Signal transduction of chondroitin sulfate proteoglycans and neuronal network formation

PtPζ是一种受体型蛋白酪氨酸磷酸酶，它以肝素结合生长因子多营养素为配体，在小脑浦肯野细胞的形态发生中发挥重要作用。在这项研究中，我们研究了PtPζ和DNER之间的功能相互作用，DNER是浦肯野细胞中高表达的Delta/Notch样受体。PtP、ζ和DNER在浦肯野细胞的树突状突起和表达这两种蛋白的COS7细胞中呈片状共定位。免疫沉淀实验表明，在体内和体外，ζ与DNER形成了络合物。DNER胞内区的几个酪氨酸残基，包括基于酪氨酸的分选基序及其附近的酪氨酸残基被磷酸化，并很容易被ptpζ催化结构域去磷酸化。酪氨酸分选基序在dNER的内吞和胞内转运中起关键作用，酪氨酸磷酸化和破坏该基序导致dNER在pTPζ表达细胞的质膜上积聚。DNER在Neuro-2A细胞质膜上的聚集导致包括突起延长在内的形态变化。Pleiotroin抑制DNER的内吞作用，并抑制DNER诱导的Neuro-2A细胞突起生长。这些观察表明，多营养素-PTPζ信号通过改变细胞内DNER的运输来调节浦肯野细胞的形态发生。

PTPζ, a receptor-type protein tyrosine phosphatase, uses a heparin-binding growth factor pleiotrophin as a ligand and play important roles in the morphogenesis of cerebellar Purkinje cells. In this study, we studied functional interaction between PTPζ and DNER, a Delta/Notch-like receptor highly expressed in Purkinje cells. PTPζ and DNER displayed patchy co-localization in the dendritic shafts of Purkinje cells and in the COS-7 transfectants expressing both proteins. Immunoprecipitation experiments indicated that PTPζ and DNER formed complex both in vivo and in vitro. Several tyrosine residues in the intracellular region of DNER including the ones in and adjacent to the tyrosine-based sorting motif were phosphorylated, and were easily dephosphorylated by the PTPζ catalytic domain. The tyrosine-based sorting motif played critical roles in the endocytosis and intracellular transport of DNER, and tyrosine phosphorylation and destruction of this motif resulted in the accumulation of DNER on the plasma membrane of the PTPζ-expressing cells. Accumulation of DNER on the plasma membrane of Neuro-2A cells led to the morphological changes including extension of processes. Pleiotrophin suppressed the endocytosis of DNER and DNER-induced inhibition of neurite outgrowth of Neuro-2A cells. These observations suggests that pleiotrophin-PTPζ signaling regulates the morphogenesis of Purkinje cells by modifying the intracellular transport of DNER.