Part Of DNA Binding Domain Research Articles

Identification of two novel isoforms of mouse NUR77 lacking N-terminal domains.

Nur77 is a member of nuclear receptor superfamily that acts as a transcription factor and regulates expression of multiple genes. Subcellular localization of Nur77 protein plays an important role in the survival and cell death. In this study, we have predicted and confirmed alternatively spliced two new transcripts of Nur77 gene in mouse. The newly identified transcripts have their alternatively spliced first exon located upstream of published 5'-UTR of the gene. Transcription factor binding sites in the possible promoter regions of these transcripts were also analyzed. Expression of novel transcript variants was found to be significantly lower than the already published transcript. New transcript variants encode for NUR77 protein isoforms which are significantly smaller in size due to lack of transactivation domain and a part of DNA binding domain. Western blot analysis using NUR77 specific antibody confirmed the existence of these smaller variants in mouse. Localization of these new isoforms was predicted to be majorly outside the nucleus. In silico analysis of the conceptually translated proteins was performed using different bioinformatics tools. The results obtained in this study offer further insight into novel area of research on extensively studied Nur77. © 2017 IUBMB Life, 69(2):106-114, 2017.

A mineralocorticoid-like receptor in the rainbow trout, Oncorhynchus mykiss: cloning and characterization of its steroid binding domain

Using reverse transcriptase polymerase chain reaction (PCR) (RT-PCR) with degenerate primers followed by 3′ rapid amplification of cDNA ends PCR (3′Race-PCR) we have isolated a new fish steroid receptor cDNA sequence of 1806 bp from rainbow trout (Oncorhynchus mykiss) testis. This sequence has clear homology with various mineralocorticoid receptor cDNA sequences (rat, human, African toad: 68–70% amino acid identity), and encompasses the second part of DNA binding domain (C domain), the whole hinge region (D domain) and the steroid binding domain (E domain) plus 726 bp of 3′untranslated sequence. COS-1 cells transfected with a pCMV5 expression vector containing the whole E domain (pCMV5-rtMR) showed high affinity binding for cortisol ( K a = 0.53 ± 0.03 nM, K d = 1.9 nM) in the cytosol, which could not be detected in untransfected cells. Aldosterone displaced 3H-cortisol binding, though was less effective by than unlabeled cortisol ( P < 0.05). Competition experiments with other steroids gave the following hierarchy for the displacement of the 3H-cortisol from the receptor-ligand complex: cortisol = corticosterone = 11-deoxycortisol = 21-deoxycortisol > 11-deoxycorticosterone = 11β-hydroxyprogesterone = 17-hydroxyprogesterone > dexamethasone, whereas 17,20β-dihydroxy-4-pregnen-3-one and 17,20β,21β-trihydroxy-4 pregnen-3-one (two fish specific progestins) did not show any specific binding. These results strongly suggest that this cDNA sequence encodes a rainbow trout mineralocorticoid-like receptor, and represent the first description of such a receptor in teleost fish where aldosterone, the classic mineralocorticoid, is believed to be absent.