Cloning And Preliminary Study On Appetite Regulation Of PYY And POMC Gene In Siberian Sturgeon

Proopinmelanocortin(POMC)and peptide YY(PYY)is key central and peripheral factor in inhibition of food intake in central and peripheral of vertebrate.In order to study the molecular level of Siberian sturgeon appetite suppression,we employed PCR,cloning and RACE for the first time obtained the whole sequence of PYY and partial sequence of POMC-A and POMC-B of Siberian sturgeon.In order to study the role of PYY in the regulation of food intake,we employed real-time PCR technology to detect the tissue expression of PYY the effects of preprandial(pre-and post-feeding)and fasting of its expression.To study the influence of the mature peptide of PYY and POMC,tests were used PYY and a-MSH as the injection peptides,the effect on food intake was detected by a intraperitoneal injection.In this study,we first characterized a high conservation of PYY and found high expression levels of PYY mRNA in the brain and digestive tract of Siberian sturgeon.Then,we examined preprandial(pre-and post-feeding)changes of PYY mRNA expression in the brain and duodenum that showed a significantly increased in 3h post-feeding,suggesting an anorectic possible function of PYY in siberian sturgeon.Next,we examined the expression of PYY mRNA during 15 days fasting and refeed after fasting.The SsPYY mRNA expression of unfed fish had a significant 2.4,1.7,2.0,2.2,and 2.1-fold decrease compared to 1-,3-,6-,10-and 15-day ad libitum fed animals,respectively.After refeed,SsPYY mRNA significantly increased 1.9 and 4.1-fold above that of the 15-day fed and unfed fish control group.Food-deprived Siberian sturgeon had a significant difference in mRNA expression levels,that is 2.5-fold lower in the duodenum of PYY than 1-,3-,6-,10-and 15-day regularly fed controls.When 15-day fasted fish were re-fed,PYY mRNA in the duodenum was significantly increased.Furthermore,the levels of 15-day fed control group and fasted group of Siberian sturgeon respectively had an approximate 1.7-and 4.3-fold lower than 15-day refed fish.A single intraperitoneal injection of 10,100 and 200 ng/g BW SsPYYl-36 caused a reduction in the next feeding and no significant reduction in food intake was observed in fish injected with a 1 ng/g BW.It showed that PYY can inhibit the Siberian sturgeon feeding through periphery system.This test has been identified to get Siberian sturgeon POMC gene in two subtypes of POMC-A and POMC-B.The length of the Siberian sturgeon POMC-A and POMC-B fragments were 805bp and 1119bp.Sequence analysis showed that the POMC-A and POMC-B covered all of the ORF,its deduced amino acid sequence of POMC-A and POMC-B can theoretically produce ?-MSH,?-MSH,Clip,?-LPH,?-MSH and?-endorphin.However the cleavage site of ?-LPH and ?-MSH appears different between POMC-A and POMC-B.At the same time,sequence analysis also found that,compared with other sturgeon class,Siberian sturgeon ORF box of the POMC-B appeared a 312bp inserts howed that of POMC-A and POMC-B may have sub-functional.Furthermore,a single intraperitoneal injection of 1,10,100 and 200 ng/g BWa-MSH caused a reduction in the nextlh feeding.It showed that POMC can inhibit the Siberian sturgeon feeding through periphery system.In summary,Siberian sturgeon PYY and POMC is a regulator of appetite suppressant,this test PYY and POMC cleavage of a-MSH peptide can inhibit food intake through the peripheral injection showed that PYY and POMC can inhibit the Siberian sturgeon feeding through periphery system.This study provides an essential groundwork to further elucidate the appetite regulatory systems of PYY and POMC in teleosts and enriches fish feeding genes library.