Relationship Of GRP78 And MC5R With Productive Performance In The Geese With Fatty Liver

Goose fatty liver(commonly called as foie gras)is a nutritious and high-grade food.There are significant differences in the performance of foie gras production among different breeds of geese.This means that the performance can be improved by genetic selection.Currently,goose breed often used in the industry of foie gras in China is the Landes goose from abroad.To make a profit by avoiding high import cost of this breed,many enterprises prefer to purchase Landes geese with germplasm degeneration as the breed undergoes through several passages without efficient selection after import.The productivity and economic gain both have been severely impaired by this situation.An effective solution is to make a genetic selection on the breed after import However,current selection method is not efficient as it is only based on goose phenotype.Compared to the phenotypic selection,the genetic marker-assisted selection is more advantageous.This study was aimed to assessing whether GRP78 and MC5R are closely related to fatty liver formation and whether their polymorphic loci are associated with relevant productive traits in the fatty liver-producing geese,thus this study could provide a basis for marker-assisted selection.The major results of this study are as follows:1.Quantitative real-time PCR was used to determine the expression of GRP78 gene in different tissues(liver,abdominal fat and pectoral muscle)of the geese that were conventionally fed(the control group)or overfed(the overfeeding group)for 7,14 and 19 days.The results showed that the mRNA expression level of GRP78 gene in the livers of the overfed geese was lower than that in the control geese on the 7,14 and 19 days of overfeeding,but the difference in GRP78 expression between the groups was not statistically significant(P>0.05).On the contrary,the expression level of GRP78 mRNA in abdominal fat of the geese in the overfeeding group was higher than that in the control group,and the difference reached statistical significance until 14 days of overfeeding(P<0.05).However,there was no significant difference in mRNA expression of GRP78 gene in breast muscle between the two groups.Furthermore,the addition of sucrose to the feed resulted in further suppression of GRP78 expression in fatty liver,accompanied by an increase in fatty liver weight and abdominal fat weight.This result indicated that GRP78 could be involved in the formation of goose fatty liver,and its inhibited expression may promote the formation of fatty liver.Therefore,GRP78 was a good candidate gene for assistant selection marker in the fatty liver-producing goose.To assess the feasibility of using GRP78 gene as an assistant selection marker,the GRP78 gene sequence and its upstream sequence(<2.5 kb)were amplified by PCR and sequenced to screen for the presence of SNP sites.The results showed that there were only 6 SNPs in the upstream sequence far from the start codon of the gene(-1904 bp to-2484 bp),and there was no significant correlation between these sites and the relevant productive performance in the geese with fatty liver,and no other SNPs were identified in the coding sequence and intronic sequence of GRP78.The absence of a polymorphic site in the GRP78 gene sequence suggested that the GRP78 gene was subjected to a greater selection pressure due to the importance of its biological function.2.Quantitative real-time PCR was used to determine the expression of MC5R gene in different tissues(liver,abdominal fat and pectoral muscle)of the geese in the control group and the overfeeding groupon the 7,14 and 19 days of overfeeding.The results showed that the expression of MC5R gene was significantly lower in the livers of the overfed geese on the 7,14 and 19 days of overfeeding than that in the control group.The inhibition of MC5R became more obvious with the increasing amount of fat deposition in the liver.Similarly,the expression of MC5R gene in abdominal fat was also significantly lower in the overfeeding group than that in the control group.For pectoral muscles,there was no significant difference between the groups on the 7 days of overfeeding.After 14 days of overfeeding,the expression of MC5R in the overfeeding group was lower than that in the control group,while the difference between the groups was not statistically significant;and after 19 days of overfeeding,the expression of MC5R in the geese of the overfeeding group was significantly lower than that in the control group,and the inhibition was intensified.These results suggested that the inhibition of MC5R expression may promote fat deposition in the tissues,and therefore the MC5R gene is a good candidate gene for assistant selection marker in the fatty liver-producing geese.3.In order to better understand the relationship between MC5R and goose fatty liver formation,this study determined the effects of fatty liver-related factors(high-dosage of glucose,insulin,and fatty acids)on the expression of MC5R gene in goose primary hepatocytes.The hepatocytes were treated with high-dosage of glucose,insulin,palmitic acid,oleic acid,and linoleic acid for 14 hours,respectively.Data indicated that glucose and insulin induced the expression of MC5R in variable degrees in the treated cells,whereas fatty acids did not obviously affect the expression of MC5R.These results were inconsistent with the expression pattern of MC5R in goose fatty liver.It is speculated that there might have other factors or mechanisms that could affect the expression of MC5R gene in goose fatty liver.4.To investigate the possibility of MC5R being an assistant selection marker in fatty liver-producing geese,this study performed sequencing analysis on MC5R coding sequence to find SNP sites,and association analysis of SNPs with productive performance(liver weight,abdominal fat weight,and carcass weight).The data showed that a C/T polymorphic locus(807bp downstream the start codon)was found,and the polymorphism did not change the amino acid sequence of MC5R.This SNP was associated with abdominal fat weight in the geese with fatty liver.The average abdominal fat weight of the geese with CT genotype was significantly greater than that with CC genotype(P<0.05),the difference was 52.67 g;and the average abdominal fat weight of the geese with TT genotype was not significantly different from those with the CC and CT genotypes;the SNP locus was also associated with the carcass weight of the geese with fatty liver.The average abdominal fat weight of the geese with CC genotype was significantly greater than that with CT genotype(P<0.05),and the difference was 630g;and the average abdominal fat weight of the geese with CT genotype was not significantly different from those with the CC and TT genotypes.However,there was no obvious association of the SNP with liver weight in the geese with fatty liver.The results from association analysis suggested that MC5R could be used in the marker assisted selection for the fatty liver-producing geese,ie.,selection on the abdominal fat weight and body weight in the geese with fatty liver without significantly affecting the liver weight.In summary,by making clear the expression profiles of GRP78 and MC5R genes in different tissues of Landes geese along with the time of overfeeding,the effects of supplemented sugar into diet and fatty liver-related factors on gene expression,as well as the association of SNPs with the productive performance in the fatty liver-producing geese,this study suggests that the inhibition on the expression of GRP78 and MC5R genes in the liver contributes to the formation of geese fatty liver,and both of the genes are good candidate gene for marker-assisted selection in the fatty liver-producing goose.The SNP identified MC5R coding sequence may be used as a genetic marker for selection on abdominal fat weight and carcass weight in the geese with fatty liver.