Alfalfa is valued worldwide for high biomass, protein-rich forage and biological nitrogen fixation, but its productivity and feed quality are tightly linked. Longer, thicker stems support yield, whereas excessive cell-wall deposition and lignification reduce digestibility. Carbon–nitrogen allocation also determines whether plants invest resources in soluble sugars, proteins or structural fiber. Although hormones and transcription factors are known to influence these traits, far less is understood about epitranscriptomic regulation—the chemical modification of RNA after it is produced. In particular, the role of m6A, the most abundant internal modification in eukaryotic mRNA, has remained largely unexplored in perennial forage legumes. Because of these challenges, deeper investigation is needed into how RNA methylation coordinates alfalfa growth and nutrient metabolism.
Researchers from China Agricultural University, Shanxi Agricultural University, the Shandong Academy of Agricultural Science and the Xinjiang Uygur Autonomous Region Academy of Agricultural Sciences reported (DOI: 10.1093/hr/uhag146) the study in Horticulture Research on April 16, 2026. They identified MsMTA as a nuclear m6A methyltransferase that connects RNA stability with stem development and forage composition in alfalfa (Medicago sativa). By combining transgenic experiments with transcriptome-wide RNA modification mapping, the team showed that lowering MsMTA expression disrupts metabolic gene stability and shifts the plant toward shorter growth, higher soluble sugar levels and lower fiber content.
The team generated alfalfa lines in which MsMTA was reduced through RNA interference (RNAi), alongside overexpression lines and wild-type controls. In three RNAi lines, MsMTA expression fell by 39.14%–52.84%, while total m6A abundance dropped by 43.41%–58.35%. These plants developed shorter internodes, thinner stems and less dry matter, but their leaves were largely unchanged. Nutritionally, soluble sugar increased, acid detergent fiber decreased by 9.30%–15.89%, and crude protein declined by only 5.11%–7.55%.
Oxford Nanopore Technologies direct RNA sequencing (ONT–DRS) showed that m6A marks were concentrated mainly in the 3′ untranslated region (3′ UTR), and silencing MsMTA reduced the number of modified sites from 8,089 to 5,314. RNA sequencing identified 549 genes with altered expression. Integrating both datasets revealed 110 genes with changes in methylation and abundance, including 59 that lost m6A and were downregulated. The team then used m6A immunoprecipitation followed by quantitative polymerase chain reaction (m6A-IP–qPCR), reverse-transcription quantitative polymerase chain reaction (RT–qPCR), and actinomycin D decay assays. These tests showed that MsSDR1, MsLOX3, MsSCPL22, MsATPC2, MsGS2 and MsAPT1 transcripts decayed faster when MsMTA was suppressed, directly linking RNA methylation to metabolic transcript stability.
“Our results suggest that MsMTA acts as a molecular coordinator rather than a simple growth switch,” the authors said. “When its activity is reduced, alfalfa does not merely become shorter; it reallocates carbon and nitrogen, changes fiber and sugar accumulation, and destabilizes transcripts that support energy production, redox control and nitrogen assimilation. This trade-off is important because improving one forage trait can come at the expense of another. The study therefore provides a clearer target for designing precise strategies that protect biomass while improving nutritional value.”
The work points toward precision breeding rather than complete suppression of MsMTA. Direct silencing raised sugar and lowered fiber, but it also reduced plant height, dry matter and crude protein, making whole-plant knockdown unsuitable as an immediate breeding solution. More selective approaches—such as tissue-specific, developmental-stage-specific or moderate adjustment of m6A activity—could help retain stem growth while improving digestibility and energy content. Downstream targets involved in carbon–nitrogen metabolism, photosynthetic energy distribution or cell-wall formation may offer additional routes with fewer growth penalties. Field trials will still be needed to test whether these laboratory and greenhouse effects remain stable across environments, harvest stages and alfalfa genetic backgrounds before the mechanism can support cultivar development.
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References
DOI
10.1093/hr/uhag146
Original Source URL
https://doi.org/10.1093/hr/uhag146
Funding information
We extend our gratitude for the financial assistance provided by the National Natural Science Foundation of China, under grant number 32071870.
About Horticulture Research
Horticulture Research is an open access journal of Nanjing Agricultural University and ranked number one in the Horticulture category of the Journal Citation Reports ™ from Clarivate, 2023. The journal is committed to publishing original research articles, reviews, perspectives, comments, correspondence articles and letters to the editor related to all major horticultural plants and disciplines, including biotechnology, breeding, cellular and molecular biology, evolution, genetics, inter-species interactions, physiology, and the origination and domestication of crops.