Expression and characterization of a phospholipid hydroperoxide glutathione peroxidase gene in Schistosoma japonicum.
Phospholipid hydroperoxide glutathione peroxidase (PHGPx, GPx4) is a major antioxidant enzyme, which plays unique roles in the protection of cells against oxidative stress by catalysing reduction of lipid hydroperoxides. We isolated and characterized a full-length cDNA sequence encoding GPx gene from a blood fluke, Schistosoma japonicum (designated SjGPx), which contained an in-frame TGA codon for selenocysteine (Sec) and a concurrent Sec insertion sequence in its 3'-untranslated region. Protein encoded by SjGPx demonstrated a primary structure characteristic to the PHGPx family, including preservation of catalytic domains and absence of the subunit interaction domains. Semi-quantitative reverse transcription PCR and Western blotting showed that the SjGPx was mainly expressed in the female adults and eggs. RNA interference approach was employed to investigate the effects of knockdown of SjGPx. SjGPx expression level was significantly reduced on the 5th day post-RNAi. Significantly reduction in GPx enzyme activities, as well as obvious changes in morphology of intrauterine eggs followed the reduction in SjGPx transcript level. We observed a 63·04% reduction in GPx activity and the eggs severely deformed. Our results revealed that SjGPx protein might be involved in the provision of enzyme activity during egg production.
Department of Medical Genetics,Wuhan University School of Basic Medical Sciences,Wuhan 430071,China.
Ying
Y
Zhang
Department of Medical Parasitology,Wuhan University School of Basic Medical Sciences,Wuhan 430071,China.
Yuan
Y
He
Department of Medical Parasitology,Wuhan University School of Basic Medical Sciences,Wuhan 430071,China.
L I
LI
He
Department of Medical Parasitology,Wuhan University School of Basic Medical Sciences,Wuhan 430071,China.
Hongying
H
Zong
Department of Medical Parasitology,Wuhan University School of Basic Medical Sciences,Wuhan 430071,China.
Guobin
G
Cai
D000595:Amino Acid Sequence; D000818:Animals; D015153:Blotting, Western; D002681:China; D005260:Female; D015971:Gene Expression Regulation, Enzymologic; D055785:Gene Knockdown Techniques; D005979:Glutathione Peroxidase / Q000737:chemistry / Q000145:classification / Q000235:genetics*; D008054:Lipid Peroxides / Q000378:metabolism*; D008297:Male; D051379:Mice; D008807:Mice, Inbred BALB C; D010063:Ovum / Q000201:enzymology; D010084:Oxidation-Reduction; D018384:Oxidative Stress; D000080662:Phospholipid Hydroperoxide Glutathione Peroxidase; D010802:Phylogeny; D034622:RNA Interference; D012330:RNA, Double-Stranded; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012549:Schistosoma japonicum / Q000201:enzymology* / Q000235:genetics*; D012554:Schistosomiasis japonica / Q000469:parasitology; D012908:Snails / Q000469:parasitology; D013047:Specific Pathogen-Free Organisms
D008054:Lipid Peroxides; D012330:RNA, Double-Stranded; D000080662:Phospholipid Hydroperoxide Glutathione Peroxidase; D005979:Glutathione Peroxidase
RNA interference; Schistosoma japonicum; egg production; phospholipid hydroperoxide GPx; selenium-dependent GPx