Elucidating microbial-induced cold tolerance gene expression pathways in Arabidopsis thaliana by SA-producing...

Low temperature severely limits crop productivity, yet the molecular mechanisms by which beneficial microbes enhance plant cold tolerance remain insufficiently defined, particularly regarding the role of microbial-derived salicylic acid (SA) in host signaling. This study investigated whether bacterial SA contributes to cold adaptation and how it modulates plant regulatory networks. Arabidopsis thaliana wild-type (Col-0), the SA-signaling mutant npr1-1, and the SA-degrading line nahG were inoculated with the SA-producing bacterium Sphingomonas paucimobilis ZJSH1 or its SA-deficient mutant (ΔpchB), followed by exposure to 4 ℃ for 14 days. Endogenous SA was quantified by UPLC-MS, physiological parameters were measured, and transcriptomic changes were analyzed using RNA sequencing. S. paucimobilis strain ZJSH1 inoculation significantly increased endogenous SA levels (2.40-3.37 µg g⁻¹ protein), chlorophyll content (11.65-13.12 µg g⁻¹ protein), and proline accumulation (up to 7.33 µg g⁻¹ protein), enhanced antioxidant enzyme activities (SOD up to 346.24 U mg⁻¹ protein; POD up to 5402.22 U g⁻¹ protein, and CAT up to 1051.11 U mg⁻¹ protein), and reduced lipid peroxidation compared with ΔpchB and non-inoculated controls. RNA-seq identified 8,307, 5,341, and 6,953 differentially expressed genes in Col-0, npr1-1, and nahG, respectively, enriched in photosystem stabilization, ICE-CBF-COR signaling, MAPK pathways, and redox homeostasis. Growth-related hormone pathways (auxin, cytokinin, gibberellin) were activated, whereas abscisic acid (ABA)-associated responses were repressed. Comparative genotype analysis revealed that photosynthetic protection and antioxidant activation were largely SA-driven and independent of NPR1, while NPR1 mediated transcriptional fine-tuning and proteostasis regulation. These findings define a dual SA regulatory mechanism underlying microbe-induced cold tolerance and provide mechanistic support for the use of S. paucimobilis strain ZJSH1 as a microbial strategy to enhance plant resilience under low-temperature stress.