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Targeted disruption of the type I phospatidylinositol phosphate kinase gamma gene and the effect on focal adhesion formation in a mouse genetic background


Phosphatidylinositol-4, 5-bisphosphate (PI-4,5-P2) is a lipid molecule that contributes to the formation and maintenance of focal adhesions, by promoting the formation of an indirect physical association between integrin molecules and the actin cytoskeleton.

A splice variant of Type I phosphatidylinositol phosphate kinase gamma (PIPKIg), a key enzyme in the synthesis of PI-4.5-P2 has recently been demonstrated to localize to focal adhesions and may contribute to local synthesis of PI-4.5-P2 at the sites of focal adhesions.

The goal of this proposal is to generate a targeted gene disruption of this splice variant of PIPKIg in a mouse genetic model, with the specific aim of studying the putative requirement for localized PI-4.5-P2 production in the formation and maintenance of focal adhesions.

The phenotype of mice harbouring a disruption of the PIPKIg gene will be analysed using standard techniques, and their morphology and histology will be compared to mouse models containing disruptions of various focal adhes ion components.

Most of the models that will be used for comparison have already been generated and characterized by the host laboratory. A cell biological approach will also be used, to study the formation and architecture of focal adhesions.

Fibroblasts harbouring the targeted disruption of PIPKIg will be generated and the formation and composition of focal adhesions will be studied using immuno fluorescence, live cell imaging techniques and biochemical techniques.

In this manner the specific PI-4,5-P2-depen dent steps leading to focal adhesion formation will be elucidated. An understanding of how focal adhesions are formed and maintained will advance our understanding of the processes leading to cancer formation and metastasis, immune cell malfunction and imp aired wound healing.

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