Functional Analysis Of MAPEG Superfamily Members AccMGST1 And AccMGST2 In Apis Cerana Cerana

Apis cerana cerana is a unique bee species in China.It makes important contribution to the agricultural economic development and ecological stability.However,with the increasingly serious environmental problems,Apis cerana cerana has been suffering from damages caused by adversity,its survival has been severely tested,and the number of bee colonies has sharply decreased.Therefore,it is of great significance to explore the molecular mechanism of antioxidation to improve the survival ability and maintain the population development of Apis cerana cerana.MAPEG family is a superfamily that can mediate the synthesis of inflammatory factors.MGST1 and MGST2,members of it,play important role in lipid peroxidation protection and regulation of oxidative stress pathway,respectively.Thus,we studied the molecular mechanism and action mechanism of antioxidant under stress of AccMGST1,AccMGST2 through molecular biology technology.The main results are as follows:AccMGST1 and AccMGST2 genes were isolated for the first time by RT-PCR,their biological characteristics were analyzed by bioinformatics.The coding region sequence of AccMGST1 gene isolated by RT-PCR was 450 bp long,which encodes a peptide composed of149 amino acids,its predicted relative molecular weight of the coding protein was 17.143 kDa,the isoelectric point was 9.74,and its structure is a transmembrane trimer which was composed of ?-helices.The coding sequence of AccMGST2 gene is 462 bp in length and encodes a peptide composed of 152 amino acids,its predicted relative molecular weight of the coding protein was 17.517 kDa,the isoelectric point was 8.33,and its structure is also a transmembrane trimer which was composed of ?-helices.Amino acid sequence between AccMGST2 and AmMGST2 in Apis mellifera have high similarity,the similarity reached96.08%.The expression characteristics of AccMGST1 and AccMGST2 in different tissues were explored by qRT-PCR.The results showed that AccMGST1 gene was highly expressed in the epidermis but low in the head,abdomen and muscle while AccMGST2 gene was highlyexpressed in the head and muscle.The expression characteristics of AccMGST1 and AccMGST2 in different abiotic stresses were explored by qRT-PCR.qRT-PCR analysis showed that the expression of AccMGST1 and AccMGST2 can respond to oxidative stress caused by different abiotic stress.Western blot analysis showed that AccMGST1 can respond to oxidative stress caused by different abiotic stress at the protein level.Function of recombinant AccMGST1 protein was explored by bacteriostatic experiment and DNA protection experiment.The recombinant AccMGST1 protein not only help E.coli resist oxidative damage to cumene hydroperoxide but also reduce the oxidative damage to DNA double helix structure.Through RNA interference,AccMGST1 and AccMGST2 were silenced in Apis cerana cerana,their roles in antioxidation were analyzed.After silencing AccMGST1 gene in Apis cerana cerana by RNA interference,the expression level of related antioxidant genes were increased.After silencing AccMGST2 gene in Apis cerana cerana by RNA interference,qRT-PCR analysis revealed that the expression level of some MAPK genes and antioxidant genes were decreased.This study shows that AccMGST1 has the function of oxidative defense,and AccMGST2 is involved in the regulation of oxidative stress of Apis cerana cerana.