CONICET | Buscador de Institutos y Recursos Humanos

von Hippel?Lindau (VHL) disease is characterized by development of highly vascularized tumors. VHL protein drives the degradation of hypoxia-inducible transcription factors (HIFs) in normoxia. Mutations of VHL lead to a deregulation of HIFs promoting tumor development. The molecular mechanisms describing VHL mutation loss of function are still an open question. We found that RSUME is expressed in VHL-derived tumors. At molecular level, we demonstrated that RSUME interacts with VHL leading to HIFs- stabilization. The functional inhibition was also found in representative Type2 mutants (VHLY112H; VHLR167Q; VHLL188V), in which RSUME potentiates their loss of function activity. In this work we aimed to determine the mechanism of action of RSUME over VHL Type2 mutants.By co-transfecting RCC-786-O cells (VHL -/-) with RSUME expression reporter vector (RSUME-Luc) and VHL or VHL representative mutants we demonstrate that VHL exert an inhibitory action on RSUME expression which is not produced by VHL mutants, highlighting a permissive setting for the RSUME levels deregulation. In addition by co-immunoprecipitation we demonstrated RSUME interacts with each VHL type 2 mutant and HIF-2 in the same complex and displaces the binding between HIF-2 and each VHL mutant. To understand if RSUME actions are mediated by summoylation, we generated VHL171 mutant, unable to be sumoylated and we evidence that RSUME interacts with VHL171 mutant and leads HIF-2 stabilization as seen for VHLwt. Additionally we observed the interaction RSUME - non-sumoylated VHL in cellular extract treated with Gam enzyme (inhibitor of generalized sumoylation). Even more, we establish that RSUME interacts with non-sumoylables Type 2 VHL mutants (VHL mutants carrying additional 171 mutation) and allows HIF-2 stabilization. Taken together these results support a role of RSUME participation in Type2 VHL mutations functional impact.