Proteomic Analysis of Protein Expression Affected by Peroxiredoxin V Knock-Down in Hypoxic Kidney

Abstract

Peroxiredoxin V, an atypical thioredoxin peroxidase, is widely expressed in mammalian tissues. In addition, Prdx V is localized in mitochondria, peroxisome, cytosol, and the nucleus. Prdx V has been reported to protect a wide range of cellular environments as an antioxidant enzyme, and its dysfunctions may be implicated in several diseases, such as cancer, inflammation, and neurodegenerative disease. Identification and relative quantification of proteins affected by Prdx V may help identify novel signaling mechanisms that are important for oxidative stress response. However, the role of Prdx V in the modulation of hypoxia-related cellular response is not studied yet. To examine the function of endogenous Prdx V in hypoxic condition in vivo, we generated a transgenic mouse model with Prdx V siRNA expression controlled by U6 promoter. Of many tissues, the knockdown of Prdx V expression was displayed in the kidney, lung, and liver but not the spleen and skin. We conducted on the basis of nano-UPLC-MS(E) proteomic study to identify the Prdx V-affected protein networks in hypoxic kidneys. In this study, we identified protein networks associated with oxidative stress, fatty acid metabolism, and mitochondrial dysfunction. Our results indicated that Prdx V affected to regulation of kidney homeostasis under hypoxia stress.

Proteome Analyses of Hydrogen-producing Hyperthermophilic Archaeon Thermococcus onnurineus NA1 in Different One-carbon Substrate Culture Conditions

“…Compared with previous approaches, the nano-UPLC-MS E method based on label-free quantitative analysis enables large scale comparison of all detectable proteins in complex samples separated by one-dimensional PAGE (21,23).…”

“…To normalize comparative proteomic data, the "auto normalization" function of ProteinLynx GlobalServer was used. Protein identification was allowed only if the confidence was greater than 95% on the basis of the IDENTITY E algorithm (21). The Expression software generated results as an exact mass retention time table with quantitative protein and peptide values.…”

Proteomic Analysis of Gingival Tissue and Alveolar Bone during Alveolar Bone Healing

“…The separated proteins were then visualized by staining with Coomassie Blue R-250, and the gels were washed three times in distilled water prior to tryptic digestion. For efficient trypsinization, the separated proteins were collected from the gels by excising 10 slices of gel for each patient sample, as described previously (26). The gel slices were then chopped into 1-mm 3 pieces and destained overnight via incubation in 50% acetonitrile and 50 mM ammonium bicarbonate.…”

Hypoxia elicits broad and systematic changes in protein subcellular localization

“…Such studies have provided useful insights into transcriptional regulatory programs. Likewise, a number of proteomic studies have been performed to identify changes in protein levels induced by altered oxygen levels in various organisms (5,10,11,17,30,37,47). However, such studies are limited by the sensitivity of the techniques used, mainly shotgun proteomic or even less sensitive methods like two-dimensional differential gel electrophoresis (2D DIGE) (1).…”

“…However, such studies are limited by the sensitivity of the techniques used, mainly shotgun proteomic or even less sensitive methods like two-dimensional differential gel electrophoresis (2D DIGE) (1). They often identify changes in abundant enzymes, structural proteins, and chaperones, but they rarely identify low-level regulatory proteins such as nuclear proteins and signaling proteins (5,10,11,17,30,37,47). Hence, more sensitive methods that can directly monitor nuclear and signaling proteins should enable us to glean important regulatory and signaling events.…”