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Lafora disease (LD) is a fatal childhood dementia characterized by progressive myoclonic epilepsy manifesting in the teenage years, rapid neurological decline, and death typically within ten years of onset. Mutations in either EPM2A, encoding the glycogen phosphatase laforin, or EPM2B , encoding the E3 ligase malin, cause LD. Whole exome sequencing has revealed many EPM2A variants associated with late-onset or slower disease progression. We established an empirical pipeline for characterizing the functional consequences of laforin missense mutations in vitro using complementary biochemical approaches. Analysis of 26 mutations revealed distinct functional classes associated with different outcomes that were supported by clinical cases. For example, F321C and G279C mutations have attenuated functional defects and are associated with slow progression. This pipeline enabled rapid characterization and classification of newly identified EPM2A mutations, providing clinicians and researchers genetic information to guide treatment of LD patients., Competing Interests: M.S.G. is a consultant for Maze Therapeutics, Enable Therapeutics, Glut1 Deficiency Syndrome Foundation, and Chelsea’s Hope. M.S.G. and C.V.K. are founders of Atterogen, LLC., (© 2021 The Authors.)

Glycosylation defects are a hallmark of many nervous system diseases. However, the molecular and metabolic basis for this pathology is not fully understood. In this study, we found that N-linked protein glycosylation in the brain is metabolically channeled to glucosamine metabolism through glycogenolysis. We discovered that glucosamine is an abundant constituent of brain glycogen, which functions as a glucosamine reservoir for multiple glycoconjugates. We demonstrated the enzymatic incorporation of glucosamine into glycogen by glycogen synthase, and the release by glycogen phosphorylase by biochemical and structural methodologies, in primary astrocytes, and in vivo by isotopic tracing and mass spectrometry. Using two mouse models of glycogen storage diseases, we showed that disruption of brain glycogen metabolism causes global decreases in free pools of UDP-N-acetylglucosamine and N-linked protein glycosylation. These findings revealed fundamental biological roles of brain glycogen in protein glycosylation with direct relevance to multiple human diseases of the central nervous system., Competing Interests: Declaration of interests M.S.G. is a consultant for Maze Therapeutics, Enable Therapeutics, Glut1-Deficiency Syndrome Foundation, and Chelsea's Hope. M.S.G., R.C.S., C.W.V.K., and R.C.B. are founders of Atterogen, LLC., (Copyright © 2021 Elsevier Inc. All rights reserved.)

Nuclear glycogen was first documented in the early 1940s, but its role in cellular physiology remained elusive. In this study, we utilized pure nuclei preparations and stable isotope tracers to define the origin and metabolic fate of nuclear glycogen. Herein, we describe a key function for nuclear glycogen in epigenetic regulation through compartmentalized pyruvate production and histone acetylation. This pathway is altered in human non-small cell lung cancers, as surgical specimens accumulate glycogen in the nucleus. We demonstrate that the decreased abundance of malin, an E3 ubiquitin ligase, impaired nuclear glycogenolysis by preventing the nuclear translocation of glycogen phosphorylase and causing nuclear glycogen accumulation. Re-introduction of malin in lung cancer cells restored nuclear glycogenolysis, increased histone acetylation, and decreased growth of cancer cells transplanted into mice. This study uncovers a previously unknown role for glycogen metabolism in the nucleus and elucidates another mechanism by which cellular metabolites control epigenetic regulation., (Copyright © 2019 Elsevier Inc. All rights reserved.)

Lafora disease (LD) is a fatal childhood epilepsy caused by recessive mutations in either the EPM2A or EPM2B gene. A hallmark of LD is the intracellular accumulation of insoluble polysaccharide deposits known as Lafora bodies (LBs) in the brain and other tissues. In LD mouse models, genetic reduction of glycogen synthesis eliminates LB formation and rescues the neurological phenotype. Therefore, LBs have become a therapeutic target for ameliorating LD. Herein, we demonstrate that human pancreatic α-amylase degrades LBs. We fused this amylase to a cell-penetrating antibody fragment, and this antibody-enzyme fusion (VAL-0417) degrades LBs in vitro and dramatically reduces LB loads in vivo in Epm2a -/- mice. Using metabolomics and multivariate analysis, we demonstrate that VAL-0417 treatment of Epm2a -/- mice reverses the metabolic phenotype to a wild-type profile. VAL-0417 is a promising drug for the treatment of LD and a putative precision therapy platform for intractable epilepsy., (Copyright © 2019 Elsevier Inc. All rights reserved.)

Glycogen is the major mammalian glucose storage cache and is critical for energy homeostasis. Glycogen synthesis in neurons must be tightly controlled due to neuronal sensitivity to perturbations in glycogen metabolism. Lafora disease (LD) is a fatal, congenital, neurodegenerative epilepsy. Mutations in the gene encoding the glycogen phosphatase laforin result in hyperphosphorylated glycogen that forms water-insoluble inclusions called Lafora bodies (LBs). LBs induce neuronal apoptosis and are the causative agent of LD. The mechanism of glycogen dephosphorylation by laforin and dysfunction in LD is unknown. We report the crystal structure of laforin bound to phosphoglucan product, revealing its unique integrated tertiary and quaternary structure. Structure-guided mutagenesis combined with biophysical and biochemical analyses reveal the basis for normal function of laforin in glycogen metabolism. Analyses of LD patient mutations define the mechanism by which subsets of mutations disrupt laforin function. These data provide fundamental insights connecting glycogen metabolism to neurodegenerative disease., (Copyright © 2015 Elsevier Inc. All rights reserved.)

Septins are GTPases involved in cytokinesis. In yeast, they form a ring at the cleavage site. Using FRAP, we show that septins are mobile within the ring at bud emergence and telophase and are immobile during S, G2, and M phases. Immobilization of the septins is dependent on both Cla4, a PAK-like kinase, and Gin4, a septin-dependent kinase that can phosphorylate the septin Shs1/Sep7. Induction of septin ring dynamics in telophase is triggered by the translocation of Rts1, a kinetochore-associated regulatory subunit of PP2A phosphatase, to the bud neck and correlates with Rts1-dependent dephosphorylation of Shs1. In rts1-Delta cells, the actomyosin ring contracts properly but cytokinesis fails. Together our results implicate septins in a late step of cytokinesis and indicate that proper regulation of septin dynamics, possibly through the control of their phosphorylation state, is required for the completion of cytokinesis.