Silencing of the methionine sulfoxide reductase A gene results in loss of mitochondrial membrane potential and increased ROS production in human lens cells

Abstract

Accumulation of methionine sulfoxide (Met(O)) is a significant feature of human cataract and previous studies have shown that methionine sulfoxide reductase A (MsrA), which acts to repair Met (O), can defend human lens cells against oxidative stress induced cell death. A key feature of oxidative stress is increased reactive oxygen species (ROS) in association with loss of mitochondrial function. Here, we sought to establish a potential role for MsrA in the accumulation of ROS in lens cells and the corresponding mitochondrial membrane potential in these cells. Targeted gene silencing was used to establish populations of lens cells expressing different levels of MsrA, and the mitochondrial membrane potential and ROS levels of these cell populations were monitored. Decreased MsrA levels were found to be associated with loss of cell viability, decreased mitochondrial membrane potential, and increased ROS levels in the absence of oxidative stress. These effects were augmented upon oxidative stress treatment. These results provide evidence that MsrA is a major determinant for accumulation of ROS in lens cells and that increased ROS levels in lens cells are associated with a corresponding decrease in mitochondrial membrane potential that is likely related to the requirement for MsrA in lens cell viability.

Mitochondrial function and redox control in the aging eye: Role of MsrA and other repair systems in cataract and macular degenerations

“…Viability in this work was measured using the MTS assay which measures the activity of mitochondrial enzymes indicating that MsrA is important in protecting mitochondrial function in the lens cell. Marchetti et al, (2006) carried out further studies on the effect of silencing the MsrA gene. They showed that decreased MsrA levels not only reduced cell viability but also allowed increased ROS production in the cell and a decrease in mitochondrial membrane potential even in the absence of oxidative stress, these effects were augmented by TBHP-induced oxidative stress.…”

Dual Sites of Protein Initiation Control the Localization and Myristoylation of Methionine Sulfoxide Reductase A

“…Bacteria grow well when methionine is replaced by its carbon-containing analogue, norleucine, but the cells are much more susceptible to killing by oxidative stresses (3). Several investigations demonstrated that modulation of MsrA activity altered the level of reactive oxygen species; overexpression lowered the level in PC-12 cells (13), whereas lens cells lacking MsrA had an increased level (14). Increased expression of either MsrA in WI-38 fibroblasts or MsrB in MOLT-4 leukemia cells reduced the steady state level of oxidized proteins after a hydrogen peroxide challenge (15,16).…”

Two Novel Approaches Providing Cardiac Protection Against Oxidative Stress

“…In contrast, knocking out MsrA makes mammalian cells more sensitive to oxidative stress. MsrA and MsrB knockout in lens and retinal cells make these cells more sensitive to oxidation (Marchetti et al, 2006;Kantorow et al, 2004). There have been reports on MsrA knockout mice in which it is clear that these animals are very sensitive to increased oxygen tension, although there is conflicting data on their life span and the presence of a neurological defect (Moskovitz et al, 2001, Salmon et al, 2009).…”

“…There are now several cell culture studies to support this mechanism. In PC12 cells over-expression of MsrA lowers the level of ROS in the cells and in lens and retinal cells knocking out of MsrA leads to higher ROS levels (Yermolaieva et al, 2004;Marchetti et al, 2006).…”