Label-free quantitative proteomic analysis of alfalfa in response to microRNA156 under high temperature
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RESEARCH ARTICLE
Open Access
Label-free quantitative proteomic analysis of alfalfa in response to microRNA156 under high temperature Muhammad Arshad1,2*, Alpa Puri1,3, Aaron J. Simkovich1,3, Justin Renaud1, Margaret Y. Gruber4, Frédéric Marsolais1,3 and Abdelali Hannoufa1,3*
Abstract Background: Abiotic stress, including heat, is one of the major factors that affect alfalfa growth and forage yield. The small RNA, microRNA156 (miR156), regulates multiple traits in alfalfa during abiotic stress. The aim of this study was to explore the role of miR156 in regulating heat response in alfalfa at the protein level. Results: In this study, we compared an empty vector control and miR156 overexpressing (miR156OE) alfalfa plants after exposing them to heat stress (40 °C) for 24 h. We measured physiological parameters of control and miR156OE plants under heat stress, and collected leaf samples for protein analysis. A higher proline and antioxidant contents were detected in miR156OE plants than in controls under heat stress. Protein samples were analyzed by label-free quantification proteomics. Across all samples, a total of 1878 protein groups were detected. Under heat stress, 45 protein groups in the empty vector plants were significantly altered (P < 0.05; |log2FC| > 2). Conversely, 105 protein groups were significantly altered when miR156OE alfalfa was subjected to heat stress, of which 91 were unique to miR156OE plants. The identified protein groups unique to miR156OE plants were related to diverse functions including metabolism, photosynthesis, stress-response and plant defenses. Furthermore, we identified transcription factors in miR156OE plants, which belonged to squamosa promoter binding-like protein, MYB, ethylene responsive factors, AP2 domain, ABA response element binding factor and bZIP families of transcription factors. Conclusions: These results suggest a positive role for miR156 in heat stress response in alfalfa. They reveal a miR156-regulated network of mechanisms at the protein level to modulate heat responses in alfalfa. Keywords: Alfalfa, Heat stress, miR156, Proteomic, LC-MS/MS
Background Alfalfa (Medicago sativa L.) is an important leguminous crop that is grown worldwide as forage for livestock feed, and contributes to improved soil quality. Another important feature of this crop is the potential for multiple harvests throughout the growing season, allowing for abundant biomass yield. Perennial nature of alfalfa * Correspondence: [email protected]; [email protected] 1 Agriculture and Agri-Food Canada, 1391 Sandford Street, London, Ontario N5V 4T3, Canada Full list of author information is available at the end of the article
and its rapid biomass production make it a suitable source for bioenergy purpose. However, these benefits are offset by alfalfa’s susceptibility to environmental stresses including heat, which exerts adverse effects on its growth and productivity [1]. High temperature can negatively affect plant growth and development including reduced seed germination [2], damage to leaves
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