Abstract
Cdh1 is an activator of the anaphase-promoting complex/cyclosome and contributes to mitotic exit and G1 maintenance by targeting cell cycle proteins for degradation. However, Cdh1 is expressed and active in postmitotic or quiescent cells, suggesting that it has functions other than cell cycle control. Here, we found that homozygous Cdh1 gene-trapped (Cdh1GT/GT) mouse embryonic fibroblasts (MEFs) and Cdh1-depleted HeLa cells reduced stress fiber formation significantly. The GTP-bound active Rho protein was apparently decreased in the Cdh1-depleted cells. The p190 protein, a major GTPase-activating protein for Rho, accumulated both in Cdh1GT/GT MEFs and in Cdh1-knockdown HeLa cells. Cdh1 formed a physical complex with p190 and stimulated the efficient ubiquitination of p190, both in in vitro and in vivo. The motility of Cdh1-depleted HeLa cells was impaired; however, codepletion of p190 rescued the migration activity of these cells. Moreover, Cdh1GT/GT embryos exhibited phenotypes similar to those observed for Rho-associated kinase I and II knockout mice: eyelid closure delay and disruptive architecture with frequent thrombus formation in the placental labyrinth layer, respectively. Furthermore, the p190 protein accumulated in the Cdh1GT/GT embryonic tissues. Our data revealed a novel function for Cdh1 as a regulator of Rho and provided insights into the role of Cdh1 in cell cytoskeleton organization and cell motility.
Supplemental material for this article may be found at http://mcb.asm.org/.
We thank H. Sabe for the pEGFP full-length p190 expression vector, P. P. Pandolfi and T. Maeda for the pWZL-hyg-SV40 large T retrovirus vector, T. Shinoda for help in the quantitative analysis of fluorescent intensity, I. Ishimatsu and M. Nakata for technical assistance in the histological analyses, Y. Ito and N. Suzuki for animal maintenance, K. Arai for secretarial assistance, and M. Amano, S. Nakada, and all members of the Saya lab for helpful comments and discussions. We also thank the members of the Gene Technology Center and CARD at Kumamoto University for technical assistance and animal maintenance.
This study was supported by grants from the Ministry of Education, Culture, Sports, Science, and Technology (MEXT) of Japan (to S.K. [20590315 and 20058033] and H.S. [17013070]).