TIM, a Dbl-related protein, regulates cell shape and cytoskeletal organization in a Rho-dependent manner.

The Dbl-like guanine nucleotide exchange factors (GEFs) have been implicated in direct activation of the Rho family of small GTPases. We previously isolated transforming immortalized mammary (TIM) as a Dbl-like protein. Here, we show that, when expressed in cells, TIM was a potent activator of RhoA. Like activated Rho proteins, expression of TIM potentiated the serum response factor (SRF)- and AP-1-regualted transcriptional activities and activated the SAPK/JNK signaling pathway. In NIH 3T3 cells, TIM induced transforming foci, which was inhibited by the ROCK inhibitor Y-27632 or the dominant negative mutants of Rho proteins. Expression of TIM led to pronounced changes in cell shape and organization of the actin cytoskeleton, including the formation of thick stress fibers at the cell periphery and cell rounding. TIM also promoted redistribution of vinculin-enriched focal adhesions at the cell periphery and increased the phosphorylation of myosin light chain (MLC). These results, taken together, suggest that TIM acts as an upstream regulator for the RhoA/ROCK-mediated cellular functions.