Transcriptome-IPMS analysis reveals a tissue-dependent miR156/SPL13 regulatory mechanism in alfalfa drought tolerance
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RESEARCH ARTICLE
Open Access
Transcriptome-IPMS analysis reveals a tissue-dependent miR156/SPL13 regulatory mechanism in alfalfa drought tolerance Biruk A. Feyissa1,2, Justin Renaud2, Vida Nasrollahi1,2, Susanne E. Kohalmi1 and Abdelali Hannoufa1,2*
Abstract Background: We previously reported on the interplay between miR156/SPL13 and WD40–1/DFR to improve response to drought stress in alfalfa (Medicago sativa L.). Here we aimed to investigate whether the role of miR156/ SPL13 module in drought response is tissue-specific, and to identify SPL13-interacting proteins. We analyzed the global transcript profiles of leaf, stem, and root tissues of one-month old RNAi-silenced SPL13 (SPL13RNAi) alfalfa plants exposed to drought stress and conducted protein-protein interaction analysis to identify SPL13 interacting partners. Result: Transcript analysis combined with weighted gene co-expression network analysis showed tissue and genotype-specific gene expression patterns. Moreover, pathway analysis of stem-derived differentially expressed genes (DEG) revealed upregulation of genes associated with stress mitigating primary and specialized metabolites, whereas genes associated with photosynthesis light reactions were silenced in SPL13RNAi plants. Leaf-derived DEG were attributed to enhanced light reactions, largely photosystem I, II, and electron transport chains, while roots of SPL13RNAi plants upregulated transcripts associated with metal ion transport, carbohydrate, and primary metabolism. Using immunoprecipitation combined with mass spectrometry (IPMS) we showed that SPL13 interacts with proteins involved in photosynthesis, specialized metabolite biosynthesis, and stress tolerance. Conclusions: We conclude that the miR156/SPL13 module mitigates drought stress in alfalfa by regulating molecular and physiological processes in a tissue-dependent manner. Keywords: Alfalfa, Drought, IPMS, Medicago sativa, miR156, SPL13, Transcriptome
Background The frequent and extreme weather events in present-day are correlated with climate change, which aggravates crop losses [38, 42]. To cope with these weather events, plants respond by developing different resilience strategies at the phenotypic, physiological, and molecular levels [23, 51]. Among the molecular strategies, the role of microRNAs in regulating various plant processes to * Correspondence: [email protected] 1 Department of Biology, University of Western Ontario, 1151 Richmond Street, London, Ontario N6A3K7, Canada 2 Agriculture and Agri-Food Canada, 1391 Sandford Street, London, Ontario N5V 4T3, Canada
enhance stress tolerance has been reported in a variety of crops [21]. Of the hundreds of plant microRNAs, microRNA156 (miR156) is highly conserved across species and regulates plant development as well as tolerance to biotic and abiotic stresses [2, 4, 5, 15, 17, 21, 23, 35]. Recent findings showed moderate levels of miR156 overexpression enhances drought tolerance in alfalfa by silencing SPL13 [2] and enhancing the downstream dihydroflavonol-4-reductase (DFR) wh
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