MULTIPLE REQUIREMENTS FOR SHPTP2 IN EPIDERMAL GROWTH FACTOR-MEDIATED CELL-CYCLE PROGRESSION

Using transient overexpression and microinjection approaches, we exami ned SHPTP2's function in growth factor signaling. Overexpression of ca talytically inactive SHPTP2 (PTP2CS) but not catalytically inactive SH PTP1, inhibited mitogen-activated protein (MAP) kinase activation and Elk-1 transactivation following epidermal growth factor (EGF) stimulat ion of 293 cells. An SHPTP2 mutant with both C-terminal tyrosyl phosph orylation sites converted to phenylalanine (PTP2YF) was also without e ffect; moreover, PTP2YF rescued PTP2CS-induced inhibition of EGF-induc ed Elk-1 transactivation. PTP2CS did not inhibit transactivation by ac tivated Ras, suggesting that SHPTP2 acts upstream of or parallel to Pa s. Neither PTP2CS nor PTP2YF inhibited platelet-derived growth factor (PDGF)-induced Elk-1 transactivation. Thus, protein tyrosine phosphata se activity, but not tyrosyl phosphorylation of SHPTP2, is required fo r the immediate-early responses to EGF but not to PDGF. To determine w hether SHPTP2 is required later in the cell cycle, we assessed S-phase entry in NIH 3T3 cells microinjected with anti-SHPTP2 antibodies or w ith a glutathione S-transferase (GST) fusion protein encoding both SH2 domains (GST-SH2). Microinjection of anti-SHPTP2 antibodies prior to stimulation inhibited EGF- but not PDGF- or serum-induced S phase entr y. Anti-SHPTP2 antibodies or GST-SH2 fusion protein could inhibit EGF- induced S-phase entry for up to 8 h after EGF addition. Although MAP k inase activation was detected shortly after EGF stimulation, no MAP ki nase activation was detected around the restriction point. Therefore, SHPTP2 is absolutely required for immediately and late events induced by some, but not all, growth factors, and the immediate-early and late signal transduction pathways regulated by SHPTP2 are distinguishable.