Molecular Mechanism Of Myzus Persicae (Hemiptera:Aphididae) In Response To UV-B Stress

Myzus persicae(Sulzer)(Hemiptera: Aphididae)is a worldwide pest that seriously harms >400 plants such as tobacco,cruciferous vegetables,peppers,and melons.As an important environmental stress factor,UV-B radiation is unavoidable for M.persicae,which live in direct sunlight all year round.However,it is still unclear that the molecular mechanism of response to UV-B stress in M.persicae.In this study,8 MpHsp70 s genes(MpHsp70,MpHsp70-1,MpHsp70-2,MpHsp70A1,MpHsp70B2,MpHsc70-4,MpHsp68 a,and MpHsp68b)were cloned in full length and analyzed the expression levels under different developmental stages and different durations of UV-B stress,so the role of Hsp70 family members were explored in the response of M.persicae to UV-B stress;the full-length sequences of four important genes in the MAPKs signaling pathway(MpJNK,Mpp38,MpERK1,and MpERK2)were cloned,and their expression patterns under different development and different durations of UV-B stress were analyzed;based on nanomaterial-mediated RNAi technology,we studied their function in M.persicae in response to UV-B stress,and explored the role of MAPK signaling pathway in insect response to UV-B stress.The results of this study laid the foundation for further exploring the molecular mechanism of M.persicae in response to UV-B stress.The corresponding results are as follows:1.Cloning and expression of MpHsp70 s and its response to UV-B stressThe 8 MpHsp70 s genes(MpHsp70,MpHsp70-1,MpHsp70-2,MpHsp70A1,MpHsp70B2,MpHsc70-4,MpHsp68 a and MpHsp68b)sequences were cloned by RT-PCR and RACE technology,which code 654,638,639,641 respectively,624,662,647,and 660 amino acids,and the amino acid terminal sequences are all EEVD,indicating that these proteins have the amino acid characteristics of cytoplasmic heat shock proteins,and all belong to cytoplasmic heat shock proteins.The relative molecular weight of the encoded protein ranged from 68.35 to 71.99 kD.All 8Mphsp70 s contained 3 Hsp70 family tag sequence features,indicating that the cloned genes all belong to the Hsp70 family.Phylogenetic analysis showed that MpHsp70-1,MpHsp70-2,MpHsp70A1,MpHsp70B2,MpHsp68 a,and MpHsp68 b belong to inducible Hsp70,and MpHsp70 and MpHsc70-4 belong to constitutive Hsc70.The expression patterns of eight Mphsp70 s genes at different developmental stages and under different long UV-B stresses were detected by fluorescent quantitative PCR.The results showed that the expression patterns of 8 MpHsp70 s genes at different developmental stages of M.persicae were different,indicating that they were in during the growth and development of M.persicae,they all drive different functions.Under UV-B stress,the expression of the 8 MpHsp70 s genes of M.persicae increases significantly,and all show a trend of first increasing and then decreasing with time,respectively at 30,120,90,60,120,60,120,and 60 min reached the maximum expression level,which was 2.63,102.09,416.35,346.70,13.39,12.72,1355.41,and212.63 times that of the control.The differential expression of 8 Mphsp70 s at different ages and under UV-B stress indicated that they played an important role in the molecular mechanism of response to UV-B stress.2.Cloning and expression of MAPK signaling pathway genes and its response to UV-B stressThe gene sequences of M.persicae MpJNK,Mpp38,MpERK1,and MpERK2 were cloned by RT-PCR and RACE technology.Their protein sequences were 397,633,438,and 361 amino acids,respectively,and the relative molecular weights of the encoded proteins were 45.588,42.023,48.271,and 41.424 kD.Phylogenetic analysis showed that MpJNK,Mpp38,MpERK1,and MpERK2 have high homology with other insects.Fluorescence qPCR was used to detect the expression patterns of MpJNK,Mpp38,MpERK1,and MpERK2 at different developmental stages of M.persicae.The results showed that: MpJNK and MpERK2 expressed the highest in the wingless adult stage,Mpp38 expressed the highest in the fourth instar nymph stage,and the expression of MpERK1 is highest in the first instar nymph stage and the wingless adult stage.Under UV-B stress,the expression levels of MpJNK,Mpp38,MpERK1,and MpERK2 all increased significantly,and the expression levels of MpJNK,Mpp38,and MpERK2 genes showed a trend of first increasing and then decreasing with time,and all reached their peaks at 60 min,which were 3.70,2.08,and 7.19 times that of the control,respectively;the expression of the MpERK1 gene showed an increasing trend with time,reaching the maximum at 150 min,which was5.04 times that of the control.This study showed that the expression levels of MpJNK,Mpp38,MpERK1 and MpERK2 of M.persicae under UV-B stress were up-regulated,and the JNK,p38,ERK1 and ERK2 signaling pathways were activated to respond to UV-B stress.This is to further study the response of M.persicae to UV-B stress.The molecular mechanism provides clues.3.The role of M.persicae MAPK signaling pathway in response to UV-B stressThe interference treatment of 1-day-old wingless adults of M.persicae with MpJNK,Mpp38,MpERK1,and MpERK2 genes was performed by the nanocarrier mediated ds RNA spot assay,and the target gene expression was measured at 24,48,and 72 h,respectively.The results showed that the expression level of MpJNK gene decreased by 42.83%,64.96%,and 17.37% at 24,48,and 72 h,respectively,indicating that the expression of MpJNK gene was significantly suppressed at 24 and48 h;the expression level of Mpp38 gene,the decrease of 86.24% and 46.23% at 48 and 72 h indicates that the expression of Mpp38 gene was significantly suppressed at48 and 72 h,respectively;the expression level of MpERK1 gene decreased by24.16%,50.90%,and 56.89% at 24,48 and 72 h,respectively,indicating that the expression of MpERK1 gene was significantly inhibited at 24,48,and 72 h,and the decline was most obvious at 72 h;the expression level of MpERK2 gene decreased at24,48,and 72 h,respectively 96.95%,92.89%,and 86.04%,indicating that the expression of MpERK2 gene was significantly inhibited at 24,48,and 72 h.MpJNK,Mpp38,MpERK1,and MpERK2 experiments based on RNAi technology showed that after interfering with Mpp38,MpERK1,and MpERK2 genes,the survival rate of wingless adults of M.persicae under UV-B irradiation was significantly lower than that of the control group(GFP)(P< 0.0001);the total number of nymphs produced,the total number of nymphs produced per unit and the average lifespan were significantly reduced(P<0.05).The results showed that Mpp38,MpERK1,and MpERK2 genes play an important role in the response of M.persicae to UV-B stress.This paper studied the cloned 8 MpHsp70 s genes and 4 MAPK signaling pathway-related genes(MpJNK,Mpp38,MpERK1,and MpERK2)sequences are full-length,and their relative expression changes in different developmental stages and different durations of M.persicae under UV-B stress were detected;RNAi technology was used to study the roles of MpJNK,Mpp38,MpERK1,and MpERK2 genes in response to UV-B stress of M.persicae.The results revealed the role of 8MpHsp70 s genes and 4 MAPK signaling pathway-related genes in the response of M.persicae to UV-B stress,laying a foundation for further exploration of the molecular mechanism of insect response to UV-B stress.