Sulfate availability critically influences plant growth, yet the role of small RNAs, particularly microRNAs (miRNAs), in regulating responses to sulfate deficiency remains poorly understood. Here, we conducted a temporal analysis of sulfate deficiency-responsive miRNAs in the roots and leaves of Solanum lycopersicum (tomato), using an updated miRNA annotation in the SL4.0 genome. We found 40 differentially expressed miRNAs, including 2 novel, tomato-specific miRNAs. Tomato miRNAs showed an important time- and organ-specific regulation, similar to the described response of the mRNA transcriptome. Integration with transcriptomic data and Degradome-seq analysis highlighted both canonical and non-canonical targets for sulfate-responsive miRNAs. miR395, the most extensively studied miRNA, was found to control not only its conserved targets involved in sulfate transport and assimilation, but also genes involved in redox homeostasis, photosynthesis and chloride transport. Notably, most targets were repressed in leaves, suggesting miRNA-mediated downregulation of energy-intensive processes, while root targets were predominantly upregulated, including genes related to protein remodeling and antioxidant defense. Comparative analysis with Arabidopsis thaliana revealed a broader functional repertoire in tomato, suggesting species-specific adaptations to sulfate deficiency. Overall, our results underscore the critical role of miRNAs in fine-tuning organ-specific metabolic reprogramming during nutrient stress, expanding the current understanding of the regulatory landscape underlying sulfate deficiency in plants.

Mutualistic interactions between plants and beneficial fungi rely on extensive transcriptional reprogramming in both partners, yet the underlying regulatory mechanisms coordinating these responses remain incompletely understood. Here, we combined a transcriptomics analysis with a gene regulatory network (GRN) inference to dissect the interaction between Arabidopsis thaliana and the growth-promoting fungus Trichoderma atroviride. At an early but established stage of colonization (72 h post-inoculation), we identified widespread transcriptional changes in both of the organisms, including host activation of hypoxia, stress and root development-related pathways in Arabidopsis, and fungal reprogramming of membrane transport and primary metabolism. Using DNA-binding motifs and GENIE3-based regulatory inference, we reconstructed interaction-specific GRNs for each species. The subnetworks focused on the main differentially expressed biological processes and uncovered ERF-, WRKY-, NAC- and DOF-centered modules linking hypoxia responses with developmental remodeling in the plant, whereas the putative orthologs of TFs involved in developmental and stress-related TFs in fungi, such as CrzA, RME1, NsdC, PacC and RPN4, formed a regulatory core coordinating fungal transport and metabolic adjustment. In parallel, we uncovered contrasting sRNA dynamics between the partners. While the Arabidopsis sRNA changes were limited, T. atroviride exhibited a strong induction of 20–22 nt sRNAs, including a small set of high-confidence sRNA–mRNA interactions targeting host genes involved in root function and immunity. Together, our results extend previous pathway-based descriptions of the Arabidopsis–Trichoderma mutualism and provide a systems-level, testable framework for how coordinated regulatory programs in both of the partners support the interaction.

Sulfur is an essential element for biological molecules, with sulfate as the primary sulfur source for plants. Under sulfate-deficiency, plants undergo physiological, metabolic, transcriptional, and post-transcriptional changes. Although Solanum lycopersicum (tomato) exhibits extensive transcriptome reprogramming during sulfate deficiency, the role of microRNAs (miRNAs) in this response remains poorly explored. In this study, we analyzed miRNAs expression in tomato roots and leaves at 3- and 4-weeks post-sowing under sulfate-sufficient and sulfate-deficient conditions, leveraging a de novo annotation of miRNA genes in the SL4.0 genome. We identified several miRNAs showing tissue- and time-specific expression changes in response to sulfate deficiency, with a potential role in processes such as lignin, chlorophyll, and polysaccharides metabolism. Consistent with other plants, miR395 emerged as a key sulfur metabolism and transport regulator. Notably, we identified a novel miR395 target, SlERF2a, an Ethylene-Responsive Factor predicted to regulate numerous sulfate deficiency-responsive genes, including genes involved in plastid organization, detoxification, sulfur assimilation and starvation response. These results suggest a broader role for miR395 in sulfate-deficiency responses in tomato compared to other plants. This work provides new insights into the post-transcriptional regulation of sulfate-deficiency in tomato, providing an updated miRNA annotation for future studies on miRNA-mediated regulation in this relevant crop.