Dhurgham ALFAHAD, Philip DASH, Salem ALHARETHI, Bandar ALHARBI, Khatab MAWLOOD

PtdIns(4,5)P2 and PtdIns(3,4,5)P3 dynamics during focal adhesions assembly and disassembly in a cancer cell line

Focal adhesions (FAs) are large assemblies of proteins that mediate intracellular signals between the cytoskeleton and theextracellular matrix (ECM). The turnover of FA proteins plays a critical regulatory role in cancer cell migration. Plasma membranelipids locally generated or broken down by different inositide kinases and phosphatase enzymes to activate and recruit proteins tospecific regions in the plasma membrane. Presently, little attention has been given to the use of phosphatidylinositol 4,5-bisphosphate(PtdIns(4,5)P2) and Phosphatidylinositol 3,4,5-trisphosphate (PtdIns(3,4,5)P3) fluorescent biosensors in order to determine thespatiotemporal organisation of PtdIns(4,5)P2 and PtdIns(3,4,5)P3 within and around or during assembly and disassembly of FAs. Inthis study, specific biosensors were used to detect PtdIns(4,5)P2, PtdIns(3,4,5)P3, and FAs proteins conjugated to RFP/GFP in orderto monitor changes of PtdIns(4,5)P2 and PtdIns(3,4,5)P3 levels within FAs. We demonstrated that the localisation of PtdIns(4,5)P2and PtdIns(3,4,5)P3 were moderately correlated with that of FA proteins. Furthermore, we demonstrate that local levels of PtdIns(4,5)P2 increased within FA assembly and declined within FA disassembly. However, PtdIns(3,4,5)P3 levels remained constant within FAsassembly and disassembly. In conclusion, this study shows that PtdIns(4,5)P2 and PtdIns(3,4,5)P3 localised in FAs may be regulateddifferently during FA assembly and disassembly.

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