A calmodulin-like protein suppresses RNA silencing and promotes geminivirus infection by degrading SGS3 via the autophagy pathway in Nicotiana benthamiana.

Fangfang Li,Xueping Zhou,Xiuling Yang,Aiming Wang,Xiongbiao Xu,Zhenghe Li,Nan Zhao,Yaqin Wang,Shu-Sheng Liu,David M Bisaro

doi:10.1371/journal.ppat.1006213

Abstract

A recently characterized calmodulin-like protein is an endogenous RNA silencing suppressor that suppresses sense-RNA induced post-transcriptional gene silencing (S-PTGS) and enhances virus infection, but the mechanism underlying calmodulin-like protein-mediated S-PTGS suppression is obscure. Here, we show that a calmodulin-like protein from Nicotiana benthamiana (NbCaM) interacts with Suppressor of Gene Silencing 3 (NbSGS3). Deletion analyses showed that domains essential for the interaction between NbSGS3 and NbCaM are also required for the subcellular localization of NbSGS3 and NbCaM suppressor activity. Overexpression of NbCaM reduced the number of NbSGS3-associated granules by degrading NbSGS3 protein accumulation in the cytoplasm. This NbCaM-mediated NbSGS3 degradation was sensitive to the autophagy inhibitors 3-methyladenine and E64d, and was compromised when key autophagy genes of the phosphatidylinositol 3-kinase (PI3K) complex were knocked down. Meanwhile, silencing of key autophagy genes within the PI3K complex inhibited geminivirus infection. Taken together these data suggest that NbCaM acts as a suppressor of RNA silencing by degrading NbSGS3 through the autophagy pathway.

Highlights

Post transcriptional gene silencing (PTGS) is an important RNA interference (RNAi)-based defense mechanism against foreign nucleic acid invasion and is involved in silencing a wide range of endogenous genes in plants
A betasatellite molecule associated with geminivirus induced high levels of an endogenous RNA silencing suppressor, calmodulin-like protein (CaM), to counter host defenses
Yeast transformants containing AD-NbRDR6 and BD-NbSGS3 or AD-NbSGS3 and BD-NbSGS3 grew on the selection plates (S1A and S1B Fig), consistent with an interaction between Suppressor of Gene Silencing 3 (SGS3) and RDR6 and self-interaction of SGS3, which was observed for AtSGS3 and AtRDR6 [12, 13]

Summary

Introduction

Post transcriptional gene silencing (PTGS) is an important RNA interference (RNAi)-based defense mechanism against foreign nucleic acid invasion and is involved in silencing a wide range of endogenous genes in plants. Many reports have shown that RDR6-deficient plants (e.g. Nicotiana benthamiana) are more susceptible to infection by some positivesense single-stranded RNA viruses, viroids and DNA viruses [7,8,9,10], strongly supporting the role of RDR6 in the host antiviral response. In these processes, a plant-specific RNA binding protein, Suppressor of Gene Silencing 3 (SGS3), functions together with RDR6 as a chaperone protein [4,5,6]. Whether SGS3 from N. benthamiana plays a similar chaperone role with NbRDR6 is still obscure

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Conclusion