Phospholipase C-γ (PLC-γ) is an important mediator of the mitogenic actions of several growth factor receptors; overexpression of the PLC-γ1 isoform induces DNA synthesis and tumorigenicity. Its phospholipase activity, though, isn't required for this — it is the Src-homology-3 (SH3) domain that is responsible, but the mechanism underlying this action has so far remained unsolved. Now, Snyder and his colleagues report that PLC-γ1 — through its SH3 domain — has physiological guanine nucleotide exchange factor (GEF) activity towards a nuclear GTPase that activates nuclear phosphatidylinositol 3′-kinase (PI3K) activity.

The nuclear GTPase in question is PI3K enhancer (PIKE), identified two years ago. PIKE has proline-rich domains, so pull-down assays were used to 'fish' for potential binding partners — proteins that contain SH3 domains. PLC-γ was found to interact strongly with the third proline-rich domain of PIKE, so the authors then tested whether PLC-γ affected PIKE's GEF activity. They showed that the SH3 domain of PLC-γ (residues 790–850) stimulated GTP loading and GDP dissociation, whereas a proline to leucine mutation at 842 abolished this. If PLC-γ does indeed have GEF activity then, like other GEFs, it should bind strongly and then dissociate from its target GTPase — in this case, PIKE — which indeed it did.

PIKE is a nuclear protein, so can PLC-γ be found in the nucleus, too? Cell fractionation studies and immunofluorescent staining confirmed that stimulation with nerve growth factor (NGF) increased the level of PLC-γ in the nucleus. So how does stimulation with NGF and subsequent nuclear translocation of PLC-γ regulate the activities of PIKE and PI3K? In cells transfected with wild-type PLC-γ and treated with NGF, PIKE activity and nuclear PI3K activity were markedly enhanced. Deletion of the SH2 or catalytic domains of PLC-γ had no effect on this, but deleting or mutating the SH3 domain blocked the activation of both proteins.

Next, the authors showed that the SH3 domain of PLC-γ is necessary for mitogenesis in response to growth factors. This is consistent with previous findings in which cells containing SH3-domain-deletion constructs failed to grow in the absence of serum. Together, these results provide a mechanism by which PLC-γ — through PIKE — can mediate mitogenic actions in the absence of its lipase activity, and further contribute to the possibility that the SH3 domain is not just a binding motif, but in fact has some enzymatic activity.