Molecular characterization of GPR50 gene and study of its comparative genetic variability in sheep breeds adapted to different thermo-contrasting climatic regimens.
GPR50, formerly known as a melatonin-related receptor, is one of the three subtypes of melatonin receptor subfamily, together with MTNR1A and MTNR1B. GPR50, despite its high identity with the melatonin receptor family, does not bind melatonin and is considered to be an ortholog of MTNR1C in mammals. GPR50-expressing cells have been found in the dorsomedial nucleus of the hypothalamus, the periventricular nucleus, and the median eminence. Genetic and functional evidence have been recently investigated linking GPR50 to adaptive thermogenesis and torpor, but still, it is an orphan receptor and is yet to be studied conclusively. The aims of the study were to characterize the GPR50 gene of sheep and to study the sequence variability of the gene in Indian sheep breeds of two different thermo-varied agroclimatic conditions. Genomic DNA isolation was done and a 791-bp sequence was amplified using self-designed primers and SNP profiling done out of samples of all the breeds to study the relative frequency of SNPs in each of the breed. Five important non-synonymous mutations were observed in the various breeds studied. T698G, G1097A, G1270A, G1318A, and C1334G lead to the following substitution: valine by glycine, arginine by glutamine, threonine by alanine, isoleucine by valine, and serine by cytosine, respectively. Two synonymous mutations (T663G and C888T) were also observed in some of the studied breeds. G1270A and C888T were the most prevalent SNPs observed in nearly all of the breeds. C888T SNPs were observed in higher prevalence in Chokla, Marwari, and Magra in comparison to Gaddi and Bharat Merino. A PolyPhen-2 analysis, which is used to assess the potential damaging nature of an SNP, revealed that mutation T698G and G1270A were benign while G1097A, G1318A, and C1334G were damaging with a score of 0.987, 0.993, and 0.739, respectively. A 3-D homology model of the protein was prepared using c4zwjA (UniProt sequence ID) as a template using the online version of Phyre2 protein modeling software. The structure demonstrated closed similarity with other G-coupled receptor and it had a 45 % α-helical content. G1270A and C888T may be taken up for SNP correlation in a larger population study for their association with heat stress protection.
Molecular Physiology Laboratory, Division of Animal Physiology and Biochemistry, ICAR-Central Sheep and Wool Research Institute, Avikanagar, 304501, Malpura, Tonk Distt, Rajasthan, India. drvijaysaxena@gmail.com.
Vijay Kumar
VK
Saxena
Molecular Physiology Laboratory, Division of Animal Physiology and Biochemistry, ICAR-Central Sheep and Wool Research Institute, Avikanagar, 304501, Malpura, Tonk Distt, Rajasthan, India.
Davendra
D
Kumar
Molecular Physiology Laboratory, Division of Animal Physiology and Biochemistry, ICAR-Central Sheep and Wool Research Institute, Avikanagar, 304501, Malpura, Tonk Distt, Rajasthan, India.
S M K
SM
Naqvi
D000222:Adaptation, Physiological; D000818:Animals; D002980:Climate; D008958:Models, Molecular; D009419:Nerve Tissue Proteins / Q000737:chemistry / Q000235:genetics*; D020641:Polymorphism, Single Nucleotide; D011487:Protein Conformation; D043562:Receptors, G-Protein-Coupled / Q000737:chemistry / Q000235:genetics*; D012756:Sheep / Q000235:genetics* / Q000502:physiology
D009419:Nerve Tissue Proteins; D043562:Receptors, G-Protein-Coupled
Endocrinology. 1996 Aug;137(8):3469-77
8754776
Curr Biol. 2012 Jan 10;22(1):70-7
22197240
Nat Protoc. 2015 Jun;10(6):845-58
25950237
Neuron. 1994 Nov;13(5):1177-85
7946354
Neuron. 1995 Nov;15(5):1003-15
7576645
J Neuroendocrinol. 1998 Sep;10(9):651-61
9744482
Front Biosci. 2003 Sep 01;8:d1093-108
12957828
Endocrine. 2005 Jul;27(2):101-10
16217123
Genet Mol Res. 2015 Jan 30;14(1):680-90
25730005
FEBS Lett. 1996 May 20;386(2-3):219-24
8647286
Reprod Domest Anim. 2015 Jun;50(3):517-21
25708518
Biomed Pharmacother. 2016 Mar;78:121-8
26898433
BMC Evol Biol. 2008 Apr 09;8:105
18400093
J Neuroendocrinol. 2012 May;24(5):798-808
22512326
J Neuroendocrinol. 2001 May;13(5):453-8
11328456
Proc Natl Acad Sci U S A. 1994 Jun 21;91(13):6133-7
7517042
Exp Physiol. 2008 Jul;93(7):773-97
18469069
EMBO J. 2006 Jul 12;25(13):3012-23
16778767
Curr Biol. 2012 Jan 10;22(1):R17-8
22240472
Mol Endocrinol. 1996 Nov;10(11):1478-87
8923472
J Genet. 2015 Dec;94(4):715-21
26690527
Meta Gene. 2014 Jun 28;2:450-8
25606429
Biol Reprod. 2000 Apr;62(4):1096-101
10727283
Am J Physiol Regul Integr Comp Physiol. 2011 Nov;301(5):R1207-28
21900642