The role of RPTPzeta in models of Congenital Muscular Dystrophies

描述(申请人提供)：与O-甘露醇糖基化有关的酶的突变会产生一组毁灭性的疾病，其特征是先天性肌营养不良(CMD)、II型无脑畸形和眼睛异常。以前使用动物模型的研究已经证明，α-营养不良糖链的低糖基化在这些疾病的发病机制中起着关键作用。然而，越来越多的证据表明，O-甘露醇糖基化的其他底物可能导致在动物模型和CMDs患者中发现的特定大脑异常。不幸的是，以前的研究未能确定O-甘露糖基化的额外神经底物，这些底物有助于这些大脑病理。利用糖基转移酶POMGNT1被敲除的CMD动物模型，我们发现了RPTP？作为大脑中O-甘露醇糖基化的另一种重要底物。受体酪氨酸磷酸酶Zeta/beta是一种主要在中枢神经系统中发现的受体磷酸酶，在神经发育的关键阶段高度表达。该蛋白的膜结合受体和分泌变体都在大脑中表达，是许多发育重要的黏附分子、生长因子和细胞外基质蛋白的高亲和力配体。我们假设RPTP6的低糖基化是由于O-甘露糖化被干扰而改变了RPTP？与这些关键的配体有关，并导致大脑发育异常。与这一假设一致，我们发现POMGNT1基因敲除后培养细胞的皮质神经元发育异常，当它们被电镀在已知RPTP？配基。在这个提议中，我们研究了RPTP？在O-甘露醇中，糖基化突变的动物改变了其配体结合特性，从而导致异常的细胞相互作用和异常的脑发育。该提案集中于以下3个具体目标：1)确定RPTP的糖基化是否改变？调节其配体结合特性。2)确定是否干扰了RPTP的糖基化？改变细胞发育。3)鉴定CMDs动物模型中1-营养不良糖非依赖性神经异常，并确定这些异常是否是由于RPTP？/磷酸多糖糖基化改变所致。为了探索这些问题，我们将利用各种遗传小鼠模型系统，在这些系统中，我们将扰乱O-甘露糖糖基化、α-葡聚糖表达或RPTP？表达。将在体外和体内进行研究，以确定O-甘露糖聚糖如何调节RPTP？功能，以及这是否导致在CMDS动物模型中发现的大脑异常。这些研究将确定RPTP是否？是CMD发病的重要因素，并将为这些疾病的神经表型的病理机制提供重要的见解。

DESCRIPTION (provided by applicant): Mutations in enzymes involved in protein O-mannosyl glycosylation produce a group of devastating diseases characterized by congenital muscular dystrophy (CMD), type II lissencephaly, and eye abnormalities. Previous studies using animal models have demonstrated that hypoglycosylation of a-dystroglycan critically contributes to the pathogenesis of these disorders. A growing body of evidence, however, suggests that other substrates for O-mannosyl glycosylation likely contribute to particular brain abnormalities found both in animal models and in patients with CMDs. Unfortunately, previous studies have failed to identify the additional neural substrates of O-mannosylation that contribute to these brain pathologies. Using an animal model of CMD in which the glycosyltransferase POMGnT1 is knocked out we identified RPTP? as another important substrate for O-mannosyl glycosylation in the brain. Receptor tyrosine phosphatase zeta/beta is a receptor phosphatase that is found predominately in the central nervous system and is highly expressed during key stages of neural development. Both membrane-bound receptor and secreted variants of the protein are expressed in the brain and are high affinity ligands for a number of developmentally important adhesion molecules, growth factors and extracellular matrix proteins. We hypothesize that the hypoglycosylation of RPTP6 due to disrupted O-mannosylation alters the interactions of RPTP? with these key ligands and contributes to abnormal brain development. Consistent with this hypothesis we found that cortical neuron development in cultured cells from POMGnT1 knockouts is abnormal when they are plated on a subset of known RPTP? ligands. In this proposal we investigate the hypothesis that the hypoglycosylaton of RPTP? in O-mannosyl glycosylation mutant animals alters its ligand-binding characteristics thereby leading to aberrant cellular interactions and abnormal brain development. The proposal is focused on the following 3 specific aims: 1) To determine whether altered glycosylation of RPTP? modulates its ligand binding characteristics. 2) To determine whether disrupted glycosylation of RPTP? alter cell development. And 3) To identify the 1-dystroglycan-independent neural abnormalities in animal models of CMDs and to determine if these abnormalities are due to altered RPTP?/phosphacan glycosylation. To explore these questions, we will utilize a variety of genetic mouse model systems in which we will disrupt O-mannosyl glycosylation, a-dystrglycan expression or RPTP?expression. Studies will be conducted both in vitro and in vivo to determine how O-mannosyl glycans modulate RPTP? function and if this contributes to brain abnormalities found in animal models of CMDs. These studies will determine if RPTP? is an important contributor to CMD pathogenesis and will provide important insights into the pathomechanisms of the neural phenotypes in these disorders.