Analysis On Gagalaxin Function Of Galaxea Astreata

It is very important to study the mechanism of coral calcification and bleaching to protect and restore coral reefs.In this study,PCR and RACE technology were used to amplify the total length of Gagalaxin mRNA of Galaxea astreata.After the protein was expressed in vitro using the prokaryotic expression technology,the proteins interacting with Gagalaxin were detected by pull down technology,and the function of Gagalaxin binding protein were analyzed to infer the function of Gagalaxin using bioinformatics.Secondly,the Gene Expression Omnibus(GEO)database was excavated to understand the influence of high temperature on coral bleaching by using Differential Expression Genes(DEGs)during coral bleaching process.Using the protein–protein interaction(PPI)network to analyze the relationship between DEGs,Gagalaxin and Gagalaxin binding proteins.This study aims to investigate the role of the Gagalaxin in the process of coral calcification and bleaching.The results are as follows:1.The total length of Gagalaxin mRNA(GenBank no.MF063014.1)was 1492 bp,including 41 bp 5' UTR,419 bp 3' UTR,1032 bp CDS,encoding 343 amino acids and rich in dicysteine repeat.It is predicted that the Gagalaxin protein has two internal repeat structures,and the N-terminal 1-22 AA was the signal peptide region.One intron(A length of 2100 bp)was detected within the initiator codon and PolyA of Gagalaxin.The result of multiple sequence alignment showed that Gagalaxin exhibited little variation in intra-genera but greater variation in inter-genera(Acropora and Galaxea)2.The Gagalaxin sequence without signal peptide region was successfully expressed after recombination with Glutathione S-transferase(GST)tag.The recombinant protein expression quantity increased significantly after induced by final concentration of 0.5 mM of Isopropyl ?-D-Thiogalactoside(IPTG)(37 ? for 4 h).3.The results of pull down experiment showed that there were some proteins binding with Gagalaxin obviously.After mass spectrometry identification,2 out of 8 Gagalaxin binding proteins matched the G.astreata transcriptome.These two proteins were Suppressor of Mek1(Might be derive from Symbiodinium microadriaticum)and Coadhesin-like protein.4.A total of 107 DEGs were detected in GSE16151,37 were detected between the control groups and slight bleaching groups,and 98 were detected between the controlgroups and extreme bleaching groups.These DEGs were mainly related to cell signal/transcription,cytoskeleton/extracellular matrix protein,DNA replication/cycle/proliferation,and Ribosome.Suppressor of Mek1,Gagalaxin and Coadhesin-like protein were not successfully matched in STRING.And DCAF13,DKC1 and CCT7 proteins were located in key locations of the Protein-protein Interaction network.