ABSTRACT
Arch BiochemMol Biol2019;10(4):035-051DOI: 10.26502/abmb.007Archives of Biochemistry and Molecular BiologyVol. 10 No. 4 -December2019. 36AbstractArgonaute2 (AGO2) is a core catalytic component of the RNA-induces silencing complex (RISC) that binds to small guide RNAs containing small interfering RNA (siRNA) and microRNA (miRNA). The guide RNA leads RISC to the complementary mRNA for gene suppression. We cloned the full length cDNA (2193 bp) of the Ago2gene (PxAgo2) from diamondback moth (DBM, Plutella xylostella). The predicted PxAgo2 protein hadan83 kDa molecular weight withatheoretical isoelectric point of 9.39. The phylogenetic tree showed a high similarity of PxAgo2with Bombyx moriAgo2 (BmAgo2). Western blot and RT-qPCR analyses showed a clear increase in the PxAgo2 mRNA and protein expression levels in the egg, 4thinstar larva, pre-pupa, pupa and adult. The double-stranded RNA-mediated RNAi of PxAgo2in DBM larvae was found 3 h after dsRNA injection,and the knockdown level was increased over time up to 36 h. PxAgo2silencing recovered the expression of PxBurs-?(Bursicon-? inDBM) to the normal expression level, which was suppressed bydsPxBurs-?in a DBM cell line. The overexpression of PxAgo2fundamentally enhanced the PxBurs-? silencing efficiency in DBM cells. Our findings reveal that PxAgo2is involved in the dsRNA-regulated gene silencing mechanism and performs a crucial function in the RNAi process ofDBM