Targeting of cucumber necrosis virus coat protein to the chloroplast stroma attenuates host defense response


Alam, S.B., Reade, R., Maghodia, A.B., Ghoshal, B., Theilmann, J., Rochon, D.A. (2021). Targeting of cucumber necrosis virus coat protein to the chloroplast stroma attenuates host defense response. Virology, [online] 554 106-119.

Plain language summary

This paper demonstrates that, as a result of some of the CNV coat protein (CP) going to Chloroplasts, the CP is able to interfere with the plants ability to mount a proper defense against the virus. We showed this in the context of the virus, where we made mutants that were unable to target the plant chloroplasts, and showed that these mutants displayed earlier and more severe symptoms similar to a hypersensitivity response (HR) that is typical of a plant defense response to viral infection. We also used Droplet Digital PCR (ddPCR) to show that these mutants also resulted in higher levels of expression on the plant defense related genes that we tested. In addition we did work in a semi-in vitro system where we cloned wild type (WT) CNV coat protein as well as those of several of the mutant viruses coat proteins into plant expression vectors. We were able to show that co-infiltration WT CNV CP into the leaves of plants is able to attenuate the induction of symptoms (necrosis) caused by infiltration of other viral genes, but co-infiltration with mutant CP had no effect on the necrosis. In this case two viral proteins that were known to cause necrotic symptoms when infiltrated into certain plants were used. CNVp33 replication associated gene which cause necrosis when it is infiltrated into the host N. benthamiana leaves, and TBSV p19 which induces necrosis in N. tobacum. In both cases WT CNV CP was able to significantly reduce the level of necrosis that was being caused by the other co-infiltrated viral proteins, whereas the mutant CP’s had little to no effect.


Cucumber necrosis virus (CNV) is a (+)ssRNA virus that elicits spreading local and systemic necrosis in Nicotiana benthamiana. We previously showed that the CNV coat protein (CP) arm functions as a chloroplast transit peptide that targets a CP fragment containing the S and P domains to chloroplasts during infection. Here we show that several CP arm mutants that inefficiently target chloroplasts, along with a mutant that lacks the S and P domains, show an early onset of more localized necrosis along with protracted induction of pathogenesis related protein (PR1a). Agroinfiltrated CNV CP is shown to interfere with CNV p33 and Tomato bushy stunt virus p19 induced necrosis. Additionally, we provide evidence that a CP mutant that does not detectably enter the chloroplast stroma induces relatively higher levels of several plant defense-related genes compared to WT CNV. Together, our data suggest that targeting of CNV CP to the chloroplast stroma interferes with chloroplast-mediated plant defense.

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