Functional Characterizations Of Oncogene TBC1D3

TBC1D3, a primate specific gene, is amplified in 15% of primary prostate tumors and in more than half of metastatic prostate tumors. The gene amplification refers to the selective increase of gene copy number, which is resulted from the chromosomal abnormalities including double minutes (DMs) or homogeneously staining chromosomal regions (HSRs). The reason for amplification and the oncogenic mechanism of TBC1D3 have yet to be elucidated. Hippo pathway is a newly established signaling pathway that plays a major role in organ size control under physical conditions and its dysregulation leads to tumorigenesis. The typical Hippo signaling pathway is consisted of Mstl/2 kinases, Latsl/2 kinases and downstream effect proteins transcriptional cofactors YAP/TAZ in mammals. Here I report that oncogene TBC1D3, a putative GAP for Rab5 that has been undergone remarkable gene replication very recently in evolution, inhibits Hippo signaling pathway. My results showed that overexpression of TBC1D3C activate YAP and TAZ. Domain mapping revealed that the N-terminal TBC domain comprised of 1-353 amino acids of TBC1D3C is responsible for YAP/TAZ activation. To gain a better understanding of how Hippo pathway is regulated by TBC1D3C, we established a TBC1D3C Tet-on inducible expression system in HEK293A cell line. By using this cell line, I found that TBC1D3C binds directly to GEF-H1, a guanine nucleotide exchange factor for the small G protein RhoA. This interaction promotes GEF-H1 release into cytosol from microtubules and its subsequent activation, which in turn activates RhoA. Active RhoA promotes actin filament formation leading to inactivation of the kinases Latsl/2, thereby activating YAP and TAZ. Taken together, my studies suggested that TBC1D3 plays an unanticipated and unique role in the regulation of Hippo pathway through its activation of GEF-H1 and RhoA, which might provide a new insight into the mechanism of TBC1D3 tumorigenesis. This work revealed a new activation mode of GEF-H1 and deepened our understanding on the signaling coupling between microtubules and actin cytoskeleton.