Expression analysis of host defense responses against the 8K (KDa) cysteine-rich viral silencing suppressor protein in Nicotiana benthamiana

Abstract

Potato mop-top virus (PMTV) encodes the 8K cysteine-rich viral suppressor of RNA silencing. To gain an insight into N. benthamiana defense mechanisms against 8K suppressor protein, we expressed two 8K suppressors from Peruvian isolates in Nicotiana benthamiana and assessed the expression of its defense genes involved in autophagy (ATG6, ATG2 and ATG7, AGO1), salicylic acid (SA) (ICS1, NPR1 and PR1) and jasmonic acid (JA) (OPR3, COI1 and PDF1.2) pathways. To do this, the 8K cDNAs of two Peruvian PMTV isolates were cloned in pGWB17 vector with a C-terminal myc tag and N-terminal 35S promoter using Gateway technology. Agrobacterium cultures harboring PMTV 8K were syringe infiltrated into the abaxial side of N. benthamiana leaves. The expression levels of defense genes were examined in N. benthamiana leaves infiltrated with P1 8K, P11 8K and the control constructs at 2 and 5 days post infiltration in response to PMTV 8Ks using q-PCR technique. Our results showed that the expression levels of ATG6, ATG2, ATG7, ICS1, OPR3, NPR1, PR1, COI1 and PDF1.2 were increased in response to both 8K suppressors. However, the transcript level of Argonaute1 (AGO1) was decreased in response to both 8K suppressors compared with the control. These results indicated that 8K suppressor proteins can alter the expression of autophagy, SA and JA signaling pathway genes in N. benthamiana. Taken together, it seems that despite the 8K role in virus pathogenicity, it can also induce host defense responses to modulate plant-virus interactions and fine-tune host-virus coexistence.