Effect Of ALDH1A1 Gene On Lipid Metabolism And Genetic Variation Analysis In Yanhuang Cattle

Aldehyde Dehydrogenase 1A1 is a member of the ALDH gene superfamily and is one of the key gene involved in the metabolism of retinal,the activation of retinoic acid metabolic pathway and the regulation of fatty acid metabolism pathway.In this study,the ALDH1A1 gene was constructed on the PMD18-T vector to obtain the cDNA sequence of the ALDH1A1 gene,and the physicochemical properties and protein structure of the gene were analyzed by bioinformatics software,and the expression of ALDH1A1 gene in various tissues of Yanhuang cattle was detected by qRT-PCR.The results showed that the ALDH1A1 gene of Yanhuang cattle encodes 501 amino acids,the theoretical isoelectric point is 7.16,which belongs to basic amino acids and plays a major role in the cytoplasm;It is a hydrophilic protein not a secreted protein,and there are 31 amino acid phosphorylation sites;The prediction results of the secondary and tertiary structures showed four structures,namely α-helix,β-sheet,β-turn and random coil.The expression of ALDH1A1 gene in liver,stomach,subcutaneous fat,duodenum and kidney tissues was extremely significant(P<0.01),and was significantly expressed in the longissimus dorsi muscle(P<0.05).The siRNA interference and overexpression vector of Yanhuang cattle ALDH1A1 gene was successfully constructed by molecular biological cloning technology,and then transfected into the isolated progenitor cells by liposome 2000,and the expression of ALDH1A1 mRNA and protein levels was detected by qRT-PCR and protein immunoblotting.The results showed that ALDH1A1,LPL and adipogenic key genes(C/EBPa and PPARy)showed a trend of increasing first and then decreasing when inducing differentiation of progenitor adipocytes.The ALDH1A1 gene interference group showed a trend of decreasing first and then increasing on the 0,4,and 9 days after induction of differentiation of preadipocytes,but the overexpression group showed a trend of increasing first and then decreasing on 0,4 and 9 days after induction of differentiation of preadipocytes;Overexpression of ALDH1A1 gene can significantly promote the formation of lipid droplets and triglycerides in mature adipocytes,and also significantly promote the expression of LPL and the key lipid-forming factors C/EBPa and PPARy,while the interference of ALDH1A1 gene is opposite,which significantly reduces the formation of fat cell lipid droplets and triglyceride content and significantly inhibits the expression of LPL and the key lipid-forming factors C/EBPa and PPARγ.This study demonstrates that the ALDH1A1 gene is involved in the process of differentiation of precursor adipocytes.In order to further explore the function of ALDH1A1 gene,this study used 9918-month-old Yanhuang cattle as the test object,and screened the polymorphic locus of ALDH1A1 gene by sequencing method,and then analyzed for its association with the meat traits of Yanhuang cattle.The results showed that only the mutation sites were detected in exons 4 and 11 among the 13 exons of the ALDH1A1 gene of Yanhuang cattle.Exon 4 has a T/G mutation at Chr9.49381142 and causes a change in the encoding amino acid tyrosine.Exon 11 has a T/A mutation at Chr9.49361243 that does not cause a change in the encoded amino acid.Intron 4 has a G/T mutation at Chr 9.493381356.The results of association analysis showed that the exon 4 of ALDH1A1 TT genotype was significantly correlated with the cross height,chest circumference,abdominal circumference,tube circumference and backfat thickness in Yanhuang cattle meat traits(P<0.05),and with intramuscular fat has a certain correlation(P<0.056);There was a significant correlation between the exon 11 of ALDH1A1 gene AA genotype and the cross height,official circumference,body weight and backfat thickness in Yanhuang cattle meat traits(P<0.05)In summary,this study successfully cloned the ALDH1A1 gene coding region sequence of Yanhuang cattle,and studied the expression difference of the gene in different tissues of Yanhuang cattle and its effect on lipid metabolism and genetic polymorphism,which provided theoretical basis and scientific basis for the function of the ALDH1A1 gene and the screening of meat gene in Yanhuang cattle.