Homodimerization of the deleted in liver cancer 2 (DLC2) is not mediated by the SAM domain

Abstract

Deleted in liver cancer 2 (DLC2) is a novel tumor suppressorgene which is found to be frequently deleted in hepatocellularcarcinoma. Although DLC2 is consisted of a RhoGAP homologydomain, a START domain and a sterile-a-motif (SAM) domain,the role of DLC2 in cellular function remains elusive. To gaininsight on the function of DLC2, we expressed and purified arecombinant13C and15N doubled DLC2 SAM domain. Circulardichromism analysis showed that DLC2 SAM contains high heli-ces. We further determined the structure of the DLC2 SAMdomain by 2D and 3D NMR experiments together with simula-ted annealing calculations. Our data revealed that the structureof DLC2 SAM is slightly different from other known SAMdomains and adapts a monomeric structure with four alpha heli-ces in solution. Consistent with the structural data, cross-linkingexperiment also suggested that DLC SAM exists in predomin-antly monomeric form in vitro. On the other hand, co-immuno-precipitation experiments suggested that the DLC2 protein ishomodimerized. Analysis of the deletion mutants revealed thatthe homodimerization is not mediated by the SAM domain, butthrough a distinct region on the DLC2 protein.