miR7122-3p is a key regulating factor of the photosynthetic process in grapevine

Abstract

Plant microRNAs are small regulatory RNAs that control gene expression and play key roles in plant development and stress adaptation. The miR7122 locus produces two strands: miR7122-5p, previously associated with the synthesis of phased small interfering RNAs, and miR7122-3p, whose biological function is still unclear. To gain experimental evidence on the role of vvi-miR7122-3p, an in-depth physiological and molecular characterization of transgenic grapevines was carried out, in which this miRNAwas either over-accumulated or sequestered using the short tandem target mimic approach. Transgenic lines of two Vitis vinifera cultivars, ‘Bragat rosa’ and ‘Chardonnay’, overexpressing (OE) or silencing (STTM) vvi-miR7122-3p, were subjected to water stress and recovery, and their physiological and molecular responses were analyzed. Modulation of vvi-miR7122-3p significantly affected leaf gas exchanges upon drought. Particularly, STTM plants exhibited reduced carbon assimilation and abnormal high substomatal CO₂ levels, pointing to a strong impairment of carbon fixation. Notably, these alterations occurred without significant changes in photosystem II efficiency, suggesting that the limitation was mainly biochemical and associated with the Calvin cycle rather than photochemical damage. Such physiological adjustments were genotype-dependent, as ‘Bragat rosa’ STTM plants displayed stronger stress sensitivity than ‘Chardonnay’ STTM vines. Additionally, under water stress, silencing of vvi-miR7122-3p triggered a large-scale transcriptional reprogramming, leading to the repression of genes involved in photosynthesis and carbon assimilation together with activation of respiration, secondary metabolism, and defense-associated pathways. Network and correlation analyses identified the 3-deoxy-D-arabino-heptulosonate 7-phosphate synthase (VvDAHPS), a key enzyme of the shikimate pathway, as main target of vvi-miR7122-3p. Experimental validation confirmed that the miRNAdirectly regulates this gene, further supporting a model in which vvi-miR7122-3p contributes to drought acclimation through regulation of the shikimate pathway. When the miRNAamounts are reduced, VvDAHPS is derepressed, activating shikimate-derived secondary metabolism and redox-related pathways. This metabolic shift could reallocate carbon sources toward the production of antioxidant and defensive compounds, therefore lowering the photosynthetic processes and leading to elevated internal CO₂ concentrations.

Acknowledgements

This study was carried out in the frame of the project “TEA4IT_Tecnologie di Evoluzione Assistita per le filiere agroalimentari italiane” funded by the Italian Ministry of Agriculture, Food Sovereignty and Forestry (MASAF). Part of the work was carried out within the project “SHIELD4GRAPE: Breeding and integrated pest management strategies to reduce reliance on chemical pesticides in grapevine,” funded by the European Union through the Horizon Europe Research and Innovation Programme (Grant number 101135088). Views and opinions expressed are, however, those of the authors only. Neither the European Union nor the granting authority nor the European Commission can be held responsible for them.