During the past funding period, we have identified a new nuclear target for heregulin, an activator of the Neu/ErbB2 tyrosine kinase, which enables this growth factor to impart a previously unappreciated mode of regulation of gene expression. The heregulin-responsive target is the heterodimeric RNA cap-binding protein complex, designated the CBC (consisting of the CBP20 and CBP80 subunits), which binds the 7methyl guanosine cap structures on RNAs transcribed by RNA polymerase II. CBC activity is essential for an early step in the splicing of precursor messenger RNA, as well as for the nuclear export of U snRNAs which are required for spliceosome assembly. We find that heregulin (as well as nerve growth factor) strongly stimulates the binding of capped RNA to the CBC and its ability to initiate splicing activity. Thus, we have uncoverered a heregulin-coupled signaling pathway to the nucleus which can regulate gene expression at the level of RNA processing: In this competitive-renewal, we plan to take advantage of this finding to obtain new insights into the signaling activities that mediate growth factor- stimulated nuclear events and the potential roles for regulated RNA processing in cell growth and differentiation. Three major experimental aims are proposed. The first is to delineate the signaling pathway leading from heregulin binding at the cell surface to the activation of the CBC in the nucleus. Given that the GTP-binding protein Cdc42 is an effective activator of the CBC, we will set out to establish how receptor tyrosine kinases promote the activation of this GTP-binding protein, as well as identify the Cdc42 signaling target(s) necessary for stimulating CBC activity.
The second aim focuses on understanding the molecular mechanism by which the CBC is stimulated to bind capped RNA. Based on our recent findings suggesting a link between a rapamycin-sensitive phosphorylation of CBP80 and CBC activation, we will test the idea that the phosphorylation of CBP80 stimulates the CBC to exchange processed mRNA for a precursor mRNA molecule.
The third aim will center on examining the cellular consequences of the growth factor-- stimulated activation of the CBC. We will establish whether CBC activation by heregulin and nerve growth factor reflects an important role in some aspect of cellular differentiation and/or if there are conditions where CBC regulation is involved in malignant transformation or cell survival. The expectation is that these studies will highlight important new participants in receptor tyrosine kinase-coupled signaling pathways and novel targets for therapeutic intervention against cancer.
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