Dottorato in SCIENZE DELLA VITA

Titolo della tesi: Using Drosophila melanogaster to dissect the role of the oncoprotein GOLPH3 in cytokinesis and cellular growth

Golgi phosphoprotein 3 (GOLPH3) is a Phosphatidylinositol-4- phosphate [PI(4)P] binding Golgi protein required for Golgi architecture, vesicle trafficking and glycosylation. Overexpression of human GOLPH3 has been reported in several solid tumors and correlates with poor prognosis in breast cancer and glioblastoma multiforme. The oncogenic activity of GOLPH3 has been linked to enhanced signaling downstream of mechanistic target of rapamycin (mTOR), an evolutionarily conserved serine/threonine protein kinase However, the molecular pathways through which GOLPH3 acts in malignant transformation remain to be clarified. It has been suggested that GOLPH3 protein might prevent tumorigenesis by (i): regulating Golgi-to-plasma secretory trafficking of growth factors, (ii): controlling the recycling or glycosylation of cancer relevant glycoproteins. Moreover, research studies in Drosophila have involved GOLPH3 protein in cytokinesis. This is highly relevant in GOLPH3 cancer biology because cytokinesis failures lead to ploidy increase, which in turn promotes chromosome instability, tumorigenesis and resistance to anti-cancer therapy. My PhD studies have been focused on the analysis of the cellular functions of GOLPH3 protein and its molecular partners using Drosophila melanogaster as a model organism. In the first chapter I investigate the role of Drosophila GOLPH3 (dGOLPH3) during the early steps of cytokinesis. dGOLPH3 accumulates at the cleavage furrow of dividing cells and interacts with the cytokinetic machinery and vesicle trafficking proteins. I demonstrate that GOLPH3 cooperates with non-muscle myosin II to regulate centralspindlin stabilization and contractile ring structure at the cleavage site. In the second chapter I use affinity purification coupled with mass spectrometry to identify the protein-protein interaction network (interactome) of Drosophila GOLPH3 in testes. Consistent with previous findings that dGOLPH3 controls membrane trafficking during cytokinesis, the dGOLPH3 interactome revealed an enrichment of proteins involved in cell cycle regulation and vesicle-mediated trafficking. In the last chapter I demonstrate that dGOLPH3 is required for tissue growth in Drosophila by regulating Rheb-mediated mTOR activation. Overall, my results shed light on the mechanisms underlying the role of GOLPH3 in cellular transformation and suggest possible therapeutic intervention for tumor pathologies characterized by GOLPH3 upregulation.