Investigating the isoform-specific role of phosphoinositide 3- kinase p110delta

Phosphoinositide 3-kinases (PI3Ks) are a family of enzymes that regulate a diverse array of biological functions in every cell type by generating lipid second messengers. The PI3K family is comprised of multiple isoforms that are divided into three main classes; class I, II and III. All class I PI3Ks are heterodimers consisting of a ~110 kDa catalytic subunit associated with a regulatory subunit. Class I PI3Ks are further subdivided into class IA and class IB, which are activated downstream of tyrosine kinases or G protein-coupled receptors, respectively. The mammalian catalytic subunits of class IA PI3Ks are p110α, p110β and p110δ. PI3Ks are involved in multiple cellular processes, and deregulation of PI3K signalling pathways have been linked to a number of diseases such as cancer, inflammation and immunity and diabetes. Targeting PI3K itself or downstream effectors of PI3K is an approach that has the potential to be of huge therapeutic benefit, however, PI3K signalling is also important in normal cell homeostasis and inhibition of all the PI3K isoforms with non-selective PI3K inhibitors is toxic to the cell. To enable individual PI3K isoforms to be targeted therapeutically, it is first important to understand the specific signalling mediated by each individual PI3K isoforms. This work focuses on addressing the isoform-specific role of p110δ. Unlike p110α and p110β, p110δ has a more restricted tissue distribution and is most abundantly expressed in leukocytes. However, there are instances where cells of non-haematopoietic origin have high levels of p110δ expression, such as breast cells, melanocytes and microglia. Using cell-based models to explore p110δ signalling, our work has revealed that coupling of the tyrosine kinase receptor, c-kit, to p110δ appears to remain largely unaltered in primary leukocytes that stably express the transforming polyoma middle T antigen. Stable overexpression of p110δ in a non-leukocyte cell line, NIH 3T3, has revealed that p110δ overexpression results in the downregulation of p110α and p110β expression and has a striking impact on cell morphology. Finally, we have identified a potential p110δ gene promoter region that mediates leukocyte-specific p110δ gene expression