The phosphatidylinositol-3-kinase (PI3K) pathway is upregulated in a variety of human cancers, such as hepatocellular carcinoma (HCC), and is thought to promote proliferation, motility and survival of tumor cells. Class la PI3Ks are lipid kinases consisting of two subunits: a p85 regulatory subunit and a p110 catalytic subunit. The molecular mechanisms responsible for down regulation of active p110 are not known. Recently, we discovered a novel kringle containing transmembrane protein we call zPA that harbors a p85-1ike domain in its C-terminal, intracellular portion. Our preliminary experiments show that zPA associates with the p110 subunit of PI3K possibly through its p85-1ike domain and that it inhibits the enzymatic activity of p110. We demonstrate that induced zPA overexpression downregulates cell growth and cell survival in liver and other tumor cells in vitro and in vivo. In human liver tumor tissues (i.e. hepatocellular carcinoma [HCC] and hepatoblastoma [HB]), zPA protein levels are altered as compared to adjacent or non-malignant liver in western blot analysis. The molecular mechanisms underlying primary human liver cancer, such as HCC and HB, are ill-defined, but aberrant tyrosine kinase and PI3K signaling may be involved. Based on these and other preliminary data, we hypothesize that zPA binds to the p110 subunit of PI3K to downregulate its enzymatic activity, and accordingly, functions as a negative growth regulator. To test this hypothesis in liver growth and neoplasia, we propose three specific aims:
Aim 1 --to determine the molecular mechanism(s) through which zPA exerts its effect on liver cell growth regulation. We will characterize the molecular interaction of zPA with the pl 10 subunit of PI3K in liver cells, analyze the proteolytic processing of zPA protein, and identify proteins other than p110 that interact with zPA.
Aim 2 --to determine the expression and association patterns of zPA and p110 as well as the mutational profiles of the zPA gene in normal, diseased and malignant human liver tissues.
Aim 3 --to examine the in vivo function of zPA in liver growth, regeneration and tumorigenesis through gain-of-function and loss-of-function animal models. We have generated transgenic mice to overexpress zPA in the liver under the transcriptional regulation of the albumin promoter/enhancer. We propose to produce a liver specific knock out mouse model of zPA. Together, these studies should provide clues about the role of zPA and the PI3K pathway in normal and aberrant liver growth.
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