Combined screening of thymocytes using apoptosis-specific cDNA array and promoter analysis yields novel gene targets mediating TCDD-induced toxicity.
We have used pathway-specific cDNA arrays coupled with analysis of gene promoter regions to identify novel genes that may mediate the toxic effects of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) in the thymus. C57BL/6 mice were injected ip with 50 microg/kg TCDD, and 3, 6, or 24 h later, RNA was extracted from the thymus and subjected to microarray analysis. Several members of the TNF and TNFR family were induced following TCDD exposure, including receptor/ligand pairs Ltbeta-R/LIGHT, OX40/OX40L and TNF-alpha/TNFR1. In addition, Fas and CD30 were also upregulated. Pro-apoptotic bcl-2 gene family members Bax and Hrk, among others, were also induced, as were pro-survival bcl-2 family genes Bcl-x and Bcl-w. Cell-cycle regulator p21Cip1 was also induced. In addition, we analyzed the promoter regions of genes induced by TCDD for the presence of dioxin-responsive elements (DREs). The Fas and LIGHT gene promoters were found to contain DREs as analyzed by Matinspector Web-based search algorithm. Furthermore, binding of the aryl hydrocarbon receptor (AhR) to the DREs present on these genes was confirmed by chromatin immunoprecipitation. Given that several of the genes, including Fas, LIGHT, and CD30 are involved in negative selection of T cells in the thymus, our studies suggest that TCDD-induced upregulation of these genes may enhance negative selection leading to thymic atrophy.
Fisher
Michael T
MT
Department of Pharmacology and Toxicology, Medical College of Virginia Campus, Virginia Commonwealth University, Richmond, Virginia 23113, USA.
Nagarkatti
Mitzi
M
Nagarkatti
Prakash S
PS
D000818:Animals; D017209:Apoptosis / Q000502:physiology*; D002453:Cell Cycle / Q000187:drug effects / Q000235:genetics; D002843:Chromatin / Q000235:genetics; D018076:DNA, Complementary / Q000096:biosynthesis* / Q000235:genetics; D004785:Environmental Pollutants / Q000633:toxicity*; D005260:Female; D015870:Gene Expression / Q000187:drug effects*; D019254:Genes, bcl-2 / Q000235:genetics; D051034:Inhibitor of Apoptosis Proteins; D008024:Ligands; D051379:Mice; D008810:Mice, Inbred C57BL; D020411:Oligonucleotide Array Sequence Analysis; D000072317:Polychlorinated Dibenzodioxins / Q000633:toxicity*; D011233:Precipitin Tests; D011401:Promoter Regions, Genetic / Q000235:genetics*; D011506:Proteins / Q000235:genetics; D012333:RNA, Messenger / Q000096:biosynthesis; D018336:Receptors, Aryl Hydrocarbon / Q000187:drug effects / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013601:T-Lymphocytes / Q000187:drug effects*; D013950:Thymus Gland / Q000166:cytology / Q000187:drug effects
D002843:Chromatin; D018076:DNA, Complementary; D004785:Environmental Pollutants; D051034:Inhibitor of Apoptosis Proteins; D008024:Ligands; D000072317:Polychlorinated Dibenzodioxins; D011506:Proteins; D012333:RNA, Messenger; D018336:Receptors, Aryl Hydrocarbon