Lafora disease proteins malin and laforin are recruited to aggresomes in response to proteasomal impairment

Department of Biological Sciences and Bioengineering, Indian Institute of Technology, Kanpur, 208016, India.
Human Molecular Genetics (Impact Factor: 6.39). 05/2007; 16(7):753-62. DOI: 10.1093/hmg/ddm006
Source: PubMed


Lafora disease (LD), an autosomal recessive neurodegenerative disorder, is characterized by the presence of cytoplasmic polyglucosan inclusions known as Lafora bodies in several tissues including the brain. Laforin, a protein phosphatase, and malin, an ubiquitin ligase, are two of the proteins that are known to be defective in LD. Malin interacts with laforin and promotes its polyubiquitination and degradation. Here we show that malin and laforin co-localize in endoplasmic reticulum (ER) and that they form centrosomal aggregates when treated with proteasomal inhibitors in both neuronal and non-neuronal cells. Laforin/malin aggregates co-localize with gamma-tubulin and cause redistribution of alpha-tubulin. These aggregates are also immunoreactive to ubiquitin, ubiquitin-conjugating enzyme, ER chaperone and proteasome subunits, demonstrating their aggresome-like properties. Furthermore, we show that the centrosomal aggregation of laforin and malin is dependent on the functional microtubule network. Laforin and malin form aggresome when expressed together or otherwise, suggesting that the two proteins are recruited to the centrosome independent of each other. Taken together, our results suggest that the centrosomal accumulation of malin, possibly with the help of laforin, may enhance the ubiquitination of its substrates and facilitate their efficient degradation by proteasome. Defects in malin or laforin may thus lead to increased levels of misfolded and/or target proteins, which may eventually affect the physiological processes of the neuron. Thus, defects in protein degradation and clearance are likely to be the primary trigger in the physiopathology of LD.


Available from: Shuchi Mittal, Jan 22, 2015
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    • "We and others have previously reported a role for Lafora disease proteins laforin and malin in cellular stress response [9] [29] [30], proteolytic processes [7] [8] [31], glucose metabolism [32e34] and RNA metabolism [35]. In this report, we attempt to propose a link for the LD proteins in the HIPK2-p53-mediated cell death pathway. "
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    ABSTRACT: Lafora disease (LD) is an autosomal recessive, progressive, and fatal form of a neurodegenerative disorder characterized by the presence of Lafora polyglucosan bodies. LD is caused by defects in either the laforin protein phosphatase or the malin E3 ubiquitin ligase. Laforin and malin were shown play key roles in proteolytic processes, unfolded stress response, and glycogen metabolism. Therefore, the LD proteins laforin and malin are thought to function as pro-survival factors and their loss thus could result in neurodegeneration. To understand the molecular pathway leading to the cell death in LD, in the present study, we investigated the possible role of LD proteins in the p53-mediated cell death pathway. We show that loss of laforin or malin results in the increased level and activity of p53, both in cellular and animal models of LD, and that this is primarily due to the increased levels of Hipk2, a proapoptotic activator of p53. Overexpression of laforin or malin confers protection against Hipk2-mediated cell death by targeting the Hipk2 to the cytoplasmic compartment. Taken together, our study strengthens the notion that laforin and malin are pro-survival factors, and that the activation of Hipk2-p53 cell death pathway might underlie neurodegeneration in LD. Copyright © 2015. Published by Elsevier Inc.
    Biochemical and Biophysical Research Communications 06/2015; 464(1). DOI:10.1016/j.bbrc.2015.06.018 · 2.30 Impact Factor
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    • "It should be recalled that large deposits of glycogen particles have long been reported in some cells by TEM, with special reference to clear-cell neoplasms [67], although UPS components were never tested in such structures. The presence of glycogen and glycogen synthase in PaCSs may be explained by several recently discovered relations between the UPS and glycogen-related proteins: (1) the UPS regulation of proteins such as laforin, acting in complex with E3 ligase malin [68], the protein targeting glycogen [69] and AMP kinase [70]; (2) direct involvement of the laforin–malin complex in misfolded protein degradation [71]; and (3) the recruitment of this complex in aggresome-like cytoplasmic structures, together with ubiquitin and glycogen [72]. In addition, a role has been proposed for Rab25/AKT-activated glycogen stores in promoting cancer cell survival through aerobic-glycolysis-mediated ATP synthesis, a crucial requirement for enhanced UPS function [73], [74]. "
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    ABSTRACT: A variety of ubiquitinated protein-containing cytoplasmic structures has been reported, from aggresomes to aggresome-like induced structures/sequestosomes or particle-rich cytoplasmic structures (PaCSs) that we recently observed in some human diseases. Nevertheless, the morphological and cytochemical patterns of the different structures remain largely unknown thus jeopardizing their univocal identification. Here, we show that PaCSs resulted from proteasome and polyubiquitinated protein accumulation into well-demarcated, membrane-free, cytoskeleton-poor areas enriched in glycogen and glycosaminoglycans. A major requirement for PaCS detection by either electron or confocal microscopy was the addition of osmium to aldehyde fixatives. However, by analyzing living cells, we found that proteasome chymotrypsin-like activity concentrated in well-defined cytoplasmic structures identified as PaCSs by ultrastructural morphology and immunocytochemistry of the same cells. PaCSs differed ultrastructurally and cytochemically from sequestosomes which may coexist with PaCSs. In human dendritic or natural killer cells, PaCSs were induced in vitro by cytokines/trophic factors during differentiation/activation from blood progenitors. Our results provide evidence that PaCS is indeed a novel distinctive cytoplasmic structure which may play a critical role in the ubiquitin-proteasome system response to immune, infectious or proneoplastic stimuli.
    PLoS ONE 12/2013; 8(12):e82560. DOI:10.1371/journal.pone.0082560 · 3.23 Impact Factor
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    • "The implication of laforin and malin in glycogen metabolism - either by means of a direct glycogen dephosphorylation executed by laforin, by the ubiquitination of glycogenic regulators through the laforin-malin complex, or by a balanced participation of both processes - has guided research in LD over the last several years. Additionally, recent work has described novel relations between laforin, malin and other proteins involved in protein clearance systems [5], [26]–[28]. Moreover, the effects of laforin-malin abnormalities produce autophagy impairment and ER stress in animal models [29]–[32]. "
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    ABSTRACT: Laforin, encoded by a gene that is mutated in Lafora Disease (LD, OMIM 254780), is a modular protein composed of a carbohydrate-binding module and a dual-specificity phosphatase domain. Laforin is the founding member of the glucan-phosphatase family and regulates the levels of phosphate present in glycogen. Multiple reports have described the capability of laforin to form dimers, although the function of these dimers and their relationship with LD remains unclear. Recent evidence suggests that laforin dimerization depends on redox conditions, suggesting that disulfide bonds are involved in laforin dimerization. Using site-directed mutagenesis we constructed laforin mutants in which individual cysteine residues were replaced by serine and then tested the ability of each protein to dimerize using recombinant protein as well as a mammalian cell culture assay. Laforin-Cys329Ser was the only Cys/Ser mutant unable to form dimers in both assays. We also generated a laforin truncation lacking the last three amino acids, laforin-Cys329X, and this truncation also failed to dimerize. Interestingly, laforin-Cys329Ser and laforin-Cys329X were able to bind glucans, and maintained wild type phosphatase activity against both exogenous and biologically relevant substrates. Furthermore, laforin-Cys329Ser was fully capable of participating in the ubiquitination process driven by a laforin-malin complex. These results suggest that dimerization is not required for laforin phosphatase activity, glucan binding, or for the formation of a functional laforin-malin complex. Cumulatively, these results suggest that cysteine 329 is specifically involved in the dimerization process of laforin. Therefore, the C329S mutant constitutes a valuable tool to analyze the physiological implications of laforin's oligomerization.
    PLoS ONE 07/2013; 8(7):e69523. DOI:10.1371/journal.pone.0069523 · 3.23 Impact Factor
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