Identification of a new protein that binds Src and PKC

Beth Israel Deaconess Medical Center, Boston MA

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Abstract

DESCRIPTION (provided by applicant): The broad long-term objective of this application is to determine the function of a newly identified protein that may serve as a scaffolding protein or adapter molecule for Protein Kinase C-delta (PKCdelta) and Src, two important signaling molecules that may contribute to salivary and epithelial functions. Our hypothesis is that we have identified a protein that is the product of a newly identified novel gene, and we propose a model to explain how PKCdelta and Src interact with this protein. If verified, this will serve as a basis upon which future research can be conducted. Salivary gland cells function to secrete fluids and proteins that are important to the health of the oral cavity. The proper biological activity of these cells depends on multiple proteins, including kinases that phosphorylate their appropriate substrates. One way that kinases associate with their substrates is through scaffolding proteins. Protein kinases C plays important roles in biological events in salivary glands, and understanding how PKC activities are controlled and directed will aid our understanding of salivary function, which is compromised in Sj[unreadable]gren's Syndrome and other situations. In salivary cells, PKCdelta is phosphorylated on tyrosine residues in response to neurotransmitters and conditions that alter fluid secretion and the permeability of tight junctions. The tyrosine phosphorylation of PKCdelta is mediated by Src or Src-related tyrosine kinases. We posit that a protein that we call "p130" is a transmembrane protein that represents a new type of anchoring protein for PKCdelta and Src. It is likely to play an important role in multiple types of epithelia. Using a Mass Spectrometry approach, we identified p130 by searching sequences in public databases. A recent report suggests that a newly identified gene encodes p130. The Specific Aims include: (1) To characterize the functional interrelationships among PKCdelta, Src, and p130; (2) To expand our studies of p130 in multiple types of salivary cells, including parotid and submandibular cells; and (3) To generate antibodies to p130 and identify and localize endogenous p130 within cells. These Aims should be achievable within the 2-year timetable of this project and allow us to determine the veracity of our hypothesis. Future efforts will determine if this new protein alters cell morphology, tight junction formation, or other functions in salivary cells.