The Roles Of Dicer-1 And Argonaute-1 In The Immune Responses Of The Pea Aphid,Acyrthosiphon Pisum

The pea aphid Acyrthosiphon pisum causes seriously damage to legumes and herbages as an important pest in agriculture and forestry.Using natural enemies and pathogenic microorganisms to control pea aphid has become a new research focus.It is the premise of the biological control to study the immune defense mechanism of pea aphid against pathogen infection.A large number of immune-related genes were missed in pea aphid,but the genes in the micro RNA(mi RNA)pathway in pea aphid were retained.Dicer-1 is an endonuclease that performs cleavage function during the synthesis of mi RNA,which is crucial for the synthesis process.Argonaute-1(Ago-1)is an important component of the RNA-inducing silencing complex(RISC)in the mi RNA pathway,which is closely related to the function of mi RNA.Studies have shown that Dicer-1 and Ago-1 play roles in insect immunity against bacterial infection.In order to investigate whether Dicer-1 and Ago-1 are involed in the immune response of the pea aphid against pathogen infection,gram-positive bacteria Staphylococcus aureus,gram-negative bacteria Escherichia coli and fungus Beauveria bassiana were used to infect the pea aphids.First,we detected the changes in the expression levels of Dicer-1 and Ago-1after bacterial and fungal infections.Next I silenced Dicer-1 and Ago-1 genes by RNA interference(RNAi).After the target genes was interfered,bacterial or fungal infection was carried out.And the proliferation of bacteria or fungi in pea aphid was detected,also the survival rate of pea aphid after infection was calculated.The main results are as follows:1.Protein domain analysis showed that Dicer-1 has one RNA helicase domain,one PAZ domain,two Rnase Ш domains and one double stranded RNA binding domain.Ago-1protein has one DUF 1785 domain,one PAZ domain and one Piwi domain.These domains are highly conserved.2.After infected with S.aureus,E.coli and B.bassiana repectively,the expression levels of Dicer-1 and Ago-1 were upregulated at different time points,indicating that Dicer-1 and Ago-1 were responsive to the infection by bacteria and fungi.3.After dsRNA injection,Dicer-1 and Ago-1 of pea aphid were mostly silenced,and the effect was significant and persistent.4.E.coli,S.aureus and B.bassiana were used to infect the Dicer-1 and Ago-1silenced pea aphids.Both bacterial cells and fungal spores increased significantly in the pea aphids.5.After knockdown of Dicer-1,and infection by S.aureus or E.coli,the survival rate of pea aphid was lower than the control group,and with a very significant difference when infected with E.coli;after infected with B.assiana,the survival rate of pea aphid decreased significantly compared to the control group.After knockdown of Ago-1,the survival rate of pea aphid was lower than the control group with a significant difference;after infected with S.aureus and E.coli,the survival rate was lower than that of the control group,and with a significant difference when infected with S.aureus;after infected with B.assiana,the survival rate of pea aphid decreased significantly compared to the control group.Conclusions: Dicer-1 and Ago-1 are all involved in the immune defense response of the pea aphid against bacterial and fungal infections.Both of them play a critical role in the anti-B.assiana response of pea aphid,and are involved in the immune response of pea aphid against S.areus and E.coli invasion in some degrees.These results further indicate that these mi RNAs depending on Dicer-1 and Ago-1 may play a potential regulatory role in the immune response of pea aphid against bacterial and fungal infections.