Growth factors play key roles in normal cell physiology and tissue development and in malignant transformation and tumor development. One of these factors is transforming growth factor-a (TGF-a), which binds to the EGF/TGF-a tyrosine kinase receptor. TGF-a is considered the prototype of a family of transmembrane growth factors characterized by a conserved six-cysteine motif in the ectodomain. Transmembrane TGF-a is often the most predominant TGF-a form. Soluble TGF-a is released into the medium as a result of the regulated proteolytic cleavage of the ectodomain. Whereas both soluble and transmembrane TGF-a activate the EGF receptor, other functions are likely to be associated with the transmembrane form. In this grant application, it is proposed to characterize the molecular identity and function of the transmembrane TGF-a-associated protein complex. He has previously obtained evidence that at least several proteins associate with transmembrane TGF-a. He hypothesizes that transmembrane TGF-a associates with a regulated protein complex that is involved in the inducible ectodomain cleavage of transmembrane TGF-a, the presentation of transmembrane TGF-a at the cell surface, and/or signaling events associated with the cytoplasmic domain. The proposed research is subdivided into two large aims.
In Aim 1, the applicant will focus on the molecular identification and characterization of proteins that associate with transmembrane TGF-a. Their identification will be pursued by a combination of approaches, including protein purification, interaction cDNA cloning and cDNA cloning of homologs of proteins which in Drosophila genetically interact with TGF-a homologs. After the association of these proteins with transmembrane TGF-a has been established in vivo, the applicant will pursue in Aim 2 i.e., the functional characterization of these proteins in several contexts: the regulated proteolysis of the transmembrane TGF-a ectodomain, the affinity of transmembrane TGF-a for the EGF receptor, presentation of transmembrane TGF-a at the cell surface, and possible signaling events associated with the cytoplasmic side of transmembrane TGF-a. The molecular and functional characterization of the transmembrane TGF-a associated-protein complex should provide insight into the role of this transmembrane growth factor and other related factors in the physiology and development of normal and tumor tissues and in various epithelial disorders.
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