Roles of the phosphoinositides PtdIns3P, PtdIns(3,5)P2 and PtdIns5P in cell signalling and trafficking; implication in human disease. – PHOSPHOINOPATHO

Phosphoinositides (PIs) are quantitatively minor membrane glycerophospholipids which metabolism is highly regulated by a set of specific kinases and phosphatases. It has been shown that some PIs such as phosphatidylinositol 3,4,5 trisphosphate (PtdIns(3,4,5)P3) and PtdIns(4,5)P2 play a critical role in the spatiotemporal organization of many signalling complexes involved in the regulation of cellular functions such as cytoskeleton dynamics, membrane trafficking, cell proliferation and survival. Recent discoveries indicate that mutations in several PI phosphatases and kinases take part in the development of human diseases including cancer, myotubular myopathy or Lowe syndrome. Several of these enzymes are involved in the metabolism of low abundance PIs such as PtdIns3P, PtdIns5P and PtdIns(3,5)P2. Besides the well studied PtdIns(3,4,5)P3 and PtdIns(4,5)P2, the metabolism and functions of PtdIns3P, PtdIns5P and PtdIns(3,5)P2 are much less documented. PtdIns3P is a spatial regulator through its interaction with FYVE or PX domains, present in proteins involved in vesicular trafficking, endosome fission and membrane remodeling. PtdIns(3,5)P2 is predicted to play a role in vacuolar/lysosomal membrane integrity, recycling and turnover. The level of these two PIs can be decreased by the 3-phosphatases from the myotubularin family, whose founding member, MTM1, is mutated in the X-linked myotubular myopathy. PtdIns3P is the precursor of PtdIns(3,5)P2 via the action of the 5-kinase PIKfyve which localizes to late endosomes and is necessary for endosome integrity. The mechanisms in which they are implicated remain elusive and further characterization of their role and metabolism is needed. The metabolic pathway and roles of the newly discovered PtdIns5P are still poorly understood. Recent data define PtdIns5P as a potential new lipid second messenger. PtdIns5P can be synthesized by phosphorylation of PtdIns via the 5-kinase PIKfyve. It can also be generated by dephosphosphorylation of PtdIns(3,5)P2 via members of the myotubularin family. Recent studies suggest that PtdIns5P is involved in i) cell survival through activation of the Akt pathway in case of bacterial infection by the pathogen Shigella flexneri or ii) in regulating chromatin modifications to modulate gene expression, replication or DNA repair through its binding to the plant homeodomain (PHD) of inhibitor of growth protein 2 (ING2). The regulation and putative role of PtdIns5P in the intracellular membranes compartments has not been documented yet. In conclusion, studying the function of PtdIns3P, PtdIns(3,5)P2 and PtdIns5P in normal and pathological conditions raises challenging issues in both basic and applied sciences and represents an extremely competitive scientific field. The aim of this project is to develop tools to specifically study the metabolism of PtdIns3P, PtdIns5P and PtdIns(3,5)P2 and to analyse their dynamic, localization and function. We will further study how defects in their metabolism can lead to some human pathologies. Our objectives are : (i) to produce a set of fully characterized new tools to study and manipulate PtdIns3P, PtdIns5P and PtdIns(3,5)P2 metabolism and function; (ii) to identify and characterize new effectors, especially for PtdIns5P; (iii) to characterize the impact of myotubularins and PIKfyve in the metabolism of these PIs in yeast, C. Elegans and mammalian cells; (iiii) to study the consequences of deregulating these PIs on membrane trafficking, signal transduction and in fine on the physiopathology of human disease. We set up a consortium of three teams with complementary expertises to cover efficiently the different aspects of this project. Partner 1 (B. Payrastre & co-workers, Inserm U563, Toulouse) has a long-standing experience in the field of signal transduction and is centered on biochemistry of phosphoinositides with a strong interface with clinicians. Partner 2 (S. Friant & co-workers, Strasbourg) is an expert in the field of membrane trafficking and identified two yeast endosomal specific PtdIns(3,5)P2 effectors, the Ent3 and Ent5 Epsins. Partner 3 (J. Laporte & co-workers, IGBMC, Illkirch) has a long-standing experience in the field of genetic, physiopathology and models of phosphoinositides related diseases. The generation of new methods and tools to study and manipulate these phosphoinositides will benefit to the scientific community. We shall obtain a better understanding of the regulation and of the molecular, cellular and physiological roles of these phosphoinositides, and possibly reveal new functions. Moreover, we will gain insight into the molecular defects linked to myotubularin deficiency in myopathy, and PIKfyve / PtdIns5P roles in oncogenesis.