Lafora disease (LD) is an autosomal recessive myoclonus epilepsy with onset in the teenage years leading to death within a decade of onset. LD is characterized by the overaccumulation of hyperphosphorylated, poorly branched, insoluble, glycogen-like polymers called Lafora bodies. The disease is caused by mutations in either , encoding laforin, a dual specificity phosphatase that dephosphorylates glycogen, or , encoding malin, an E3-ubiquitin ligase. While glycogen is a widely accepted laforin substrate, substrates for malin have been difficult to identify partly due to the lack of malin antibodies able to detect malin . Here we describe a mouse model in which the malin gene is modified at the C-terminus to contain the c-myc tag sequence, making an expression of malin-myc readily detectable. Mass spectrometry analyses of immunoprecipitates using c-myc tag antibodies demonstrate that malin interacts with laforin and several glycogen-metabolizing enzymes. To investigate the role of laforin in these interactions we analyzed two additional mouse models: malin-myc/laforin knockout and malin-myc/LaforinCS, where laforin was either absent or the catalytic Cys was genomically mutated to Ser, respectively. The interaction of malin with partner proteins requires laforin but is not dependent on its catalytic activity or the presence of glycogen. Overall, the results demonstrate that laforin and malin form a complex , which stabilizes malin and enhances interaction with partner proteins to facilitate normal glycogen metabolism. They also provide insights into the development of LD and the rescue of the disease by the catalytically inactive phosphatase.

中文翻译：

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拉福拉病中马林表达及其与伴侣蛋白的相互作用受损

拉福拉病（LD）是一种常染色体隐性遗传性肌阵挛癫痫，在青少年时期发病，导致发病后十年内死亡。 LD 的特点是过度积累过度磷酸化、支链不良、不溶性糖原样聚合物（称为拉福拉体）。该疾病是由编码laforin（一种使糖原去磷酸化的双特异性磷酸酶）或编码malin（一种E3-泛素连接酶）的突变引起的。虽然糖原是一种广泛接受的 Laforin 底物，但 Malin 的底物很难鉴定，部分原因是缺乏能够检测 Malin 的 Malin 抗体。在这里，我们描述了一种小鼠模型，其中 malin 基因在 C 末端被修饰以包含 c-myc 标签序列，使得 malin-myc 的表达易于检测。使用 c-myc 标签抗体对免疫沉淀物进行质谱分析表明，malin 与 laforin 和几种糖原代谢酶相互作用。为了研究laforin在这些相互作用中的作用，我们分析了另外两种小鼠模型：malin-myc/laforin敲除模型和malin-myc/LaforinCS模型，其中laforin要么不存在，要么催化Cys基因组突变为Ser。马林与伴侣蛋白的相互作用需要拉福林，但不依赖于其催化活性或糖原的存在。总体而言，结果表明 laforin 和 malin 形成复合物，可稳定 malin 并增强与伴侣蛋白的相互作用，以促进正常的糖原代谢。他们还为 LD 的发展和通过催化失活磷酸酶拯救该疾病提供了见解。