The Study On The Origin And Evolution Of Vertebrate Apolipoprotein Gene And The Antibacterial Function Of Lamprey LAL2

Apolipoprotein(APO),an important component of plasma lipoprotein,which is mainly synthesized in the liver and small intestine,can combine with and transport blood lipids to various tissues of the body for metabolism and utilization.In addition,apolipoproteins also play a role in the defense of pathogens.In the long history of evolution,little is known about the evolution of the apolipoprotein multigene family and its original functional characteristics.Therefore,it will be of great significance of the identification of the apolipoprotein molecules LAL1 and LAL2 in lamprey to the evolution of the vertebrate apolipoprotein family molecules.Furthermore,that exploring the structure and function of the lamprey apolipoprotein molecule LAL2 will not only revivificate the most primitive feature of apolipoprotein,but also lay the foundation for the study of lamprey serum immunology.The main contents of this paper include the following aspects:The lal2 gene was expressed in HEK293 T cells by cell transfection,the analysis of eukaryotic transcriptome sequencing and transcriptome data results were completed.By differentially expressing gene clustering,differentially expressing gene functions were enriched and analyzed by GO classification,KOG classification and KEGG classification.The potential biological functions of LAL2 were predicted,the results indicated that LAL2 may play an important role in the growth and development of lamprey,participate in the immune response of pathogen invasion and lipid synthesis and metabolism.Lamprey plasma apolipoproteins LAL2 and LAL1 were found in the genome database of Lethenteron reissneri.LAL2,LAL1 and the apolipoprotein A,C,E family amino acid sequences of other species were analyzed by colinearity,phylogenetic tree and motif using Genomicus online website,MEGA7.0 software,and MEME online website,which proved lampreys LAL2 and LAL1 are the ancestors of vertebrate apolipoproteins.However,LAL2 is in the original evolutionary position relative to LAL1,its motifs 4 and 5 from Branchiostoma belcheri APOA1,which are inserted into another Scaffold to form lal1 gene by transposition and the loss of motif 5.It is speculated that the ancestral apolipoprotein genes may be subjected to different selection pressures at the same time during the early differentiation process,and the evolutionary order may be: LAL2 / LAL1-APOA1 / APOA4 /APOE-APOA2 / APOC1 / APOC2-APOC3 / APOA5-APOC4.It was found that the level of LAL2 protein in serum was significantly up-regulated after S.aureus,V.anguillarum,and Poly I: C immunization,proving that LAL2 is involvedimmune response of bacteria and viruses.Scanning electron microscopy images showed that some S.aureus and E.coli cells were cracked,contents were leaked out,and death were occurred after incubation with LAL2.Western blot and ELISA results showed that LAL2 could bind to S.aureus,B.cereus,and E.coli,and interact with surface components LTA,LPS,PGN,and MDP.This study certifies that the vertebrate apolipoprotein A,C,E gene family are formed through the gene replication and recombination of LAL1,LAL2 in lamprey,which not only help in drawing the evolutionary history of the vertebrate apolipoprotein multigene family,but also provide new insight into the important and diversified functional properties of apolipoprotein.It also revealed that lamprey LAL2,as an effector molecule in sera,participates in immune response,pattern recognition,and antibacterial processes,laying a foundation for the exploration of the sera immune defense system of lamprey.