Characterization of ena/VASP domains with dominant negative function

MARIANA V. HERNÁNDEZ, D. LORENA FRANCO & M. FERNANDA CERIANI.

Huerta Grande, Cordoba

Congreso; II Reunión Conjunta de Neurociencias; 2010

Neurodegeneration is a process known to occur in metazoans. In previous work we found that the gene enabled (ena), encoding a protein involved in cytoskeleton remodeling, is implicated in progressive neurodegeneration in Drosophila. Silencing ena ortholog genes (the ena/VASP family) in mouse hippocampal neurons triggered neurite retraction and concominant neuronal cell death through an apoptotic pathway. Since our ultimate goal is to confirm these results in a mouse model of late onset progressive neurodegeneration through deregulation of ena/VASP family members, we characterized potential dominant negative (DN) versions to identify the most effective one. Ena/VASP proteins share three well-defined domains, the EVH1 domain, a central proline-rich region, and EVH2. Thus we analyzed the expression of EVH1-GFP, EVH2-GFP and GFP (control) in transfected NIH 3T3 cells. Both constructs delocalized VASP with variable efficiency. Preliminary results after DN overexpression in mouse hippocampal neurons led to neurite retraction, recreating what was observed with RNAi pools. Further experiments are needed to determine which of the DN has the most inhibitory potential on the function of Ena/VASP family members.