Molecular cloning and characterization of a phospholipid hydroperoxide glutathione peroxidase gene from a blood fluke Schistosoma japonicum.
Phospholipid hydroperoxide glutathione peroxidase (PHGPx) is a major antioxidant enzyme and plays critical roles in the protection of cells against oxidative stress by catalysing reduction of lipid hydroperoxides. A full-length cDNA sequence corresponding to GPx gene from Schistosoma japonicum (designated SjGPx) was isolated and characterized. SjGPx 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. Phylogenetic analysis revealed that the SjGPx was highly related to the other PHGPx-related members, and clustered into the trematode subclade II. Semi-quantitative reverse transcription PCR and western blotting showed that the SjGPx was mainly expressed in the female adults and eggs. RNA interference was employed to investigate the effects of knockdown of SjGPx. SjGPx expression level was significantly reduced on the 5th day post-RNAi. We observed a 53.86% reduction in total GPx activity and the eggs severely deformed. Oxidative stimulation of viable worms with H2O2 or paraquat resulted in 1.6- to 2.1-fold induction of the GPx activity. Our results revealed that the SjGPx protein is selenium-dependent PHGPx, which might actively participate in the detoxification of oxidative damage during egg production.
Department of Medical Genetics, Wuhan University School of Basic Medicial Sciences, Wuhan, China.
Ying
Y
Zhang
Department of Medical Parasitology, Wuhan University School of Basic Medicial Sciences, Wuhan, China.
Yuan
Y
He
Department of Medical Parasitology, Wuhan University School of Basic Medicial Sciences, Wuhan, China.
Li
L
He
Department of Medical Parasitology, Wuhan University School of Basic Medicial Sciences, Wuhan, China.
Hong-Ying
HY
Zong
Department of Medical Parasitology, Wuhan University School of Basic Medicial Sciences, Wuhan, China. Electronic address: gbcai2011@whu.edu.cn.
Guo-Bin
GB
Cai
D000818:Animals; D001483:Base Sequence; D003001:Cloning, Molecular; D018388:Codon, Terminator; D018076:DNA, Complementary / Q000235:genetics / Q000302:isolation & purification; D018509:DNA, Helminth / Q000235:genetics / Q000302:isolation & purification; D005260:Female; D055785:Gene Knockdown Techniques; D005979:Glutathione Peroxidase / Q000145:classification / Q000235:genetics / Q000378:metabolism*; D015801:Helminth Proteins / Q000145:classification / Q000235:genetics / Q000378:metabolism*; D006861:Hydrogen Peroxide / Q000378:metabolism; D008297:Male; D051379:Mice; D008807:Mice, Inbred BALB C; D010063:Ovum / Q000378:metabolism; D018384:Oxidative Stress / Q000502:physiology; D000080662:Phospholipid Hydroperoxide Glutathione Peroxidase; D010802:Phylogeny; D034622:RNA Interference; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012549:Schistosoma japonicum / Q000201:enzymology* / Q000235:genetics; D012643:Selenium / Q000737:chemistry; D017279:Selenocysteine / Q000737:chemistry; D012908:Snails / Q000469:parasitology
D018388:Codon, Terminator; D018076:DNA, Complementary; D018509:DNA, Helminth; D015801:Helminth Proteins; D017279:Selenocysteine; D006861:Hydrogen Peroxide; D000080662:Phospholipid Hydroperoxide Glutathione Peroxidase; D005979:Glutathione Peroxidase; D012643:Selenium
Oxidative stress; Phospholipid hydroperoxide glutathione peroxidase (GPx); RNA interference; Schistosoma japonicum; Selenium-dependent GPx