TitleIdentification and expression of alternatively spliced aryl hydrocarbon nuclear translocator 2 (ARNT2) cDNAs from zebrafish with distinct functions.
Publication TypeJournal Article
Year of Publication2000
AuthorsTanguay, RL, Andreasen, E, Heideman, W, Peterson, RE
JournalBiochim Biophys Acta
Volume1494
Issue1-2
Pagination117-28
Date Published2000 Nov 15
ISSN0006-3002
KeywordsAlternative Splicing, Amino Acid Sequence, Animals, Aryl Hydrocarbon Receptor Nuclear Translocator, Basic Helix-Loop-Helix Transcription Factors, Cloning, Molecular, COS Cells, Dimerization, Dioxins, DNA, DNA-Binding Proteins, Gene Expression Profiling, Genes, Reporter, Molecular Sequence Data, Organ Specificity, Oxygen, Polychlorinated Dibenzodioxins, Protein Binding, Protein Isoforms, Receptors, Aryl Hydrocarbon, Response Elements, RNA, Messenger, Sequence Alignment, Signal Transduction, Trans-Activators, Transcription Factors, Transcriptional Activation, Transfection, Zebrafish, Zebrafish Proteins
Abstract

In order to further establish zebrafish as a vertebrate model for studying the mechanism of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) toxicity it is necessary to characterize the aryl hydrocarbon receptor/aryl hydrocarbon receptor nuclear translocator (AhR/ARNT) signaling pathways in this species. In this study, three zfARNT2 cDNAs were isolated, expressed, and characterized and named zfARNT2b, zfARNT2c, and zfARNT2a. zfARNT2b, zfARNT2c, and zfARNT2a encode proteins with theoretical molecular weights of 81, 79, and 45 kDa, respectively. zfARNT2b and zfARNT2a proteins are identical over the first 403 amino acids but differ in their C-terminal domains as a result of alternative mRNA splicing. zfARNT2c is nearly identical to zfARNT2b, with the exception of an in frame 15 amino acid deletion adjacent to the basic region of zfARNT2c. Using quantitative RT-PCR methods the tissue distribution of each zfARNT2 isoform was determined. In COS-7 cells expressing zfARNT2b and zfAhR2, 10 nM TCDD causes a nine-fold induction of a dioxin responsive reporter gene. In COS-7 cells expressing zfARNT2a or zfARNT2c, TCDD does not induce reporter gene expression. In contrast, all three zfARNT2 proteins induce reporter gene activity under control of hypoxia responsive elements when cotransfected with the zebrafish endothelial specific PAS protein 1. DNA gel shift analysis suggests that the decreased function of zfARNT2a is due to inefficient binding of zfARNT2a/zfAhR2 complexes to dioxin responsive elements. These results also indicate that alternative mRNA splicing results in formation of ARNT proteins with distinct functional properties.

Alternate JournalBiochim. Biophys. Acta
PubMed ID11072074
Grant ListF32 ES005786 / ES / NIEHS NIH HHS / United States
2 T32 ES07015-21 / ES / NIEHS NIH HHS / United States
F32 ES05786-01 / ES / NIEHS NIH HHS / United States