Given the role of erbB family receptor tyrosine kinases (RTKs) in human tumors, it is of critical importance to understand the regulation of erbB signaling in normal and transformed cells. Our earlier work has characterized biochemical features of erbB receptor activation and attenuation in transformed astrocytes. The proposed set of studies will examine the roles of the SH2 domain-containing SHP2 protein-tyrosine phosphatase (PTP) and the immunoreceptor tyrosine-based inhibitory motif (ITIM)-containing SIRP (Signal-Regulatory Protein) inhibitory receptor in regulating erbB signaling in normal and transformed astrocytes. The planned experiments are part of a continuing effort to understand features of erbB signal attenuation in glial cells of the central nervous system. While much is known regarding signaling induced by RTKs, the physiologic role of PTPs is poorly understood, particularly how PTPs may regulate RTK signals. RTKs, particularly of the erbB family, are essential to the pathogenesis of human astrocytic tumors. Importance of the phosphoinositide-3-OH kinase (P13-K) signaling pathway in astrocytic tumors is illustrated by studies of Epidermal Growth Factor Receptor (EGFR) oncoproteins and the PTEN phosphatase. The focus of this proposal will be on regulation of early receptor-based events following EGFR stimulation that modulate P13-K enzymatic activation and P13-K-mediated phenotypes in normal and transformed astrocytes. Our laboratory has identified a novel mechanism of negative regulation of P13-K activation in astrocytoma cells involving SHP2 and SIRP. SHP2 associates with the SIRP inhibitory receptor and a SHP2/SIRP complex inhibits erbB-induced P13-K signaling in astrocytes. SHP2 also associates with the p85 regulatory subunit of P13-K in cells in which P13-K pathway activation is unimpaired. We have shown that tyrosine phosphorylation of the SIRP receptor and recruitment of SHP2 inhibits the SHP2/p85 association. Moreover, our data show that SHP2/SIRP complex formation inhibits cell motility, survival and transformation of erbB-containing transformed astrocytes. Given its role in negative regulation of the P13-K pathway, the PTEN phosphatase may also regulate these phenotypes with SHP2 and SIRP. The proposed experiments will examine the role of the SHP2 PTP in astrocytes since this protein mediates signaling from both erbB and SIRP receptors and may therefore integrate positive and negative cellular signals. We hope to learn how erbB and SIRP signals regulate SHP2 protein associations and PIP activity and how these events regulate the P13-K pathway. We believe the planned studies in astrocyte biology will characterize biochemically a mechanism of negative regulation of erbB-induced, P13-K-mediated cell survival and motility pathways that influence astrocyte transformation.
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