Phospholipase C – 1 : A Phospholipase and Guanine Nucleotide Exchange Factor

The phospholipase C (PLC) family of cellular proteins participates in the intracellular signaling mechanisms utilized by diverse growth factors (GFs). A wide variety of growth factor receptor tyrosine kinases (RTKs) phosphorylate and activate PLC– 1; nonetheless, the biological role and necessity of this signal transduction element in mitogenesis has remained unclear. Indeed, whereas recent results point to a more important role of PLC– 1 in GF-induced mitogenesis than was suggested by initial studies (1), the molecular mechanism by which PLC– 1 mediates GFinduced mitogenesis remains elusive. Recently, Snyder and coworkers have reported that the Src homology 3 (SH3) domain of PLC– 1 acts as a guanine nucleotide exchange factor (GEF) for PIKE [phosphatidylinositol-3' kinase (PI3K) enhancer] (2). PIKE is a nucleus-localized small guanosine triphosphatase (GTPase) that stimulates PI3K activity in the nucleus and mediates the physiological activation by nerve growth factor (NGF) of nuclear PI3K activity (3). This enzymatic activity accounts for the mitogenic properties of PLC– 1. Moreover, these results show that the mitogenic activity of PLC– 1 is entirely due to its SH3 domain rather than its phospholipase activity and its SH2 domains. These results challenge the prevailing wisdom and mechanism by which PLC– 1 was believed to mediate GFinduced mitogenesis and reintroduce the concept that SH3 domains can display enzymatic activity in addition to acting as a binding motif. PLC– 1 contains two SH2, one SH3, one pleckstrin homology (PH), and two catalytic domains. All of these domains are important for the function of PLC– 1 and the regulation of GF-induced mitogenesis (Figure 1). Through its SH2 domain, PLC– 1 can form complexes with PDGF receptors (PDGFRs) or EGF receptors (EGFRs) in vivo (4–8). The formation of these protein complexes leads to phosphorylation of PLC– 1 on tyrosine residues and an increase in its enzymatic activity (9–11). The SH3 domain of PLC– 1 has been implicated in targeting PLC– 1 to the cytoskeleton (12). The SH3 domain of PLC– 1 can also bind the GTPase dynamin in vitro and in vivo (13, 14) and to proline-rich peptides derived from the Ras GEF Son-of-sevenless (Sos) in vitro (13). However, the physiological relevance of these interactions has not been determined. The PH domain of PLC– 1 reportedly binds to phosphatidylinositol-3,4,5-trisphosphate (PIP3), suggesting that PLC– 1 can be targeted to the membrane in response to GF stimulation (15). Although the SH3 domain of PLC– 1 might be responsible for mediating GF-induced mitogenesis (16, 17), what is most striking about the paper by Snyder and colleagues is the suggestion that PLC– 1 itself can directly act as a physiologically important GEF in the nucleus through its SH3 domain and this GEF activity accounts for the role of PLC– 1 in mediating NGFstimulated cell mitogenesis. Before the publication of the Snyder lab’s results (2), the role of PLC– 1 in GF-induced mitogenesis had been supported by various studies. Most of these studies indicated that the mitogenic property of PLC– 1depends on its interaction with RTKs or on the activation of its enzymatic activity and subsequent signaling cascades (1, 18). The intracellular injection of PLC– 1-specific antibodies blocks serumand Ras-stimulated DNA synthesis, indicating that PLC– 1 is required for serum-induced mitogenesis and that PLC– 1 is downstream of Ras (19). Another report has showed that inhibition of Ras activity—by the use of the dominant-negative Ras mutant N17Ras—has no effect on GFinduced activation of PLC– 1 (20). Recently, it has been shown that PLC– 1 is required for PDGF-induced DNA synthesis and c-Fos expression (21), and that homozygous disruption of the PLC– 1 gene (Plc 1) results in embryonic lethality at approximately embryonic day (E) 9.0 (22). Additionally, PLC– 1 enzymatic activity is required for GF-induced mitogenesis, and the function of PLC– 1 in GF-induced mitogenesis may be to produce diacylglycerol (DAG), which subsequently activates most protein kinase C (PKC) isoforms (23–25). Together, these results support a “phospholipase” model for PLC– 1 in GF-induced mitogenesis (Figure 2). In this model, after RTKs are activated by GFs, the SH2 domains of PLC– 1interact with the phosphotyrosine sites on RTKs—this interaction results in the translocation of PLC– 1 to the plasma membrane. The PH domain of PLC– 1 might act to stabilize the binding of PLC– 1 to the plasma membrane. Binding of PLC– 1 to RTKs results in the phosphorylation of PLC– 1 and the further activation of its enzymatic activity (18). Increased amounts of DAG are produced by activated PLC– 1 whereby PKC becomes activated and phosphorylates the protein serine-threonine kinase Raf (26–32). Activated Raf stimulates mitogen-activated protein kinase (MAPK), eventually leading to the activation of transcriptional factors c-Jun and c-Fos and DNA synthesis. Snyder and coworkers show that whereas the PLC– 1 SH2 domain and lipase activity are not required for NGF-induced cell mitogenesis, the PLC– 1 SH3 domain is necessary and sufficient to mediate NGF-induced cell mitogenesis (2, 3). PLC– 1 translocates from cytoplasm to the nucleus in response to NGF stimulation. The nuclear PLC– 1 then binds to and activates PIKE through its SH3 domain, whereupon activated PIKE then stimulates nuclear PI3K activity. Thus, PLC– 1 mediates the NGFdependent activation of nuclear PI3K activity. This enzymatic activity accounts for the mitogenic properties of PLC– 1. These results suggest a “GEF” model for PLC– 1 to mediate GF-induced Viewpoint