Preliminary Study On The Molecular Mechanism Of Low Temperature Adaptability Of Pinctada Fucata Martensii

Pinctada fucata martensii is the pearl shell of main coastal water of shellfish aquaculture in China,belonging to warm water shellfish.P.f.martensii has weak tolerance to low temperature,which limits its breeding area and thus affects the development of the seawater pearl industry.The exploration of the potential low-temperature resistance of P.f.martensii will help to select low-temperature-tolerant varieties specifically,and lay a foundation for the expansion of aquaculture in northern China.Based on the transcriptome data of P.f.martensii cold-resistant breeding population F1(SP)and control population(BP),we selected candidate genes for temperature adaptation and performed validation,cloning,and functional analysis.The main research results obtained are as follows:(1)The results of transcriptome analysis showed a total of 572 differentially expressed genes in the two populations(|log2 Ratio(BP/SP)|>1 FDR<=0.001).We found that HSP40 and HSP70 show different expression patterns in HSPs system,indicating that different HSP plays different roles in the temperature adaptation of P.f.martensii.Small amino acid systems,energy release from oxidative phosphorylation-mediated energy release and increased biosynthesis of unsaturated fatty acids may help P.f.martensii cope with low temperature stress.Some lysosomal proteins and immune-related genes are also highly expressed in SP.This suggests that the modest autophagy and immune enhancement mediated by lysosomes can help the survival of P.f.martensii in low temperature environment.These results show that the multi-system co-participates in the low temperature adaptation process of the pearl shell,which provides new insights for the recognition of shellfish adaptation to the temperature change.(2)Eight temperature-adapted candidate genes were screened based on transcriptome data(heat shock proteins 40,Pm-HSP40;heat shock proteins 70,Pm-HSP70;hexokinase,Pm-HK;phosphoenolpyruvate carboxykinase,Pm-PEPCK;delta 6-fatty acid desaturase,Pm-?6FAD;stearoyl-Co A desaturase,Pm-SCD;Cathepsin L,Pm-CTSL;Cathepsin C,Pm-CTSC),and qRT-PCR technology was used to verify whether it is temperature adaptive genes.The results showed that the selected timed expression of the 8 temperature-adapted candidate genes was significantly correlated with temperature(P<0.05)in the gill of P.f.martensii,among which Pm-HK,Pm-PEPCK Pm-?6FAD,Pm-SCD,Pm-CTSL and Pm-CTSC may play an important role in the cold stress of P.f. martensii;Pm-HSP70 may play an important role in P.f.martensii under high temperature stress;Pm-HSP40 in P.f.martensii may be plays an important role in both low temperature and high temperature stress.It was confirmed that the eight candidate genes selected were temperature-adapted related genes.(3)The full length cDNA sequence of 8 temperature-adaptation-related genes(Pm-HSP40,Pm-HSP70,Pm-HK,Pm-PEPCK,Pm-?6FAD,Pm-SCD,Pm-CTSL,Pm-CTSC)in P.f.martensii were obtained by RACE technology.The cDNA sequence was full-length and bioinformatics analysis of ORF,amino acid sequence,protein domain,transmembrane domain,tertiary structure prediction and homologous comparison was performed using software.qRT-PCR analysis showed that 8 genes were expressed in the whole tissue and the differences were significant(P<0.05),of which Pm-HSP40,Pm-HSP70,Pm-HK,Pm-PEPCK,Pm-CTSL and Pm-CTSC expressed the highest in the hepatopancreas,while Pm-?6FAD and Pm-SCD m RNA were expressed in the gonad.It is speculated that hepatopancreas play an important role in the adaptation to low temperature of P.f.martensii.Whereas Pm-?6FAD and Pm-SCD significantly expressed in the gonads which may be related to their involvement in the gonadal development of P.f.martensii.