When activated, the platelet-derived growth factor (PDGF) receptor phosphorylates itself on multiple tyrosines. These phosphorylated tyrosines serve as assembly sites for several SH2-domain-containing proteins that include phospholipase C-γ (PLCγ), a Ras-specific GTPase-activating protein (GAP), a subunit of phosphoinositide 3-kinase (PI3K), and a phosphotyrosine phosphatase (PTP) (Panel A). PDGF binding stimulates several changes in the target cell, one of which is an increase in DNA synthesis, as measured by incorporation of radioactive thymidine or bromodeoxyuridine into DNA.
To determine which of the bound proteins is responsible for activation of DNA synthesis, you construct several mutant genes for the PDGF receptor that retain individual or combinations of tyrosine phosphorylation sites. When expressed in cells that do not make a PDGF receptor of their own, each of the receptors is phosphorylated at its tyrosines upon binding of PDGF. As shown in Panel B, DNA synthesis is stimulated to different extents in cells expressing the mutant receptors.
A. From these data, which, if any, of PI3K, GAP, PTP, and PLCγ are involved in the stimulation of DNA synthesis by PDGF (2 points, state the protein(s))? Explain your answer with respect to experiment(s) that led to conclusion (3 points, 2-3 sentences).