Investigation of the role of the p110β isoform of PI 3-kinase in cell signalling and proliferation
Phosphoinositide 3-kinases (PI3Ks) are signalling enzymes that play important roles in various cellular processes and in disease, such as cancer and auto-immunity. There are several isoforms of PI3K whose individual roles have been studied extensively, amongst other to understand whether any of the PI3K isoforms may represent a target for selective pharmacological intervention in disease. In this study, I aimed to identify the mechanisms that determine the sensitivity of cancer cell growth and proliferation to inhibition of the p110β isoform of PI3K. p110β is activated downstream G-protein coupled receptors (GPCRs), and occurs in complex with a regulatory subunit named p85 which links this PI3K to tyrosine kinase signalling pathways. The parameters that determine cellular sensitivity to p110β are unclear. To characterize the role of p110β in signal transduction, its recruitment downstream a model receptor tyrosine kinase (PDGFR, platelet-derived growth factor receptor) and its binding to the different p85 isoforms was investigated. p110β was found to be recruited downstream active PDGFR alongside p110β, the other PI3K isoform that acts as the main PI3K isoform downstream such receptor. I also document that there is no preferential association of p110β with p85α or p85β, the two main p85 isoforms. In a panel of human cancer cell lines, I discovered that cancer cell lines that showed sensitivity to p110β growth and cell signalling inhibition were deficient in expression or function of IRS-1 (insulin receptor substrate-1), although they expressed IRS-1 mRNA, and this molecular trait was further investigated. Using cell transfection and RNA interference (RNAi) techniques I tried to modulate p110β sensitivity in various cellular models. Overexpression of IRS-1 protein in p110β-sensitive cancer cells could not be achieved, likely due to the same inhibitory signalling networks that prevent expression of endogenous IRS-1. Unexpectedly, downregulation of IRS expression by RNAi resulted in a positive effect of p110β inhibition on cell growth and proliferation. Such a growth-stimulating effect of inhibition of p110β was also observed under other conditions. The molecular basis for this effect is not completely clear to date and possible explanations are discussed. I conclude that p110β has a role as a signalling modulator that, upon specific circumstances, can determine both positive and negative responses in the cellular signalling circuitry and behaviour.
- Theses