Cross-kingdom sRNA Ta_sRNA1 silences PRIM2 to fine-tune Arabidopsis immunity during symbiosis with Trichoderma atroviride

Abstract

· Trichoderma spp. enhance plant growth and immunity through beneficial root colonization, however, the role of fungal small RNAs (sRNAs) in this interaction remains poorly understood. We identified Ta_sRNA1, a highly abundant T. atroviride sRNA transferred into Arabidopsis root cells, where it loads into ARGONAUTE 1 and 2 (AGO1/2) to modulate host gene expression.

· Through stem-loop RT-qPCR, AGO immunoprecipitation, transgenic lines and overexpression strains, we validated PRIM2 (encoding the DNA primase large subunit), as a functional Arabidopsis target ofTa_sRNA1. Defense and colonization phenotypes were evaluated in lines altered for Ta_sRNA1 or PRIM2expression, with or without Trichoderma pretreatment.

· Ta_sRNA1 silencing of PRIM2 restricts Trichoderma overcolonization and enhances resistance to Botrytis cinerea. This repression primes systemic immunity and boosts ROS accumulation without growth penalties. In contrast, PRIM2 overexpression suppresses ROS and promotes necrosis, supporting its role as a susceptibility (S) gene. Sustained Ta_sRNA1 expression compromises Trichoderma-induced systemic resistance and perturbs JA/ET-SA signaling balance.

· We conclude that Ta_sRNA1 is a cross-kingdom sRNA that promotes immune priming and limits colonization by repressing PRIM2. This positions PRIM2 as a key regulator of ROS and hormone-driven immune balance and highlights fungal sRNAs as fine-tuners of plant defense and mutualism.