The additional unique ability of TuMV HC-Pro in inhibiting HEN1 activity for enhancing the autophagic AGO1 degradation in RNA silencing suppression

Abstract

Genomes of potyviruses, the largest group of plant viruses, encode HC-Pro proteins that mediate RNA silencing suppression. HC-Pros may exhibit only 40% similarity between species, and induce different levels in autophagic ARGONAUTE1 (AGO1) degradation. Our data indicated that HC-Pro of turnip mosaic virus (HC-ProTu) could efficiently trigger AGO1 degradation through autophagy compared with HC-Pros of zucchini yellow mosaic virus (HC-ProZy) and tobacco etch virus (HC-ProTe). Furthermore, HC-ProTu, but not in HC-ProZy, forms a suppression body (S-body) to recruit AGO1 and HEN1, preventing those components from translocating into the nucleus. HC-ProTu, but not HC-ProZy and HC-ProTe, specifically inhibits HEN1 activity, resulting in unmethylated microRNAs (miRNAs) accumulating in the cytoplasm without loading into AGO1. Therefore, we hypothesize that HC-ProTu could enhance the autophagic AGO1 degradation due to the unique HEN1 inhibition interfering with RNA-inducing silencing complex (RISC) assembly.

The P1/HC-Pro of turnip mosaic virus has more inhibits RISC activity unlike the other potyviruses

“…3e). Furthermore, Wei et al (2022) and Shang (2020) demonstrated that P1/HC-Pro Tu /atg8a ge plants, which have an ATG8a knockout mutant, have recovered AGO1 levels. Surprisingly, the normalized in vitro RISC activity returned to 20% and 39% e ciency at 10 ng and 20 ng, respectively, in P1/HC-Pro Tu /atg8a ge plants, suggesting restoration of RISC regulation (Fig.…”

“…Therefore, we can apply the same approach to generate AGO2-speci c antibodies for AGO2-IP and in vitro RISC assays in comparative studies. Hu et al (2020) and Wei et al (2022) demonstrated that HC-Pro Tu has more e cient to trigger autophagic AGO1 degradation than other HC-Pro. AGO1 degradation was also observed in TuGR-infected Arabidopsis, suggesting that HC-Pro Tu -mediated AGO1 degradation is one approach for gene silencing suppression.…”

“…Surprisingly, P1/HC-Pro Tu−K plants, which lost 40% suppression ability (Kung et al, 2014), also showed 0.7-fold AGO1 levels similar to those of P1/HC-Pro Te plants, suggesting that HC-Pro Tu−K also has the ability to trigger AGO1 degradation but is not as e cient as wild-type HC-Pro Tu . Sanobar et al (2021) and Wei et al (2022) demonstrated that HC-Pro Tu has physical interaction with HEN1 through FRNK motif to inhibit the miRNA methylation, but not in HC-Pro Zy and HC-Pro Te , suggesting that HC-Pro Tu -mediated HEN1 inhibition could enhance the autophagic AGO1 degradation. Indeed, the viral species of the HC-Pro effect is also referred in RISC inhibition assay.…”

“…Arabidopsis thaliana Columbia ecotype (Col-0), P1/HC-Pro Tu , P1/HC-Pro Tu−K , P1/HC-Pro Zy , P1/HC-Pro Te , P1/HC-Pro Tu /atg8a ge plants, ago1-27, and ago1-36 mutants were used in this study (Morel et al, 2002;Wu et al, 2010;Kung et al, 2014;Shang, 2020;Wei et al, 2022). Arabidopsis seeds were surface sterilized and vernalized at 4°C for 1 days before sowing on Murashige and Skoog medium.…”

“…Many reports have implied that interfering with RNA-induced silencing complex (RISC) assembly triggers autophagy for AGO1 degradation (Yu et al, 2006;Derrien et al, 2012;Hacquard et al, 2022). Indeed, Wei et al (2022) and Shang (2020) demonstrated that a CRISPR-mediated ATG8a knockout mutant in a P1/HC-Pro Tu background (P1/HC-Pro Tu /atg8a ge plants) restored the AGO1 levels to near the levels of Col-0 plants, implying that P1/HC-Pro Tu triggers autophagic AGO1 degradation. The case of P1/HC-Pro Tu /atg8a ge can be succeeded because the T-DNA insertion of P1/HC-Pro Tu plants was identi ed in chromosome 1 (28,814,559 bp) (Hong, 2015).…”