Sequence analysis of retinoic acid receptor α, β and γ isoforms in the lizard, Podarcis sicula

Vitamin A and its principal biologically active derivative, retinoic acid (RA), play a fundamental role in diverse processes, such as proliferation, differentiation, morphogenesis, metabolism and apoptosis of many types of cells. In addition, RA has been shown to be involved in the regulation of testicular function. These effects are mediated by interaction with two families of nuclear receptors, retinoic acid receptor (RAR) and retinoid X receptor (RXR), each with three subtypes α, β and γ. The physiological involvement of retinoids in testicular function has been conducted mainly in mammals. Recently, we found that exogenous all-trans-retinoic acid impairs spermatogenesis and enhance testicular germ cell apoptosis in the lizard, Podarcis sicula, a seasonal breeder. To further investigate the role of retinoic acid in lizard, we focus this work principally on the characterization of lizard retinoic acid receptors (α, β and γ isoforms).RARα is 2720 bp long with a putative ORF between 699 and 2133. A Kozac sequence is present at 696 and a putative poly-adenilation site is present in position 2612. The RARα sequence shares 87% homology with mouse RARα mRNA while it has 76 and 80% homology with lizard RARβ and γ mRNAs. RARβ is 2478 bp long showing a putative ORF between 196 and 1543. A canonical Kozac sequence is present at 193 and a putative poly-adenilation site is present at 2294. RARβ shares 91% homology with mouse RARβ mRNA and has 76% homology with both RARα and γ. RARγ is 2416 bp long. With a putative ORF between 444 and 1818. A Kozac sequence is present at 441 and a putative poly-adenilation site is present at 2288. RARγ shares 86% homology with mouse RARβ mRNA and has 80 and 76% homology with both RARα and β respectively. It is worth to note that, as in mouse, the 5′UTR of all isoforms is TATA and CAAT less. Both Northern blot and PCR analyses indicate that lizard testis expresses only RARα and RARβ mRNAs, while RARγ mRNA transcript was not found. In the period analysed, RARβ was expressed during the gonadal full activity (May) and RARα was present in the post-reproductive period (August). During the autumnal recrudescence (October) RARα and RARβ are co-expressed and, as indicated by quantitative PCR analysis, RARβ mRNA levels are lower than RARα ones. Thus, the appearance and abundance of each receptor correspond to a specific phase of lizard reproductive cycle, allow us to hypothesize that each RAR subtype could play a specific role in the regulation of spermatogenetic activity.The results of the present study show, for the first time, the characterization of RAR mRNAs in the testis of lizard P. sicula, whose expression is related to the different phase of reproductive cycle. Moreover, the γ form, is principally expressed in the skin during the March–July period, having probably a role in regulating skin homeostasis and colour livery, which are important factor in mating during the reproductive cycle.