Identification of a stress-responsive Onchocerca volvulus glutathione S-transferase (Ov-GST-3) by RT-PCR differential display.

Abstract

The effects of oxidative insult on gene transcript levels in the filarial nematode Onchocerca volvulus were investigated using differential display RT-PCR. Oxidative stress was applied with the reagents paraquat, plumbagin and xanthine-xanthine oxidase. In all three cases, a cDNA fragment encoding a novel glutathione S-transferase (GST) resembling members of the theta-class was identified as upregulated (PQ29, PG112, XOD26). The subsequently isolated full-length cDNA harbors a 753-bp open reading frame encoding a GST with 268 amino acid residues and a predicted molecular mass of 31 kDa. This stress-responsive GST (Ov-GST-3) possesses only 14 and 21% sequence identity with the other O. volvulus GSTs (Ov-GST-1 and Ov-GST-2, respectively). Interestingly, Ov-GST-3 shares higher sequence identity with GSTs that are upregulated due to environmental stress. In order to confirm the specific upregulation of the Ov-GST-3 transcripts identified by differential display and to analyze the mRNA levels of the other Ov-GSTs (Ov-GST-1 and Ov-GST-2) under elevated stress conditions, a semi-quantitative polymerase chain reaction-enzyme-linked immunosorbent assay was performed. The Ov-GST-3 gene transcript level increased dramatically in response to xanthine-xanthine oxidase and to a lesser extent with paraquat and plumbagin. In contrast, Ov-GST-1 and Ov-GST-2 did not show any significant alterations in their steady-state mRNA levels in response to oxidative stress when examining the same mRNA samples. The present study clearly demonstrates that Ov-GST-3 is a critical enzyme in the defense against oxidative stress.

• Authors: E Liebau, M L Eschbach, W Tawe, A Sommer, P Fischer, R D Walter, K Henkle-Dührsen
• Journal: Molecular and biochemical parasitology (Mol Biochem Parasitol) Vol. 109 Issue 2 Pg. 101-10 (Jul 2000) ISSN: 0166-6851 [Print] Netherlands
• PMID: 10960169 (Publication Type: Journal Article, Research Support, Non-U.S. Gov't)

Chemical References

• DNA, Complementary
• RNA, Helminth
• RNA, Messenger
• Glutathione Transferase

Topics

• Amino Acid Sequence
• Animals
• Cloning, Molecular
• DNA, Complementary
• Enzyme-Linked Immunosorbent Assay
• Female
• Glutathione Transferase (chemistry, genetics, metabolism)
• Molecular Sequence Data
• Onchocerca volvulus (enzymology, physiology)
• Oxidative Stress
• RNA, Helminth (genetics, metabolism)
• RNA, Messenger (genetics, metabolism)
• Reverse Transcriptase Polymerase Chain Reaction (methods)
• Sequence Analysis, DNA
• Transcription, Genetic