苹果m6A去甲基化酶MdALKBH1A调控高温和碳饥饿胁迫抗性

doi:10.1016/j.jia.2025.01.001

摘要/Abstract

摘要：

前人的研究结果证明了m⁶A去甲基化酶在协调植物胁迫反应中的关键作用；然而，苹果m⁶A去甲基化酶在热胁迫和固定碳饥饿条件下的功能尚不清楚。本研究鉴定了苹果RNA去甲基化酶基因家族，并选择了苹果RNA去甲基化酶基因MdALKBH1A进行进一步研究。通过LC-MS/MS分析方法证明了MdALKBH1A是苹果的m⁶A去甲基化酶。过表达MdALKBH1A的转基因‘Micro Tom’番茄植株对高温更为敏感，这可能与抗氧化能力降低、膜脂过氧化作用增加、质膜稳定性降低有关。此外，过表达MdALKBH1A的番茄植株通过提高质膜稳定性、光合速率和自噬活性增强其对碳饥饿胁迫的抗性。综上所述，本研究阐明了苹果MdALKBH1A在应对高温胁迫和碳饥饿胁迫中的关键作用。

Abstract:

The pivotal role of N⁶-methyladenosine (m⁶A) demethylases in regulating plant stress responses has been widely explored; however, the function of apple m⁶A demethylases under heat stress and fixed-carbon starvation is unclear. In this study, the apple RNA demethylase gene family was identified, and the demethylase gene MdALKBH1A was selected for further analysis. Using liquid chromatography-tandem mass spectrometry (LC-MS/MS) analysis, we demonstrated that MdALKBH1A is the m⁶A demethylase of apple. Moreover, transgenic ‘Micro Tom’ tomato plants overexpressing MdALKBH1A were more sensitive to high temperature, probably due to the decreased antioxidant ability, increased membrane lipid peroxidation and reduced plasma membrane stability. However, these tomato plants overexpressing MdALKBH1A were more resistant to fixed-carbon starvation, as evidenced by the improved plasma membrane stability, enhanced photosynthetic rates and elevated autophagic activity. In summary, our results highlight the crucial role played by MdALKBH1A in the response of apple plants to high-temperature stress and fixed-carbon starvation.

Key words: MdALKBH1A , heat stress , fixed-carbon starvation , tomato , apple

Ru Bao, Tianli Guo, Zehua Yang, Chengyu Feng, Junyao Wu, Xiaomin Fu, Liu Hu, Changhai Liu, Fengwang Ma. 苹果m6A去甲基化酶MdALKBH1A调控高温和碳饥饿胁迫抗性[J]. Journal of Integrative Agriculture, 2025, 24(4): 1489-1502.

Ru Bao, Tianli Guo, Zehua Yang, Chengyu Feng, Junyao Wu, Xiaomin Fu, Liu Hu, Changhai Liu, Fengwang Ma. Overexpression of the apple m⁶A demethylase gene MdALKBH1A regulates resistance to heat stress and fixed-carbon starvation[J]. Journal of Integrative Agriculture, 2025, 24(4): 1489-1502.

参考文献

Abel S, Theologis A. 1995. A polymorphic bipartite motif signals nuclear targeting of early auxin-inducible proteins related to PS-IAA4 from pea (Pisum sativum). The Plant Journal, 8, 87–96.

Amara U, Shoaib Y, Kang H. 2022. ALKBH9C, a potential RNA m6A demethylase, regulates the response of Arabidopsis to abiotic stresses and abscisic acid. Plant, Cell & Environment, 45, 3566–3581.

Berry J A, Bjrkman O. 1980. Photosynthetic response and adaptation to temperature in higher plants. Annual Review of Plant Physiology, 31, 491–543.

Brychkova G, Yarmolinsky D, Batushansky A, Grishkevich V, Khozin-Goldberg I, Fait A, Amir R, Fluhr R, Sagi M. 2015. Sulfite oxidase activity is essential for normal sulfur, nitrogen and carbon metabolism in tomato leaves. Plants (Basel, Switzerland), 4, 573–605.

Chen P X, Zhi F, Li X W, Shen W Y, Yan M J, He J Q, Bao C, Fan T, Zhou S X, Ma F W, Guan Q M. 2022. Zinc-finger protein MdBBX7/MdCOL9, a target of MdMIEL1 E3 ligase, confers drought tolerance in apple. Plant Physiology, 188, 540–559.

Chen W, Gong L, Guo Z L, Wang W S, Zhang H Y, Liu X Q, Yu S B, Xiong L Z, Luo J. 2013. A novel integrated method for large-scale detection, identification, and quantification of widely targeted metabolites: Application in the study of rice metabolomics. Molecular Plant, 6, 1769–1780.

Duan H C, Wei L H, Zhang C, Wang Y, Chen L, Lu Z, Chen P R, He C, Jia G. 2017. ALKBH10B is an RNA N6-methyladenosine demethylase affecting Arabidopsis floral transition. Plant Cell, 29, 2995–3011.

Guo T L, Bao R, Yang Z H, Fu X M, Hu L, Wang N, Liu C H, Ma F W. 2023. The m6A reader MhYTP2 negatively modulates apple Glomerella leaf spot resistance by binding to and degrading MdRGA2L mRNA. Molecular Plant Pathology, 24, 1287–1299.

Guo T L, Liu C H, Meng F X, Hu L, Fu X M, Yang Z H, Wang N, Jiang Q, Zhang X Z, Ma F W. 2022. The m6A reader MhYTP2 regulates MdMLO19 mRNA stability and antioxidant genes translation efficiency conferring powdery mildew resistance in apple. Plant Biotechnology Journal, 20, 511–525.

Guo T L, Wang N, Xue Y C, Guan Q M, van Nocker S, Ma F W, Liu C H. 2019. Overexpression of the RNA binding protein MhYTP1 in transgenic apple enhances drought tolerance and WUE by improving ABA level under drought condition. Plant Science, 280, 397–407.

Hu J Z, Manduzio S, Kang H. 2019. Epitranscriptomic RNA methylation in plant development and abiotic stress responses. Frontiers in Plant Science, 10, 500.

Huong T T, Ngoc L N T, Kang H. 2020. Functional characterization of a putative RNA demethylase ALKBH6 in Arabidopsis growth and abiotic stress responses. International Journal of Molecular Sciences, 21, 6707.

Jia G, Fu Y, Zhao X, Dai Q, Zheng G, Yang Y, Yi C, Lindahl T, Pan T, Yang Y G, He C. 2011. N6-methyladenosine in nuclear RNA is a major substrate of the obesity-associated FTO. Nature Chemical Biology, 7, 885–887.

Jia M, Liu X, Xue H, Wu Y, Shi L, Wang R, Chen Y, Xu N, Zhao J, Shao J, Qi Y, An L, Sheen J, Yu F. 2019. Noncanonical ATG8–ABS3 interaction controls senescence in plants. Nature Plants, 5, 212–224.

Johnson M P. 2016. Photosynthesis. Essays in Biochemistry, 60, 255–273.

Kim K M, Cho H, Choi K, Kim J, Kim B W, Ko Y G, Jang S K, Kim Y K. 2009. A new MIF4G domain-containing protein, CTIF, directs nuclear cap-binding protein CBP80/20-dependent translation. Genes and Development, 23, 2033–2045.

Klionsky D J. 2004. Cell biology: Regulated selfcanni-balism. Nature, 431, 31–32.

Kurek I, Chang T K, Bertain S M, Madrigal A, Liu L, Lassner M W, Zhu G. 2007. Enhanced thermostability of Arabidopsis Rubisco activase improves photosynthesis and growth rates under moderate heat stress. Plant Cell, 19, 3230–3241.

Lee A S, Kranzusch P J, Cate J H. 2015. eIF3 targets cellprolifer-ation messenger RNAs for translational activation or repression. Nature, 522, 111–114.

Li F, Vierstra R D. 2012. Autophagy: A multifaceted intracellular system for bulk and selective recycling. Trends in Plant Science, 17, 526–537.

Liu G F, Wang J, Hou X L. 2020. Transcriptome-wide N6-methyladenosine (m6A) methylome profiling of heat stress in Pak-choi (Brassica rapa ssp. chinensis). Plants (Basel, Switzerland), 9, 1080.

Liu S, Sun B, Cao B, Lv Y, Chen Z, Xu K. 2023. Effects of soil waterlogging and high-temperature stress on photosynthesis and photosystem II of ginger (Zingiber officinale). Protoplasma, 260, 405–418.

Liu Y, Bassham D C. 2012. Autophagy: Pathways for self-eating in plant cells. Annual Review of Plant Biology, 63, 215–237.

Livak K J, Schmittgen T D. 2001. Analysis of relative gene expression data using real-time quantitative PCR and the 2–∆∆CT method. Methods, 25, 402–408.

Lobell D B, Field C B. 2007. Global scale climate-crop yield relationships and the impacts of recent warming. Environmental Research Letters, 2, 014002.

Lunn J E, Macrae E. 2003. New complexities in the synthesis of sucrose. Current Opinion in Plant Biology, 6, 208–214.

Luo G Z, MacQueen A, Zheng G, Duan H, Dore L C, Lu Z, Liu J, Chen K, Jia G, Bergelson J, He C. 2014. Unique features of the m6A methylome in Arabidopsis thaliana. Nature Communications, 5, 5630.

Ma J, Du G Y, Li X H, Zhang C Y, Guo J K. 2015. A major locus controlling malondialdehyde content under water stress is associated with Fusarium crown rot resistance in wheat. Molecular Genetics and Genomics, 290, 1955–1962.

Ma L, Li G. 2021. Arabidopsis FAR-RED ELONGATED HYPOCOTYL3 negatively regulates carbon starvation responses. Plant, Cell & Environment, 44, 1816–1829.

Martínez-Pérez M, Aparicio F, Arribas-Hernández L, Tankmar M D, Rennie S, von Bülow S, Lindorff-Larsen K, Brodersen P, Pallas V. 2023. Plant YTHDF proteins are direct effectors of antiviral immunity against an N6-methyladenosine-containing RNA virus. The EMBO Journal, 42, e113378.

Meissner R, Jacobson Y, Melamed S, Levyatuv S, Shalev G, Ashri A, Elkind Y, Levy A. 1997. A new model system for tomato genetics. The Plant Journal, 12, 1465–1472.

Meyer K D, Patil D P, Zhou J, Zinoviev A, Skabkin M A, Elemento O, Pestova T V, Qian S B, Jaffrey S R. 2015. 5´ UTR m6A promotes cap-independent translation. Cell, 163, 999–1010.

Roundtree I A, Luo G Z, Zhang Z, Wang X, Zhou T, Cui Y, Sha J, Huang X, Guerrero L, Xie P, He E, Shen B, He C. 2017. YTHDC1 mediates nuclear export of N6-methyladenosine methylated mRNAs. eLife, 6, e31311.

Salvucci M E, Crafts-Brandner S J. 2004. Inhibition of photosynthesis by heat stress: The activation state of Rubisco as a limiting factor in photosynthesis. Physiologia Plantarum, 120, 179–186.

Schofflfl F, Prandl R, Reindl A. 1998. Regulation of the heat-shock response. Plant Physiology, 117, 1135–1141.

Schwartz S, Agarwala S D, Mumbach M R, Jovanovic M, Mertins P, Shishkin A, Tabach Y, Mikkelsen T S, Satija R, Ruvkun G, Carr S A, Lander E S, Fink G R, Regev A. 2013. High-resolution mapping reveals a conserved, widespread, dynamic mRNA methylation program in yeast meiosis. Cell, 155, 1409–1421.

Shao Y, Wong C E, Shen L, Yu H. 2021. N6-methyladenosine modification underlies messenger RNA metabolism and plant development. Current Opinion in Plant Biology, 63, 102047.

Sharma D K, Andersen S B, Ottosen C O, Rosenqvist E. 2015. Wheat cultivars selected for high Fv/Fm under heat stress maintain high photosynthesis, total chlorophyll, stomatal conductance, transpiration and dry matter. Physiologia Plantarum, 153, 284–298.

Shi H, Wei J, He C. 2019. Where, when, and how: Context-dependent functions of RNA methylation writers, readers, and erasers. Molecular Cell, 74, 640–650.

Shoaib Y, Hu J, Manduzio S, Kang H. 2021. Alpha-ketoglutarate-dependent dioxygenase homolog 10B, an N6-methyladenosine mRNA demethylase, plays a role in salt stress and abscisic acid responses in Arabidopsis thaliana. Physiologia Plantarum, 173, 1078–1089.

Sun X, Wang P, Jia X, Huo L, Che R, Ma F. 2018. Improvement of drought tolerance by overexpressing MdATG18a is mediated by modified antioxidant system and activated autophagy in transgenic apple. Plant Biotechnology Journal, 16, 545–557.

Wang X, Lu Z, Gomez A, Hon G C, Yue Y, Han D, Fu Y, Parisien M, Dai Q, Jia G, Ren B, Pan T, He C. 2014. N6-methyladenosine-dependent regulation of messenger RNA stability. Nature, 505, 117–120.

Wang X, Zhao B S, Roundtree I A, Lu Z, Han D, Ma H, Weng X, Chen K, Shi H, He C. 2015. N(6)-methyladenosine modulates messenger RNA translation efficiency. Cell, 161, 1388–1399.

Xue H, Meng J, Lei P, Cao Y, An X, Jia M, Li Y, Liu H, Sheen J, Liu X, Yu F. 2022. ARF2-PIF5 interaction controls transcriptional reprogramming in the ABS3-mediated plant senescence pathway. The EMBO Journal, 41, e110988.

Yu J Y, Li Y, Wang T, Zhong X. 2018. Modification of N6-methyladenosine RNA methylation on heat shock protein expression. PLoS ONE, 13, e0198604.

Zandalinas S I, Mittler R, Balfagón D, Arbona V, Gómez-Cadenas A. 2018. Plant adaptations to the combination of drought and high temperatures. Physiologia Plantarum, 162, 2–12.

Zhao J G, Lu Z G, Wang L, Jin B. 2020. Plant responses to heat stress: Physiology, transcription, noncoding RNAs, and epigenetics. International Journal of Molecular Sciences, 22, 117.

Zhao X, Yang Y, Sun B F, Shi Y, Yang X, Xiao W, Hao Y J, Ping X L, Chen Y S, Wang W J, Jin K X, Wang X, Huang C M, Fu Y, Ge X M, Song S H, Jeong H S, Yanagisawa H, Niu Y, Jia G F, et al. 2014. FTO-dependent demethylation of N6-methyladenosine regulates mRNA splicing and is required for adipogenesis. Cell Research, 24, 1403–1419.

Zhou J, Wang J, Yu J Q, Chen Z. 2014. Role and regulation of autophagy in heat stress responses of tomato plants. Frontiers in Plant Science, 5, 174.