Simvastatin Suppresses Self-Renewal of Mouse Embryonic Stem Cells By Inhibiting Rhoa Geranylgeranylation

¹ Center for Development & Differentiation, KRIBB, 52 Eoeun-dong, Yuseong-gu, Daejeon 305-806, Republic of Korea
² Department of Biological Sciences, Korea Advanced Institute of Science and Technology, 373-1 Guseong-dong, Yuseong-gu, Daejeon 305-701, Republic of Korea

^* To whom correspondence should be addressed. E-mail: june{at}kribb.re.kr.

	Abstract

Statins, 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase inhibitors, were originally developed to lower cholesterol. Their pleiotropic (or cholesterol-independent) effects at the cellular and molecular level are highly related to numerous cellular functions such as proliferation and differentiation. However, they are hardly studied in embryonic stem cells (ESCs). In this study, we evaluated the effects of statins on mouse ESCs (J1, D3, and RW.4) to enhance our understanding of the molecular basis of ESC self-renewal. Treatment of ESCs with simvastatin, mevastatin, atorvastatin, or pravastatin induced morphological change and decreased cell proliferation. We observed that the use of simvastatin was most effective among those in all three ESCs. Loss of ESC self-renewal by simvastatin was determined by marked down-regulation of ESC markers, alkaline phosphatase, Oct-4, Nanog, Rex-1, and SSEA-1. Simvastatin effects were selectively reversed by either mevalonate or its metabolite GGPP, but not by cholesterol or FPP. These results suggest that simvastatin effects were mainly derived from depletion of intracellular pools of GGPP, the substrate required for the geranylgeranylation. Using this approach, we found that GGPP, a derivative of the mevalonate pathway, is critical for ESC self-renewal. Furthermore, we identified that simvastatin selectively blocked cytosol-to-membrane translocalization of RhoA small G protein, known to be the major target for geranylgeranylation, and lowered the levels of ROCK2 protein in ESCs. In addition, simvastatin down-regulated the ROCK activity and this effect was reversed by addition of GGPP. Our data suggest that simvastatin, independently of its choresterol-lowering properties, impairs the ESC self-renewal by modulating RhoA/ROCK-dependent cell-signaling.

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