Argonaute proteins(AGOs) are central to RNA silencing pathways and play critical roles in plant antiviral defense.However,the functions of individual AGOs in rice remain incompletely understood.In this study,we demons...Argonaute proteins(AGOs) are central to RNA silencing pathways and play critical roles in plant antiviral defense.However,the functions of individual AGOs in rice remain incompletely understood.In this study,we demonstrate that rice AGO2contributes to resistance against rice ragged stunt virus(RRSV) through a regulatory module involving mi R167g-3p and its target gene,SNAP32.Immunoprecipitation coupled with small RNA sequencing revealed that AGO2 associates not only with virus-derived small interfering RNAs(vsi RNAs)but also preferentially associates with mi R167g-3p during RRSV infection.Functional analyses further showed that mi R167g-3p expression is induced upon infection.Transgenic rice lines overexpressing mi R167g-3p exhibited enhanced resistance,whereas knockdown lines were more susceptible.SNAP32 was validated as a direct target of mi R167g-3p through transient expression assays in Nicotiana benthamiana and dual-luciferase assays in rice protoplasts.Expression analyses confirmed that mi R167g-3p represses SNAP32 at the transcript level.Consistently,SNAP32-overexpressing plants displayed increased susceptibility to RRSV,while snap32 knockout plants showed enhanced resistance,supporting a negative role of SNAP32 in antiviral defense.Together,these findings establish a regulatory pathway in which AGO2 promotes antiviral immunity by stabilizing mi R167g-3p to repress SNAP32,thereby restricting RRSV infection.This work advances our understanding of AGO2-mediated defense in rice and highlights the use of a mi RNA 3p strand within an AGO-mi RNA-target module as an important layer of resistance against viral pathogens.展开更多
基金supported by grants from the Biological Breeding-National Science and Technology Major Project(2023ZD04070)the National Natural Science Foundation of China (Nos. 32025031,31871934,32170167)the Natural Science Foundation of Fujian Province (2025J01549)。
文摘Argonaute proteins(AGOs) are central to RNA silencing pathways and play critical roles in plant antiviral defense.However,the functions of individual AGOs in rice remain incompletely understood.In this study,we demonstrate that rice AGO2contributes to resistance against rice ragged stunt virus(RRSV) through a regulatory module involving mi R167g-3p and its target gene,SNAP32.Immunoprecipitation coupled with small RNA sequencing revealed that AGO2 associates not only with virus-derived small interfering RNAs(vsi RNAs)but also preferentially associates with mi R167g-3p during RRSV infection.Functional analyses further showed that mi R167g-3p expression is induced upon infection.Transgenic rice lines overexpressing mi R167g-3p exhibited enhanced resistance,whereas knockdown lines were more susceptible.SNAP32 was validated as a direct target of mi R167g-3p through transient expression assays in Nicotiana benthamiana and dual-luciferase assays in rice protoplasts.Expression analyses confirmed that mi R167g-3p represses SNAP32 at the transcript level.Consistently,SNAP32-overexpressing plants displayed increased susceptibility to RRSV,while snap32 knockout plants showed enhanced resistance,supporting a negative role of SNAP32 in antiviral defense.Together,these findings establish a regulatory pathway in which AGO2 promotes antiviral immunity by stabilizing mi R167g-3p to repress SNAP32,thereby restricting RRSV infection.This work advances our understanding of AGO2-mediated defense in rice and highlights the use of a mi RNA 3p strand within an AGO-mi RNA-target module as an important layer of resistance against viral pathogens.
