Aspects of the Regulation and Role of Focal Adhesion Kinase and Src in Oncogenic Transformation

Agochiya, Mahima (2000) Aspects of the Regulation and Role of Focal Adhesion Kinase and Src in Oncogenic Transformation. PhD thesis, University of Glasgow.

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Abstract

Focal adhesion kinase (FAK) is a non-receptor tyosine kinase present in cell-ECM focal adhesions. As a part of the integrin signalling complex, FAK is implicated to play a role in the mediation of integrin induced cell growth, motility and survival- processes that are also required for tumour development and progression. A role for FAK has thus been proposed in the process of tumorigenesis. In support of this, FAK is found to be upregulated in various different tumours and tumour cell lines. However, the mechanisms that lead to the upregulation of FAK in cancer cells have not been investigated thus far. The first part of this study focused on establishing some of the genetic alterations adopted by cancer cells to elevate the level of FAK protein. We used fluorescent in situ hybridisation ((FISH) to first confirm that fak localised to human chromosome 8q in normal cells. Elevated FAK protein levels in cell lines derived from invasive squamous cell carcinomas of the head and neck were accompanied by gain in fak gene copy number in all the cases examined. Moreover, increased fak gene dosage, including genetic alterations like amplification and isochromosome formation, were observed in cell lines derived from human tumours of lung, breast and colon. In addition, conversion from adenoma to carcinoma, in an in vitro model for human colon cancer progression, was accompanied by an elevation in FAK protein level and gain in fak gene copy number. Although, other genes like c-myc, lying near the fak locus, were also found to be co-amplified or increased in copy number, given the biological functions FAK is proposed to play a role, it may contribute in exerting the selective pressure for the re retention of the whole region in increased number of copies. In order to further investigate the role of FAK in tumour development, we studied its importance in cell survival. For this we used an in vitro model of apoptosis, wherein, apoptosis was induced in v-Src transformed Rat-1 cells after serum-deprivation and inhibition of v-Src activity. In particular, we examined the importance of FAK proteolysis in the induction of apoptosis. We found that although induction of apoptosis was accompanied by FAK cleavage, inhibition of FAK proteolysis was unable to promote cell survival. The in vitro model of apoptosis using v-Src transformed Rat-1 cells was also characterised further in this thesis. Like FAK, the v-Src oncoprotein also resides in the focal adhesions where its constitutive kinase activity induces disruption of focal adhesions and cell transformation. Unlike other oncoproteins like c-Myc, and v-Jun, v-Src does not induce cell death under low serum conditions. However, the data described in this thesis suggested that v-Src primes the cells to undergo apoptosis, partly through disruption of integrin signalling, while providing them with a survival signal at the same time. Removal of this survival signal, thus, induced the cells to undergo apoptosis. Further investigations indicated that the surrogate integrin survival signal provided by v-Src required the activation of the PI 3-kinase/Akt pathway, while the MAP kinase pathway did not play any role in the mediation of the survival signalling. In addition, the apoptotic response induced in v-Src transformed Rat-1 cells was shown to be accompanied by activation of caspases, and stress-activated kinase, p38. Cell death was inhibited by overexpression of the anti-apoptotic protein Bcl-2 or by the inhibtion of caspase and p38 activity simultaneously.

Item Type: Thesis (PhD)
Qualification Level: Doctoral
Additional Information: Adviser: Margaret Frame
Keywords: Molecular biology
Date of Award: 2000
Depositing User: Enlighten Team
Unique ID: glathesis:2000-75549
Copyright: Copyright of this thesis is held by the author.
Date Deposited: 19 Nov 2019 19:29
Last Modified: 19 Nov 2019 19:29
URI: http://theses.gla.ac.uk/id/eprint/75549

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