WRN modulates translation by influencing nuclear mRNA export in HeLa cancer cells.

BACKGROUND: The Werner syndrome protein (WRN) belongs to the RecQ family of helicases and its loss of function results in the premature aging disease Werner syndrome (WS). We previously demonstrated that an early cellular change induced by WRN depletion is a posttranscriptional decrease in the levels of enzymes involved in metabolic pathways that control macromolecular synthesis and protect from oxidative stress. This metabolic shift is tolerated by normal cells but causes mitochondria dysfunction and acute oxidative stress in rapidly growing cancer cells, thereby suppressing their proliferation. RESULTS: To identify the mechanism underlying this metabolic shift, we examined global protein synthesis and mRNA nucleocytoplasmic distribution after WRN knockdown. We determined that WRN depletion in HeLa cells attenuates global protein synthesis without affecting the level of key components of the mRNA export machinery. We further observed that WRN depletion affects the nuclear export of mRNAs and demonstrated that WRN interacts with mRNA and the Nuclear RNA Export Factor 1 (NXF1). CONCLUSIONS: Our findings suggest that WRN influences the export of mRNAs from the nucleus through its interaction with the NXF1 export receptor thereby affecting cellular proteostasis. In summary, we identified a new partner and a novel function of WRN, which is especially important for the proliferation of cancer cells.

Measurements of Hydrogen Peroxide and Oxidative DNA Damage in a Cell Model of Premature Aging.

Reactive oxygen species (ROS) represent a number of highly reactive oxygen-derived by-products generated by the normal mitochondrial respiration and other cellular metabolic reactions. ROS can oxidize macromolecules including lipids, proteins, and nucleic acids. Under physiological condition, the cellular levels of ROS are controlled by several antioxidant enzymes. However, an imbalance between ROS production and detoxification results in oxidative stress, which leads to the accumulation of macromolecular damage and progressive decline in normal physiological functions.Oxidative deterioration of DNA can result in lesion that are mutagenic and contribute to aging and age-related diseases. Therefore, methods for the detection of ROS and oxidative deterioration of macromolecules such as DNA in cells provide important tool in aging research. Here, we described protocols for the detection of cytoplasmic and mitochondria pools of hydrogen peroxide, and the DNA modification 8-oxoguanine, a biomarker of oxidative damage, that are applicable to cell-based studies on aging and other related areas.