Overexpression of the human ubiquitin E3 ligase CUL4A alleviates hypoxia-reoxygenation injury in pheochromocytoma (PC12) cells.

The ubiquitin E3 ligase CUL4A plays important roles in diverse cellular processes including carcinogenesis and proliferation. It has been reported that the expression of CUL4A can be induced by hypoxic-ischemic injury. However, the effect of elevated expression of CUL4A on hypoxia-reoxygenation injury is currently unclear. In this study, human CUL4A (hCUL4A) was expressed in rat pheochromocytoma (PC12) cells using adenoviral vector-mediated gene transfer, and the effects of hCUL4A expression on hypoxia-reoxygenation injury were investigated. In PC12 cells subjected to hypoxia and reoxygenation, we found that hCUL4A suppresses apoptosis and DNA damage by regulating apoptosis-related proteins and cell cycle regulators (Bcl-2, caspase-3, p53 and p27); consequently, hCUL4A promotes cell survival. Taken together, our results reveal the beneficial effects of hCUL4A in PC12 cells upon hypoxia-reoxygenation injury.

Molecular and functional characterization of sulfiredoxin homologs from higher plants.

By reducing cysteine-sulfinic acid in oxidized peroxiredoxin, sulfiredoxin (Srx) plays an important role in oxidation stress resistance in yeast and human cells. Here, we report the first molecular and functional characterization of Srx homolog from higher plants. Bioinformatic analysis revealed the presence of potential Srx encoding sequences in both monocot and dicot plant species. Putative plant Srx proteins exhibited significant identities to their orthologs from yeast and human, and contained the conserved signature sequence and residues essential for catalysis. However, unlike yeast and human orthologs, plant Srxs were all predicted to possess chloroplast transit peptide in their primary structure. The Srx proteins from Arabidopsis and rice (designated as AtSrx and OsSrx, respectively) complemented functional deficiency of Srx in the SRX1 deletion yeast cells. A GFP fusion protein of AtSrx was targeted to chloroplast in Arabidopsis mesophyll protoplast. AtSrx transcription occurred in both vegetative and reproductive organs, and the highest transcript level was detected in leaves. Under oxidation stress, AtSrx transcript level was substantially increased, which paralleled with enhanced transcription of 2-Cys peroxiredoxins that have been found essential in maintaining chloroplast redox balance. In addition to oxidation stress, osmotic/water deficit or cold treatments also raised AtSrx transcript level. Consistent with above findings, the knock-out mutant of AtSrx was significantly more susceptible to oxidation stress than wild type Arabidopsis plant. Taken together, the results of this work indicate the existence of functional Srx homolog in higher plants that is essential for plants to cope with oxidation stress.