Submitted on November 1, 2007
Accepted on December 28, 2007
RNA Suppression of ERK2 Leads to Collapse of Mitochondrial Membrane Potential with Acute Oxidative Stress in Human Lens Epithelial Cells
James M. Flynn1, Deborah A. Lannigan2, David E. Clark2, Margaret H. Garner3, and Patrick R. Cammarata4*
1 Cell Biology and Genetics, University of North Texas Health Science Center, Fort Worth, Texas, United States
2 Center for Cell Signaling and the Department of Microbiology, University of Virginia, School of Medicine, Charlottesville, Virginia, United States
3 Department of Cell Biology and Genetics, University of North Texas Health Science Center at Fort Worth, Fort Worth, Texas, United States
4 Cell Biology and Genetics, University of North Texas Hlth Sci Cntr, Fort Worth, Texas, United States
* To whom correspondence should be addressed. E-mail: pcammara{at}hsc.unt.edu.
17-β estradiol (E2) reduces oxidative stress-induced depolarization of mitochondrial membrane potential (MMP) in cultured human lens epithelial cells (HLE-B3). The mechanism by which non-genomic effects of E2 contributed to the protection against mitochondrial membrane depolarization was investigated. Mitochondrial membrane integrity is regulated by phosphorylation of BAD and it is known that phosphorylation of Ser-112 inactivates BAD and prevents its participation in the mitochondrial death pathway. We found that E2 rapidly increased both the phosphorylation of extracellular signal-regulated kinase 2 (ERK2) and Ser-112 in BAD. Ser-112 is phosphorylated by RSK, a Ser/Thr kinase, which is a downstream effector of ERK1/2. Inhibition of RSK by the RSK-specific inhibitor, SL0101, did not reduce the level of E2-induced phosphorylation of Ser-112. Silencing BAD using small interfering RNA (siRNA) did not alter mitochondrial membrane depolarization elicited by peroxide insult. However, under the same conditions, silencing ERK2 dramatically increased membrane depolarization compared to the control siRNA. Therefore, ERK2, functioning through a BAD-independent mechanism, regulates MMP in humans lens epithelial cells. We propose that estrogen-induced activation of ERK2 acts to protect cells from acute oxidative stress. Moreover, despite the fact that ERK2 plays a regulatory role on mitochondrial membrane potential, estrogen was found to block mitochondrial membrane depolarization via an ERK-independent mechanism.
