terminal CBM21 type CBM (Fig. 1A &1B). Although laforin and SEX4 have evolutionarily distinct CBMs, multiple groups have proposed that CBM20 and CBM21 may share a common evolutionary origin and suggest keeping distinct families grouped into a common CBM clan [28–31].
Confirmation of laforin predictions
Previously we reported that out of 170 eukaryotic genomes (including 94 protozoan) and 670 bacterial genomes that the laforin gene is only conserved in vertebrate genomes and in five protozoan genomes, Toxoplasma gondii, Eimeria tenella, Tetrahymena thermophila, Paramecium tetraurelia, and Cyanidioschyzon merolae  (Fig. 1C). In addition, we demonstrated that SEX4 is conserved in green algae and land plants (collectively known as Archaeplastida/Kingdom Plantae) and showed that while laforin and SEX4 are not orthologous proteins that they are functional equivalents  (Fig. 1C). Our phylogenetic analyses and examination of the biology and evolution of the five protists that have laforin led us to propose three criteria to predict if a protozoan genome would contain laforin: the organism must 1) be of red algal descent, 2) possess a true mitochondrion, and 3) produce an insoluble glucan (e.g. floridean starch, amylopectin granules, etc.) . Out of the 170 organisms that we probed, we found that if an organism lacked one of these qualities then it lacked laforin and if it possessed all of these qualities then it possessed laforin . Therefore, Plasmodium species lack laforin because they do not metabolize insoluble glucans, Cryptosporidium species lack laforin because they lack true mitochondria, and Chlamydomonas lack laforin because they are not of red-algal descent (and instead have SEX4) . Some of the genomes that we originally probed were incomplete at the time. However, based on the above three criteria, we postulated that the following four organisms currently being sequenced would contain laforin, Galdieria sulphuraria, Guillardia theta, Neospora caninum, and Sarcocystis neurona .
In light of the relatedness between CBM20 and CBM21 CBMs, we probed 122 protozoan genomes (28 more than previously) searching for a protein containing a CBM20 or CBM21 domain followed by a phosphatase domain. In order to enhance our likelihood of uncovering a laforin ortholog, we performed BLASTp and tBLASTn searches of multiple databases (Additional File 1) using human (Hs-) and C. merolae (Cm-) laforin, as C. merolae is likely the most evolutionarily ancient organism with laforin and Cm-laforin was the least identical (25%) to Hs-laforin of all the protozoan laforin orthologs previously identified . To ensure that we did not miss a laforin-like protein in these genomes, we also searched the same databases using the same search methods for proteins with a DSP domain followed by a CBM20 or CBM21, i.e. SEX4 orthologs.
Discovery of putative laforin orthologs in invertebrate genomes
Metazoans are defined as all living animals that contain tissues and are descended from the last common ancestor of Bilateria, Cnidaria (jellyfish, sea anemones, corals, hydra, etc.), Ctenophora (comb jellies), Placozoa (Trichoplax sp.), and Porifera (sponges) . Bilateralians are subdivided as either protostomes (including arthropods, nematodes, annelids, and mollusks) or deuterostomes. The three major deuterstome phyla, chordates, echinoderms (sea urchins, sea stars, etc.), and hemichordates (acorn worm), arose from a common ancestor more than 600 million years ago, followed by subsequent divergence of the chordates into three subphyla: cephalochordates, urochordates (also called tunicates), and vertebrates (summarized in Additional File 2) [39, 40].
The gene encoding laforin is an evolutionarily ancient gene, originating in a primitive red alga, or its ancestor, long before the emergence of metazoans . While laforin is an ancient gene, it has a unique evolutionary lineage. Although we previously identified laforin orthologs in five protozoan genomes, we did not find it in any non-vertebrate model organism genomes (yeast, fly, or worms), nor did we find it in the genome of any invertebrates . We postulated that invertebrates lack laforin because they do not synthesize an insoluble glucan as an energy source (as do the protozoans that contain laforin) and they do not inhibit insoluble glucan accumulation (as seen with vertebrates inhibiting LBs). However, over the course of the last two years multiple basal position metazoan genomes have been sequenced or improved and a surplus of information has been gleaned by the evo-devo community from the genomes of Branchiostoma floridae (commonly known as amphioxus, subphylum cephalochordata), Ciona intestinalis (sea squirt belonging to urochordates), Monosiga brevicollis (choanoflagellate and closest known unicellular relative to metazoans), Nematostella vectensis (sea anemone belonging to the ancient metazoan Phylum Cnidaria), and Trichoplax adhaerens (arguably the simplest free-living metazoan, Phylum Placozoa) [41–45]. While definitive conclusions concerning the origin and early radiation of these organisms in the metazoan tree of life remain unsettled, these reports have elucidated multiple aspects of metazoan evolution and the genomes of primitive metazoans regarding genome complexity, exon-intron structure, gene repertoire, and rates of molecular evolution.
To gain insight into the evolution of these putative laforin orthologs, we analyzed the gene structure and intron-exon boundaries of each and compared them with the gene encoding Hs-laforin, EPM2A. EPM2A is comprised of four exons and three introns (Fig. 3C) [9, 10]. Similarly, Bf-laforin-264244 has the same arrangement and the intron-exon boundaries are at similar locations (Fig. 3C). The other two putative Bf-laforin orthologs each contain three exons and two introns. Additionally, exon 1 from these two Bf-laforins is similar in size to exon 1 and exon 2 of both Hs-laforin and Bf-laforin-264244, suggesting possible intron loss (Fig. 3C). Finally, Nv-laforin has no introns, suggesting intron loss and potentially an increased rate of evolution at this locus in Nematostella.
Molecular evolution and conservation of lafoin.
42, 44, 48
44, 46, 48
40, 43, 44, 46, 48
42, 43, 44, 46, 47
40, 42, 43, 44, 46, 47, 48
The laforin gene in H. sapiens, T. gondii, N. caninum, E. tenella, P. tetraurelia, and B. floridae all contain at least four exons. Therefore, we propose that the laforin gene originally had four or more exons, and that the introns were lost in C. merolae and Nematostella. These results suggest that the laforin gene locus underwent a higher rate of molecular evolution than genes in Nematostella that share conserved introns with human genes. This increased molecular evolution may explain the absence of laforin in yeast, flies, worms, and the majority of invertebrates as we discuss below.
Biochemical characterization of Nv- and Bf-laforin
To determine if we had identified true laforin orthologs in invertebrates, we cloned the genes expressing Nv-laforin and Bf-laforin-264244 from Nematostella and Branchiostoma, respectively. We previously cloned and characterized Tg-laforin and found that we could only obtain soluble recombinant Tg-laforin when we added a GST tag to the amino terminus of Tg-laforin [, and unpublished data]. Even with the addition of the GST tag, the majority of GST-Tg-laforin is insoluble, suggesting that the majority of the protein does not correctly fold in bacteria. Unlike Hs-laforin but similar to Tg-laforin, both Nv- and Bf-laforin were largely insoluble with a HIS6 epitope. Therefore, we generated bacterial constructs expressing GST-Nv-laforin-HIS6 and GST-Bf-laforin-HIS6, purified the recombinant proteins (Additional File 3), and biochemically characterized them.
Bf- and Nv-laforin possess the same three in vitro biochemical properties as Hs-laforin: both utilize p-NPP as an artificial substrate, bind amylopectin, and liberate phosphate from amylopectin. Additionally, the invertebrate laforin orthologs contain the critical signature primary amino acids of both a CBM20 and DSP, and they possess the same predicted secondary structure as Hs-laforin. Thus, our bioinformatics searches for a protein containing a CBM and DSP correctly predicted the proteins biochemical properties and we identified novel laforin orthologs. Our finding of laforin orthologs in two invertebrate genomes reinforces the global function of this protein and yields insights into the evolution of this gene family.
Evolutionary lineage of glucan phosphatases
As discussed above, we previously reported that laforin is not confined to vertebrate genomes, but it is also conserved in five protozoan genomes . The gene encoding laforin is conserved in species as divergent as humans and red algae, but it is absent in the vast majority of protozoan and invertebrate genomes. SEX4 is conserved in diverse members of Archaeplastida/Kingdom Plantae and is necessary for proper starch metabolism in Arabidopsis [15, 19, 20]. These findings suggest that laforin and SEX4 are ancient proteins that regulate an aspect of energy metabolism conserved in multiple kingdoms, namely the dephosphorylation of glycogen and starch.
We previously postulated that throughout evolution organisms maintained or lost laforin depending on their "need" to manage insoluble glucans . We suggested that protists with laforin maintained it to manage insoluble glucans as an energy source . Similarly, vertebrates maintained laforin because of their "need" to combat Lafora body accumulation and used this reasoning to explain why the genome of most metazoans (i.e. animals) lack laforin . However, it is also possible that vertebrate genomes have laforin as a result of horizontal gene transfer (HGT) from a protist.
In searching for SEX4 orthologs, we identified full-length orthologs in fourteen genomes, encompassing trees, land plants, a moss (Physcomitrella patens), and a single-cell green alga (Fig. 7B). In addition, we identified several partial cDNA or protein hits of SEX4 orthologs in other Archaeplastida (data not shown). These results demonstrate that SEX4 is conserved in all branches of Archaeplastida/Kingdom Plantae and highlight the importance of this glucan phosphatase throughout Archaeplastida.
Our finding of laforin in two invertebrate genomes raises questions regarding the evolutionary lineage of laforin in metazoans/animals. It is possible that the unique evolutionary lineage of laforin was a result of HGT from a protozoan or its ancestor. If so, this event occurred earlier than the radiation of metazoans because laforin is conserved in two invertebrate genomes, including the Cnidarian Nematostella.
An alternative hypothesis, and one that we favour, postulates that in non-protozoan genomes the absence or presence of laforin was determined by rates of molecular evolution and the need to combat insoluble glucans, while in protists conservation of laforin is determined by the three criteria previously presented and discussed . This hypothesis seems likely given recent reports showing the rate of molecular evolution in Branchiostoma and Nematostella is similar to that of vertebrates and the rate is much higher in other invertebrates, as discussed above (Table 1).
This hypothesis calls into question why laforin has been conserved. Invertebrates may not need laforin because they have significantly shorter lifespans than vertebrates. In support of this thought, the only cellular pathology observed in LD patients is neuronal apoptosis . Since neurons live longer than most all other cell types, it seems plausible that invertebrates may be more like skin cells (or other cell types with shorter lifespans); they accumulate LBs but not to a detrimental state. Thus, laforin may represent a vestigial gene or perhaps a pseudogene in Nematostella and Branchiostoma.
Alternatively, Nematostella and Branchiostoma may have laforin for a purpose. Vertebrates do not utilize insoluble glucans as an energy source, but they do "combat" the accumulation of insoluble glucans, in the form of detrimental LBs. We know that many, if not all, vertebrate species suffer from LD (including, canines, felines, cattle, and birds) [60–64]. It is possible that Nematostella and Branchiostoma also have laforin in order to combat LBs.
Lastly, Nematostella and Branchiostoma may have laforin because their genomes have not evolved as rapidly as other invertebrates, as has been recently demonstrated, and thus still have/need laforin [42, 44]. If this is the case, other invertebrates (e.g flies and worms) may not need laforin because they have evolved a separate means to deal with LB-like accumulations.
Regardless of what has driven the evolutionary lineage of laforin, these results define the boundaries of glucan phosphatase conservation. These findings coupled with our previous work establish the conservation of laforin and SEX4 in evolutionary niches from protozoans to plants to algae to vertebrates and now invertebrates. Notably, laforin and SEX4 are absent in bacteria and archaea. While these groups do contain each domain that is found in glucan phosphatases (i.e. a phosphatase domain and carbohydrate binding domain), no protein in their genomes contains both. Given the completeness of bacterial genomes, it is unlikely that a glucan phosphatase exists in bacteria. Therefore, we suspect that we have identified the complete evolutionary lineage of the glucan phosphatases laforin and SEX4. The conservation of laforin across evolutionary niches, coupled with what appears to be complete conservation of SEX4 throughout all Archaeplastida/Kingdom Plantae demonstrate that phosphorylation/dephosphorylation of glucans is pervasive throughout nature.
Plasmids and Proteins
Wild type and C/S Hs-laforin in pET21a (Novagen, San Diego, CA) for use in bacterial expression were described previously [13, 65]. Recombinant HIS-tagged VHR cloning and purification have been previously described . The complete open reading frame of Bf-laforin-264224 was amplified from cDNA provided by the Branchiostoma floridea Gene Collection . The complete open reading frame of Nv-laforin was amplified from Nematostella DNA provided by Dr. Mark Q. Martindale. Bf- and Nv-laforin were cloned into pET-GSTX and pET21a, respectively . Mutations were introduced using QuickChange (Stratagene). Recombinant GST- and His-tagged proteins were expressed in Escherichia coli BL21 (DE3) CodonPlus RIL cells (Stratagene, La Jolla, CA) and purified using Ni2+-agarose (Qiagen, Germany) and/or glutathione-agarose affinity chromatography steps as described previously .
Phosphatase Activity Assays
Hydrolysis of para-nitrophenylphosphate (p-NPP) was performed in 50 μl reactions containing 1X phosphate buffer (0.1 M sodium acetate, 0.05 M bis-Tris, 0.05 M Tris-HCl, 2 mM dithiothreitol, at the appropriate pH), 50 mM pNPP, and 100–500 ng of enzyme at 37°C for 5–30 minutes. The reaction was terminated by the addition of 200 μl of 0.25 M NaOH and absorbance was measured at 410 nm. We tested the specific activity of each enzyme at pH 5.0, 5.5, 6.0, 6.5, 7.0, 7.5, and 8.0. Malachite green assays were performed as described  with the following modifications: 1X phosphate buffer, 100–500 ng of enzyme, and ≈45 μg of amylopectin in a final volume of 20 μl. The reaction was stopped by the addition of 20 μl of 0.1 M N-ethylmaleimide and 80 μl of malachite green reagent. Absorbance was measured after 30 minutes at 620 nm. We tested the specific activity of each enzyme at the same pH units as above.
Carbohydrate binding Assay
Carbohydrate binding assays were done similarly as described previously . Briefly, 50 mg of amylopectin was dissolved in 400 μl of ethanol, followed by the addition of 1 ml of water and 1 ml of 2 M NaOH, 2 ml of water was added, pH was adjusted to 6.5, and the total volume was brought to 10 ml. 1 ml of 5 mg/ml amylopectin was centrifuged at 50 K for 1.5 hours, the pellet was resuspended in 0.5 ml of buffer (50 mM Tris, pH7.5, 150 mM NaCl, 0.1% β-mercaptoethanol) with protease inhibitors, 0.5 μg of recombinant protein was added, the tube was rotated at 4°C for 1 hour, a second centrifugation was preformed at 50 K for 1.5 hours, the proteins in the supernatant were precipitated with acetone, and the pellet was resuspended in Western loading dye. Potato amylopectin was purchased from Sigma (St. Louis, MO). Recombinant proteins were detected with α-HIS-horeseradish peroxidase (HRP) antibody (Santa Cruz) and SuperSignal West Pico (Pierce).
Search strategy, sequence alignment, and phylogenetic analyses
The sequences of laforin and SEX4 orthologs were obtained by performing tBLASTn searches using the GenBank "dbEST" database or BLASTp and PSI-BLAST  searches using GenBank "eukaryote genome" and "non-redundant" (nr) databases, the Cyanidioschyzon merolae genome project, Department of Energy Joint Genome Institute Resource, The Institute for Genomic Research (TIGR), ToxoDB, GeneDB, Genoscope, UCSC Genome Browser, Tetrahymena Genome Database, and multiple organism specific databases. Accession numbers for small subunit (SSU) ribosomal RNA (rRNA), SEX4 orthologs, and laforin orthologs are listed in Additional File 5, 6, and 8, respectively. The web address for each database is listed in Additional File 4. A list of each genome that we investigated and a reason why an organism's genome lacks laforin is listed in Additional File 7. Amino-acid sequences of laforin orthologs were aligned by ClustalW  and refined manually using MacVector. SSU rRNA sequences were obtained by performing BLASTn using GenBank from "all organisms" and "nr" databases and accession numbers are listed in Additional File 5. Phylogenetic trees were generated from a ClustalW  multiple sequence alignment using PROTDIST and FITCH from the PHYLIP 3.65 software package and displayed utilizing HYPERTREE 1.0.0 .
We thank Drs. Doug Andres, Seema Mattoo, and Carolyn Worby for insightful discussions; Dr. Yutaka Satou and the Branchiostoma floridea Gene Collection for B. floridea cDNA; and Drs. Mark Q. Martindale and John R. Finnerty for N. vectensis DNA. This work was supported by National Institutes of Health grants 5R00NS061803 and 5P20RR0202171 (to M.S.G.) and University of Kentucky College of Medicine start-up funds.
- Berkovic SF, Andermann F, Carpenter S, Wolfe LS: Progressive myoclonus epilepsies: specific causes and diagnosis. N Engl J Med. 1986, 315 (5): 296-305.View ArticlePubMedGoogle Scholar
- Berkovic SF, Cochius J, Andermann E, Andermann F: Progressive myoclonus epilepsies: clinical and genetic aspects. Epilepsia. 1993, 34 (Suppl 3): S19-30.PubMedGoogle Scholar
- Minassian BA: Lafora's disease: towards a clinical, pathologic, and molecular synthesis. Pediatr Neurol. 2001, 25 (1): 21-29. 10.1016/S0887-8994(00)00276-9.View ArticlePubMedGoogle Scholar
- Van Heycop Ten Ham MW: Lafora disease, a form of progressive myoclonus epilepsy. Handbook of Clinical neurology. Edited by: Vinken PJ, Bryun GW. 1975, Holland, Amsterdam: North Holland Publishing Company, 15: 382-422.Google Scholar
- Lafora G, Glick B: Beitrag zur histopathologie der myoklonischen epilepsie. Z Ges Neurol Psychiatr. 1911, 6: 1-14. 10.1007/BF02863929.View ArticleGoogle Scholar
- Yokoi S, Austin J, Witmer F, Sakai M: Studies in myoclonus epilepsy (Lafora body form). I. Isolation and preliminary characterization of Lafora bodies in two cases. Arch Neurol. 1968, 19 (1): 15-33.View ArticlePubMedGoogle Scholar
- Sakai M, Austin J, Witmer F, Trueb L: Studies in myoclonus epilepsy (Lafora body form). II. Polyglucosans in the systemic deposits of myoclonus epilepsy and in corpora amylacea. Neurology. 1970, 20 (2): 160-176.View ArticlePubMedGoogle Scholar
- Yokoi S, Austin J, Witmer F: Isolation and characterization of Lafora bodies in two cases of myoclonus epilepsy. Journal of Neuropathology and Experimental Neurology. 1967, 26 (1): 125-127.PubMedGoogle Scholar
- Minassian BA, Lee JR, Herbrick JA, Huizenga J, Soder S, Mungall AJ, Dunham I, Gardner R, Fong CY, Carpenter S, et al: Mutations in a gene encoding a novel protein tyrosine phosphatase cause progressive myoclonus epilepsy. Nat Genet. 1998, 20 (2): 171-174. 10.1038/2470.View ArticlePubMedGoogle Scholar
- Serratosa JM, Gomez-Garre P, Gallardo ME, Anta B, de Bernabe DB, Lindhout D, Augustijn PB, Tassinari CA, Malafosse RM, Topcu M, et al: A novel protein tyrosine phosphatase gene is mutated in progressive myoclonus epilepsy of the Lafora type (EPM2). Hum Mol Genet. 1999, 8 (2): 345-352. 10.1093/hmg/8.2.345.View ArticlePubMedGoogle Scholar
- Yuvaniyama J, Denu JM, Dixon JE, Saper MA: Crystal Structure of the Dual Specificity Protein Phosphatase VHR. Science. 1996, 272: 1328-1331. 10.1126/science.272.5266.1328.View ArticlePubMedGoogle Scholar
- Ganesh S, Agarwala KL, Ueda K, Akagi T, Shoda K, Usui T, Hashikawa T, Osada H, Delgado-Escueta AV, Yamakawa K: Laforin, defective in the progressive myoclonus epilepsy of Lafora type, is a dual-specificity phosphatase associated with polyribosomes. Hum Mol Genet. 2000, 9 (15): 2251-2261.View ArticlePubMedGoogle Scholar
- Wang J, Stuckey JA, Wishart MJ, Dixon JE: A unique carbohydrate binding domain targets the lafora disease phosphatase to glycogen. J Biol Chem. 2002, 277 (4): 2377-2380. 10.1074/jbc.C100686200.View ArticlePubMedGoogle Scholar
- Ganesh S, Tsurutani N, Suzuki T, Hoshii Y, Ishihara T, Delgado-Escueta AV, Yamakawa K: The carbohydrate-binding domain of Lafora disease protein targets Lafora polyglucosan bodies. Biochem Biophys Res Commun. 2004, 313 (4): 1101-1109. 10.1016/j.bbrc.2003.12.043.View ArticlePubMedGoogle Scholar
- Gentry MS, Dowen RH, Worby CA, Mattoo S, Ecker JR, Dixon JE: The phosphatase laforin crosses evolutionary boundaries and links carbohydrate metabolism to neuronal disease. J Cell Biol. 2007, 178 (3): 477-488. 10.1083/jcb.200704094.PubMed CentralView ArticlePubMedGoogle Scholar
- Coppin A, Dzierszinski F, Legrand S, Mortuaire M, Ferguson D, Tomavo S: Developmentally regulated biosynthesis of carbohydrate and storage polysaccharide during differentiation and tissue cyst formation in Toxoplasma gondii. Biochimie. 2003, 85 (3–4): 353-10.1016/S0300-9084(03)00076-2.View ArticlePubMedGoogle Scholar
- Coppin A, Varré J, Lienard L, Dauvillée D, Guérardel Y, Soyer-Gobillard M, Buléon A, Ball S, Stanislas Tomavo: Evolution of Plant-Like Crystalline Storage Polysaccharide in the Protozoan Parasite Toxoplasma gondii Argues for a Red Alga Ancestry. Journal of Molecular Evolution. 2005, 60 (2): 257-267. 10.1007/s00239-004-0185-6.View ArticlePubMedGoogle Scholar
- Guérardel Y, Leleu D, Coppin A, Liénard L, Slomianny C, Strecker G, Ball S, Tomavo S: Amylopectin biogenesis and characterization in the protozoan parasite Toxoplasma gondii, the intracellular development of which is restricted in the HepG2 cell line. Microbes and Infection. 2005, 7 (1): 41-48. 10.1016/j.micinf.2004.09.007.View ArticlePubMedGoogle Scholar
- Niittyla T, Comparot-Moss S, Lue W-L, Messerli G, Trevisan M, Seymour MDJ, Gatehouse JA, Villadsen D, Smith SM, Chen J, et al: Similar protein phosphatases control starch metabolism in plants and glycogen metabolism in mammals. J Biol Chem. 2006, 281 (17): 11815-11818. 10.1074/jbc.M600519200.View ArticlePubMedGoogle Scholar
- Sokolov LN, Dominguez-Solis JR, Allary AL, Buchanan BB, Luan S: A redox-regulated chloroplast protein phosphatase binds to starch diurnally and functions in its accumulation. Proc Natl Acad Sci USA. 2006, 103 (25): 9732-9737. 10.1073/pnas.0603329103.PubMed CentralView ArticlePubMedGoogle Scholar
- Worby CA, Gentry MS, Dixon JE: Laforin: A dual specificity phosphatase that dephosphorylates complex carbohydrates. J Biol Chem. 2006, 281 (41): 30412-30418. 10.1074/jbc.M606117200.PubMed CentralView ArticlePubMedGoogle Scholar
- Kotting O, Santelia D, Edner C, Eicke S, Marthaler T, Gentry MS, Comparot-Moss S, Chen J, Smith AM, Steup M, et al: STARCH-EXCESS4 Is a Laforin-Like Phosphoglucan Phosphatase Required for Starch Degradation in Arabidopsis thaliana. Plant Cell. 2009, 21 (1): 334-346. 10.1105/tpc.108.064360.PubMed CentralView ArticlePubMedGoogle Scholar
- Tagliabracci VS, Turnbull J, Wang W, Girard JM, Zhao X, Skurat AV, Delgado-Escueta AV, Minassian BA, Depaoli-Roach AA, Roach PJ: Laforin is a glycogen phosphatase, deficiency of which leads to elevated phosphorylation of glycogen in vivo. Proc Natl Acad Sci USA. 2007, 104 (49): 19262-19266. 10.1073/pnas.0707952104.PubMed CentralView ArticlePubMedGoogle Scholar
- Tagliabracci VS, Girard JM, Segvich D, Meyer C, Turnbull J, Zhao X, Minassian BA, Depaoli-Roach AA, Roach PJ: Abnormal metabolism of glycogen phosphate as a cause for lafora disease. J Biol Chem. 2008, 283 (49): 33816-33825. 10.1074/jbc.M807428200.PubMed CentralView ArticlePubMedGoogle Scholar
- Boraston AB, Bolam DN, Gilbert HJ, Daview GJ: Carbohydrate-binding modules: fine-tuning polysaccharide recognition. The Biochemical Journal. 2004, 382: 769-781. 10.1042/BJ20040892.PubMed CentralView ArticlePubMedGoogle Scholar
- Coutinho PM, Henrissat B: Carbohydrate-active enzymes: an integrated database approach. Recent Advances in Carbohydrate Bioengineering. Edited by: Gilbert HJGD, Henrissat B, Svensson B. 1999, Cambridge: The Royal Society of Chemistry, 3-12.Google Scholar
- Rodriguez-Sanoja R, Oviedo N, Sanchez S: Microbial starch-binding domain. Curr Opin Microbiol. 2005, 8 (3): 260-267. 10.1016/j.mib.2005.04.013.View ArticlePubMedGoogle Scholar
- Machovic M, Janecek S: Starch-binding domains in the post-genome era. Cell Mol Life Sci. 2006, 63 (23): 2710-2724. 10.1007/s00018-006-6246-9.View ArticlePubMedGoogle Scholar
- Machovic M, Svensson B, MacGregor EA, Janecek S: A new clan of CBM families based on bioinformatics of starch-binding domains from families CBM20 and CBM21. FEBS J. 2005, 272 (21): 5497-5513. 10.1111/j.1742-4658.2005.04942.x.View ArticlePubMedGoogle Scholar
- Svensson B, Jespersen H, Sierks MR, MacGregor EA: Sequence homology between putative raw-starch binding domains from different starch-degrading enzymes. Biochem J. 1989, 264 (1): 309-311.PubMed CentralView ArticlePubMedGoogle Scholar
- Janecek S, Sevcik J: The evolution of starch-binding domain. FEBS Lett. 1999, 456 (1): 119-125. 10.1016/S0014-5793(99)00919-9.View ArticlePubMedGoogle Scholar
- Speer CA, Dubey JP, McAllister MM, Blixt JA: Comparative ultrastructure of tachyzoites, bradyzoites, and tissue cysts of Neospora caninum and Toxoplasma gondii. Int J Parasitol. 1999, 29 (10): 1509-1519. 10.1016/S0020-7519(99)00132-0.View ArticlePubMedGoogle Scholar
- Embley TM, Martin W: Eukaryotic evolution, changes and challenges. Nature. 2006, 440 (7084): 623-630. 10.1038/nature04546.View ArticlePubMedGoogle Scholar
- Cavalier-Smith T: Principles of Protein and LIpid Targeting in Secondary Symbiogenesis: Euglenoid, Dinoflagellatae, and Sporozoan Plastid Origins and the Eukaryote Family Tree. The Journal of Eukaryotic Microbiology. 1999, 46 (4): 347-366. 10.1111/j.1550-7408.1999.tb04614.x.View ArticlePubMedGoogle Scholar
- Cavalier-Smith T: Only six kingdoms of life. Proc Biol Sci. 2004, 271 (1545): 1251-1262. 10.1098/rspb.2004.2705.PubMed CentralView ArticlePubMedGoogle Scholar
- Dubey JP: Neosporosis – the first decade of research. Int J Parasitol. 1999, 29 (10): 1485-1488. 10.1016/S0020-7519(99)00134-4.View ArticlePubMedGoogle Scholar
- Dubey JP, Carpenter JL, Speer CA, Topper MJ, Uggla A: Newly recognized fatal protozoan disease of dogs. J Am Vet Med Assoc. 1988, 192 (9): 1269-1285.PubMedGoogle Scholar
- Animals. Metazoa. [http://tolweb.org/Animals/2374/2002.01.01]
- Peterson KJ, Butterfield NJ: Origin of the Eumetazoa: testing ecological predictions of molecular clocks against the Proterozoic fossil record. Proc Natl Acad Sci USA. 2005, 102 (27): 9547-9552. 10.1073/pnas.0503660102.PubMed CentralView ArticlePubMedGoogle Scholar
- Peterson KJ, Lyons JB, Nowak KS, Takacs CM, Wargo MJ, McPeek MA: Estimating metazoan divergence times with a molecular clock. Proc Natl Acad Sci USA. 2004, 101 (17): 6536-6541. 10.1073/pnas.0401670101.PubMed CentralView ArticlePubMedGoogle Scholar
- King N, Westbrook MJ, Young SL, Kuo A, Abedin M, Chapman J, Fairclough S, Hellsten U, Isogai Y, Letunic I, et al: The genome of the choanoflagellate Monosiga brevicollis and the origin of metazoans. Nature. 2008, 451 (7180): 783-788. 10.1038/nature06617.PubMed CentralView ArticlePubMedGoogle Scholar
- Putnam NH, Butts T, Ferrier DE, Furlong RF, Hellsten U, Kawashima T, Robinson-Rechavi M, Shoguchi E, Terry A, Yu JK, et al: The amphioxus genome and the evolution of the chordate karyotype. Nature. 2008, 453 (7198): 1064-1071. 10.1038/nature06967.View ArticlePubMedGoogle Scholar
- Srivastava M, Begovic E, Chapman J, Putnam NH, Hellsten U, Kawashima T, Kuo A, Mitros T, Salamov A, Carpenter ML, et al: The Trichoplax genome and the nature of placozoans. Nature. 2008, 454 (7207): 955-960. 10.1038/nature07191.View ArticlePubMedGoogle Scholar
- Putnam NH, Srivastava M, Hellsten U, Dirks B, Chapman J, Salamov A, Terry A, Shapiro H, Lindquist E, Kapitonov VV, et al: Sea anemone genome reveals ancestral eumetazoan gene repertoire and genomic organization. Science. 2007, 317 (5834): 86-94. 10.1126/science.1139158.View ArticlePubMedGoogle Scholar
- Holland LZ, Albalat R, Azumi K, Benito-Gutierrez E, Blow MJ, Bronner-Fraser M, Brunet F, Butts T, Candiani S, Dishaw LJ, et al: The amphioxus genome illuminates vertebrate origins and cephalochordate biology. Genome Res. 2008, 18 (7): 1100-1111. 10.1101/gr.073676.107.PubMed CentralView ArticlePubMedGoogle Scholar
- Kortschak RD, Samuel G, Saint R, Miller DJ: EST analysis of the cnidarian Acropora millepora reveals extensive gene loss and rapid sequence divergence in the model invertebrates. Curr Biol. 2003, 13 (24): 2190-2195. 10.1016/j.cub.2003.11.030.View ArticlePubMedGoogle Scholar
- Technau U, Rudd S, Maxwell P, Gordon PM, Saina M, Grasso LC, Hayward DC, Sensen CW, Saint R, Holstein TW, et al: Maintenance of ancestral complexity and non-metazoan genes in two basal cnidarians. Trends Genet. 2005, 21 (12): 633-639. 10.1016/j.tig.2005.09.007.View ArticlePubMedGoogle Scholar
- Raible F, Tessmar-Raible K, Osoegawa K, Wincker P, Jubin C, Balavoine G, Ferrier D, Benes V, de Jong P, Weissenbach J, et al: Vertebrate-type intron-rich genes in the marine annelid Platynereis dumerilii. Science. 2005, 310 (5752): 1325-1326. 10.1126/science.1119089.View ArticlePubMedGoogle Scholar
- Miller DJ, Ball EE: Cryptic complexity captured: the Nematostella genome reveals its secrets. Trends Genet. 2008, 24 (1): 1-4. 10.1016/j.tig.2007.10.002.View ArticlePubMedGoogle Scholar
- Zhou G, Denu JM, Wu L, Dixon JE: The catalytic role of Cys124 in the dual specificity phosphatase VHR. J Biol Chem. 1994, 269 (45): 28084-28090.PubMedGoogle Scholar
- Denu JM, Stuckey JA, Saper MA, Dixon JE: Form and Function in Protein Dephosphorylation. Cell. 1996, 87 (3): 361-10.1016/S0092-8674(00)81356-2.View ArticlePubMedGoogle Scholar
- Guo S, Stolz LE, Lemrow SM, York JD: SAC1-like domains of yeast SAC1, INP52, and INP53 and of human synaptojanin encode polyphosphoinositide phosphatases. J Biol Chem. 1999, 274 (19): 12990-12995. 10.1074/jbc.274.19.12990.View ArticlePubMedGoogle Scholar
- Hughes WE, Cooke FT, Parker PJ: Sac phosphatase domain proteins. Biochem J. 2000, 350 (Pt 2): 337-352. 10.1042/0264-6021:3500337.PubMed CentralView ArticlePubMedGoogle Scholar
- Maehama T, Dixon JE: The Tumor Suppressor, PTEN/MMAC1, Dephosphorylates the Lipid Second Messenger, Phosphatidylinositol 3,4,5-Trisphosphate. J Biol Chem. 1998, 273 (22): 13375-13378. 10.1074/jbc.273.22.13375.View ArticlePubMedGoogle Scholar
- Taylor GS, Maehama T, Dixon JE: Inaugural article: myotubularin, a protein tyrosine phosphatase mutated in myotubular myopathy, dephosphorylates the lipid second messenger, phosphatidylinositol 3-phosphate. Proc Natl Acad Sci USA. 2000, 97 (16): 8910-8915. 10.1073/pnas.160255697.PubMed CentralView ArticlePubMedGoogle Scholar
- Robinson FL, Dixon JE: Myotubularin phosphatases: policing 3-phosphoinositides. Trends Cell Biol. 2006, 16 (8): 403-412. 10.1016/j.tcb.2006.06.001.View ArticlePubMedGoogle Scholar
- Blennow A, Nielsen TH, Baunsgaard L, Mikkelsen R, Engelsen SB: Starch phosphorylation: a new front line in starch research. Trends Plant Sci. 2002, 7 (10): 445-450. 10.1016/S1360-1385(02)02332-4.View ArticlePubMedGoogle Scholar
- Yu TS, Kofler H, Hausler RE, Hille D, Flugge UI, Zeeman SC, Smith AM, Kossmann J, Lloyd J, Ritte G, et al: The Arabidopsis sex1 mutant is defective in the R1 protein, a general regulator of starch degradation in plants, and not in the chloroplast hexose transporter. Plant Cell. 2001, 13 (8): 1907-1918. 10.1105/tpc.13.8.1907.PubMed CentralView ArticlePubMedGoogle Scholar
- Csuros M, Rogozin IB, Koonin EV: Extremely intron-rich genes in the alveolate ancestors inferred with a flexible maximum-likelihood approach. Mol Biol Evol. 2008, 25 (5): 903-911. 10.1093/molbev/msn039.View ArticlePubMedGoogle Scholar
- Lohi H, Young EJ, Fitzmaurice SN, Rusbridge C, Chan EM, Vervoort M, Turnbull J, Zhao XC, Ianzano L, Paterson AD, et al: Expanded repeat in canine epilepsy. Science. 2005, 307 (5706): 81-10.1126/science.1102832.View ArticlePubMedGoogle Scholar
- Simmons MM: Lafora disease in the cow?. J Comp Pathol. 1994, 110 (4): 389-401. 10.1016/S0021-9975(08)80316-7.View ArticlePubMedGoogle Scholar
- Loupal G: [A storage disease in a parakeet (Nymphicus hollandicus) morphologically similar to the systemic myoclonic disease (Lafora disease) in man and dog]. Zentralbl Veterinarmed A. 1985, 32 (7): 502-511.PubMedGoogle Scholar
- Suzuki Y, Kamiya S, Ohta K, Suu S: Lafora-like bodies in a cat. Case report suggestive of glycogen metabolism disturbances. Acta Neuropathol. 1979, 48 (1): 55-58. 10.1007/BF00691791.View ArticlePubMedGoogle Scholar
- Cavanagh JB: Corpora-amylacea and the family of polyglucosan diseases. Brain Res Brain Res Rev. 1999, 29 (2–3): 265-295. 10.1016/S0165-0173(99)00003-X.View ArticlePubMedGoogle Scholar
- Worby CA, Gentry MS, Dixon JE: Malin decreases glycogen accumulation by promoting the degradation of protein targeting to glycogen (PTG). J Biol Chem. 2008, 283 (7): 4069-4076. 10.1074/jbc.M708712200.PubMed CentralView ArticlePubMedGoogle Scholar
- Yu JK, Wang MC, Shin IT, Kohara Y, Holland LZ, Satoh N, Satou Y: A cDNA resource for the cephalochordate amphioxus Branchiostoma floridae. Dev Genes Evol. 2008Google Scholar
- Taylor GS, Liu Y, Baskerville C, Charbonneau H: The activity of Cdc14p, an oligomeric dual specificity protein phosphatase from Saccharomyces cerevisiae, is required for cell cycle progression. J Biol Chem. 1997, 272 (38): 24054-24063. 10.1074/jbc.272.38.24054.View ArticlePubMedGoogle Scholar
- Gentry MS, Worby CA, Dixon JE: Insights into Lafora disease: malin is an E3 ubiquitin ligase that ubiquitinates and promotes the degradation of laforin. Proc Natl Acad Sci USA. 2005, 102 (24): 8501-8506. 10.1073/pnas.0503285102.PubMed CentralView ArticlePubMedGoogle Scholar
- Harder KW, Owen P, Wong LK, Aebersold R, Clark-Lewis I, Jirik FR: Characterization and kinetic analysis of the intracellular domain of human protein tyrosine phosphatase beta (HPTP beta) using synthetic phosphopeptides. Biochemical Journal. 1994, 298: 395-401.PubMed CentralView ArticlePubMedGoogle Scholar
- Altschul SF, Madden TL, Schaffer AA, Zhang J, Zhang Z, Miller W, Lipman DJ: Gapped BLAST and PSI-BLAST: a new generation of protein database search programs. Nucleic Acids Res. 1997, 25 (17): 3389-3402. 10.1093/nar/25.17.3389.PubMed CentralView ArticlePubMedGoogle Scholar
- Thompson JD, Higgins DG, Gibson TJ: CLUSTAL W: improving the sensitivity of progressive multiple sequence alignment through sequence weighting, position-specific gap penalties and weight matrix choice. Nucl Acids Res. 1994, 22 (22): 4673-4680. 10.1093/nar/22.22.4673.PubMed CentralView ArticlePubMedGoogle Scholar
- Bingham J, Sudarsanam S: Visualizing large hierarchical clusters in hyperbolic space. Bioinformatics. 2000, 16 (7): 660-661. 10.1093/bioinformatics/16.7.660.View ArticlePubMedGoogle Scholar
- Adl SM, AG Simpson, MA Farmer, RA Andersen, OR Anderson, JR Barta, SS Bowser, G Brugerolle, RA Fensome, et al: The new higher level classification of eukaryotes with emphasis on the taxonomy of protists. The Journal of Eukaryotic Microbiology. 2005, 52: 399-451. 10.1111/j.1550-7408.2005.00053.x.View ArticlePubMedGoogle Scholar
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