Oxidative stress in Caenorhabditis elegans: protective effects of the Omega class glutathione transferase (GSTO-1).
To elucidate the function of Omega class glutathione transferases (GSTs) (EC 2.5.1.18) in multicellular organisms, the GSTO-1 from Caenorhabditis elegans (GSTO-1; C29E4.7) was investigated. Disc diffusion assays using Escherichia coli overexpressing GSTO-1 provided a test of resistance to long-term exposure under oxidative stress. After affinity purification, the recombinant GSTO-1 had minimal catalytic activity toward classic GST substrates but displayed significant thiol oxidoreductase and dehydroascorbate reductase activity. Microinjection of the GSTO-1-promoter green fluorescent protein construct and immunolocalization by electron microscopy localized the protein exclusively in the intestine of all postembryonic stages of C. elegans. Deletion analysis identified an approximately 300-nucleotide sequence upstream of the ATG start site necessary for GSTO-1 expression. Site-specific mutagenesis of a GATA transcription factor binding motif in the minimal promoter led to the loss of reporter expression. Similarly, RNA interference (RNAi) of Elt-2 indicated the involvement of this gut-specific transcription factor in GSTO-1 expression. Transcriptional up-regulation under stress conditions of GSTO-1 was confirmed by analyzing promoter-reporter constructs in transgenic C. elegans strains. To investigate the function of GSTO-1 in vivo, transgenic animals overexpressing GSTO-1 were generated exhibiting an increased resistance to juglone-, paraquat-, and cumene hydroperoxide-induced oxidative stress. Specific silencing of the GSTO-1 by RNAi created worms with an increased sensitivity to several prooxidants, arsenite, and heat shock. We conclude that the stress-responsive GSTO-1 plays a key role in counteracting environmental stress.
Burmeister
Cora
C
Institute for Animal Physiology, University of Muenster, Hindenburgplatz 55, Muenster 48143, Germany.
Lüersen
Kai
K
Heinick
Alexander
A
Hussein
Ayman
A
Domagalski
Marzena
M
Walter
Rolf D
RD
Liebau
Eva
E
D000595:Amino Acid Sequence; D000818:Animals; D017173:Caenorhabditis elegans / Q000201:enzymology / Q000235:genetics / Q000378:metabolism* / Q000648:ultrastructure; D029742:Caenorhabditis elegans Proteins / Q000737:chemistry / Q000145:classification* / Q000235:genetics / Q000378:metabolism*; D050980:GATA Transcription Factors / Q000235:genetics / Q000378:metabolism; D017353:Gene Deletion; D005786:Gene Expression Regulation; D017930:Genes, Reporter / Q000235:genetics; D005982:Glutathione Transferase / Q000737:chemistry / Q000145:classification* / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D046529:Microscopy, Electron, Transmission; D016253:Microscopy, Immunoelectron; D008969:Molecular Sequence Data; D018384:Oxidative Stress; D010802:Phylogeny; D011401:Promoter Regions, Genetic / Q000235:genetics; D034622:RNA Interference; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D012680:Sensitivity and Specificity; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid
D029742:Caenorhabditis elegans Proteins; C094134:ELT-2 protein, C elegans; D050980:GATA Transcription Factors; D011994:Recombinant Proteins; C526683:GSTO-1 protein, C elegans; D005982:Glutathione Transferase