NSP2 phosphorylation modulates HYL1 nuclear import via KETCH1 to promote miRNA biogenesis in Arabidopsis

MicroRNA (miRNA) biogenesis in plants relies on the precise nuclear-cytoplasmic trafficking of the HYL1 protein. Here, we report that the NITRILE-SPECIFIER PROTEIN 2 (NSP2) is a previously unrecognized component of the Arabidopsis miRNA pathway. Through yeast two-hybrid, BiFC, Co-IP, and genetic analyses, we demonstrate that NSP2 physically interacts with HYL1 and genetically suppresses hyl1-2. NSP2 overexpression increases global miRNA abundance. Mechanistically, NSP2 acts as an adapter that bridges the importin-β family member KETCH1 to HYL1, thereby regulating HYL1 nuclear import and dicing-body (D-body) assembly. Dephosphorylation of NSP2 at a conserved threonine (T314) strengthens KETCH1-HYL1 association and further enhances pri-miRNA processing. Our study establishes NSP2 as a phosphorylation-regulated scaffold that coordinates HYL1 nuclear trafficking and microprocessor assembly, and reveals critical roles for NSP2 in miRNA biogenesis.