Functional analysis of the Drosophila RhoGAP Cv-c protein and its equivalence to the human DLC3 and DLC1 proteins

RhoGAP proteins control the precise regulation of the ubiquitous small RhoGTPases. The Drosophila Crossveinless-c (Cv-c) RhoGAP is homologous to the human tumour suppressor proteins Deleted in Liver Cancer 1-3 (DLC1-3) sharing an identical arrangement of SAM, GAP and START protein domains. Here we analyse in Drosophila the requirement of each Cv-c domain to its function and cellular localization. We show that the basolateral membrane association of Cv-c is key for its epithelial function and find that the GAP domain targeted to the membrane can perform its RhoGAP activity independently of the rest of the protein, implying the SAM and START domains perform regulatory roles. We propose the SAM domain has a repressor effect over the GAP domain that is counteracted by the START domain, while the basolateral localization is mediated by a central, non-conserved Cv-c region. We find that DLC3 and Cv-c expression in the Drosophila ectoderm cause identical effects. In contrast, DLC1 is inactive but becomes functional if the central non-conserved DLC1 domain is substituted for that of Cv-c. Thus, these RhoGAP proteins are functionally equivalent, opening up the use of Drosophila as an in vivo model to analyse pharmacologically and genetically the human DLC proteins.