ABSTRACT
We have cloned and analysed the partial putative promoter sequences of the Yellowtail Kingfish (Seriola lalandi) Kiss2 and Kiss2r genes (380 and 420 bp, respectively). We obtained in silico 1.5 kb of the zebrafish (Danio rerio) Kiss1, Kiss2, Kiss1r and zfKiss2r sequences upstream of the putative transcriptional initiation site. Bioinformatic analysis revealed promoter regulatory elements including AP-1, Sp1, GR, ER, PR, AR, GATA-1, TTF-1, YY1 and C/EBP. These regulatory elements may mediate novel roles of the Kiss genes and their receptors in addition to their established role in reproductive function.
Subject(s)
Kisspeptins/genetics , Perciformes/genetics , Receptors, G-Protein-Coupled/genetics , Regulatory Elements, Transcriptional/genetics , Zebrafish/genetics , Animals , Base Sequence , Cloning, Molecular , Computational Biology , DNA Primers/genetics , Molecular Sequence Data , Polymerase Chain Reaction , Sequence Analysis, DNAABSTRACT
The kisspeptin system plays an essential role in reproductive function in vertebrates, particularly in the onset of puberty. We investigated the kisspeptin system in two Perciform teleosts, the Southern Bluefin Tuna (SBT; Thunnus maccoyii), and the Yellowtail Kingfish (YTK; Seriola lalandi), by characterising their kisspeptin 2 receptor (Kiss2r) genes. In addition to the full length Kiss2r cDNA sequences, we have isolated from SBT and YTK a transcript variant that retained an intron. We have further obtained three ytkKiss2r transcript variants that contained deletions. In vitro functional analysis of the full length SBT and YTK Kiss2r showed higher response to Kiss2-10 than to Kiss1-10, with stronger transduction via PKC than PKA. The full length ytkKiss2r and two deletion variants were differentially expressed in the brain of male, but not in female, juvenile YTK treated with increasing doses of Kiss2-10 peptide. In the gonads, the expression level of the ytkKiss2r transcripts did not vary significantly either in the male or female fish. This is the first time that transcript variants of the Kiss2r gene that contain deletions and show responsiveness to treatments with kisspeptin have been reported in any teleost.
Subject(s)
Fish Proteins/genetics , Receptors, G-Protein-Coupled/genetics , Tuna/genetics , Amino Acid Sequence , Animals , Base Sequence , Brain/metabolism , COS Cells , Chlorocebus aethiops , Cloning, Molecular , Female , Fish Proteins/metabolism , Gene Expression , Gonads/metabolism , Kisspeptins/pharmacology , Kisspeptins/physiology , Male , Molecular Sequence Data , Organ Specificity , Perciformes/genetics , Phylogeny , Protein Isoforms/genetics , Receptors, G-Protein-Coupled/metabolism , Sequence Alignment , Sequence Homology, Amino Acid , Signal Transduction , Tuna/metabolismABSTRACT
This paper reports the isolation of two putative D(2)R promoters from grey mullet, one 5'flanking and the other an intronic sequence immediately upstream of the first coding exon. Promoter activity of the intronic sequence was confirmed in vitro through functional analysis using luciferase as reporter gene. The functional characteristics of the region flanking the 5'UTR is currently under investigation.