Superfund Research Program
Effects Related Biomarkers of Toxic Exposures
Project Leader: Terrance J. Kavanagh
Grant Number: P42ES004696
Funding Period: 1995 - 2006
The significance of the work of the project is related to the need to develop sensitive and rational biomarkers of exposure, effect and susceptibility in human populations. To that end, investigators are characterizing the sequence of the glutamylcysteinyl ligase (GLCL) genes and the factors which influence the expression and activity of this enzyme. This research will provide useful tools that will have wide applicability in the assessment of chemical exposures, especially to those agents which harm the body by causing oxidative stress.
Using primers designed from rat cDNA sequences, RT-PCR was used to amplify and clone the regulatory and catalytic subunit GLCL cDNAs from mouse kidney. A 1.3 kb portion of the 5' regulatory region of the mouse GLCL catalytic subunit gene has been cloned and sequenced. Comparison of this sequence with that published for the human gene revealed a 56 bp region of high homology approximately 300 bp upstream of the translation start site.
Researchers are investigating potential biomarkers of methylmercury exposure by examining the effects of methylmercury on the level of GSH, GLCL activity and GLCL expression in embryos and visceral yolk sacs of mice. Chronic low dose exposures (10 ppm in the drinking water) caused reproductive toxicity (lowered fecundity), fetal malformations (anopthalmia, fetal hydrops, CNS defects) and induced GLCL expression, primarily in the yolk sac. It appears that induction of GLCL, which occurred at the lower dose (3 ppm), may be a sensitive biomarker of exposure to MeHg, as no other effects on development were noted at the low dose.
In collaboration with Bristol-Meyer Squibb, researchers are also investigating the role of a peroxyvanadate compound (pVphen, an insulin mimetic) in protein tyrosine phosphatase inhibition and oxidative stress-induced tyrosine kinase activity. pVphen has been shown to both inhibit tyrosine phosphatase but also to induce oxidative stress which results in tyrosine kinase activation in Jurkat and Ramos cell lines. In another collaboration with Bristol-Meyer Squibb, rabbit polyclonal antisera with high specificity towards peptides from each subunit of GLCL have been developed and are being used to investigate the activation of GLCL in lymphocytes. GLCL epitopes may serve as a useful biomarker of chemically-induced oxidative stress.
These projects are significant in that the characterization of GLCL genes and their expression will provide useful tools for the assessment of chemical exposures, especially to those agents which harm the body by causing oxidative stress.