Cukrov Dubravka, Zermiani Monica, Brizzolara Stefano, Cestaro Alessandro, Licausi Francesco, Luchinat Claudio, Santucci Claudio, Tenori Leonardo, Van Veen Hans, Zuccolo Andrea, Ruperti Benedetto, Tonutti Pietro
Istituto di Scienze della Vita, Scuola Superiore Sant'Anna Pisa, Italy.
Dipartimento di Agronomia Animali Alimenti Risorse Naturali e Ambiente, University of Padova Padova, Italy.
Front Plant Sci. 2016 Feb 16;7:146. doi: 10.3389/fpls.2016.00146. eCollection 2016.
The ripening physiology of detached fruit is altered by low oxygen conditions with profound effects on quality parameters. To study hypoxia-related processes and regulatory mechanisms, apple (Malus domestica, cv Granny Smith) fruit, harvested at commercial ripening, were kept at 1°C under normoxic (control) and hypoxic (0.4 and 0.8 kPa oxygen) conditions for up to 60 days. NMR analyses of cortex tissue identified eight metabolites showing significantly different accumulations between samples, with ethanol and alanine displaying the most pronounced difference between hypoxic and normoxic treatments. A rapid up-regulation of alcohol dehydrogenase and pyruvate-related metabolism (lactate dehydrogenase, pyruvate decarboxylase, alanine aminotransferase) gene expression was detected under both hypoxic conditions with a more pronounced effect induced by the lowest (0.4 kPa) oxygen concentration. Both hypoxic conditions negatively affected ACC synthase and ACC oxidase transcript accumulation. Analysis of RNA-seq data of samples collected after 24 days of hypoxic treatment identified more than 1000 genes differentially expressed when comparing 0.4 vs. 0.8 kPa oxygen concentration samples. Genes involved in cell-wall, minor and major CHO, amino acid and secondary metabolisms, fermentation and glycolysis as well as genes involved in transport, defense responses, and oxidation-reduction appeared to be selectively affected by treatments. The lowest oxygen concentration induced a higher expression of transcription factors belonging to AUX/IAA, WRKY, HB, Zinc-finger families, while MADS box family genes were more expressed when apples were kept under 0.8 kPa oxygen. Out of the eight group VII ERF members present in apple genome, two genes showed a rapid up-regulation under hypoxia, and western blot analysis showed that apple MdRAP2.12 proteins were differentially accumulated in normoxic and hypoxic samples, with the highest level reached under 0.4 kPa oxygen. These data suggest that ripe apple tissues finely and specifically modulate sensing and regulatory mechanisms in response to different hypoxic stress conditions.
低氧条件会改变离体果实的成熟生理，对品质参数产生深远影响。为了研究缺氧相关过程和调控机制，将商业成熟时采收的苹果（Malus domestica，品种为澳洲青苹）果实于1°C下分别置于常氧（对照）和低氧（0.4和0.8 kPa氧气）条件下长达60天。对皮层组织的核磁共振分析鉴定出8种代谢物，其在样品间的积累存在显著差异，其中乙醇和丙氨酸在低氧和常氧处理间的差异最为明显。在两种低氧条件下均检测到乙醇脱氢酶和丙酮酸相关代谢（乳酸脱氢酶、丙酮酸脱羧酶、丙氨酸转氨酶）基因表达的快速上调，最低（0.4 kPa）氧气浓度诱导的效应更为显著。两种低氧条件均对乙烯合成酶和乙烯氧化酶转录积累产生负面影响。对低氧处理24天后采集的样品进行RNA测序数据分析发现，比较0.4 kPa和0.8 kPa氧气浓度样品时，有超过1000个基因差异表达。参与细胞壁、微量和大量碳水化合物、氨基酸和次生代谢、发酵和糖酵解的基因，以及参与转运、防御反应和氧化还原的基因似乎受到处理的选择性影响。最低氧气浓度诱导属于AUX/IAA、WRKY、HB、锌指家族的转录因子表达升高，而当苹果置于0.8 kPa氧气条件下时，MADS盒家族基因表达更高。在苹果基因组中存在的8个第VII类乙烯响应因子成员中，有两个基因在低氧条件下快速上调，蛋白质免疫印迹分析表明苹果MdRAP2.12蛋白在常氧和低氧样品中差异积累，在0.4 kPa氧气条件下达到最高水平。这些数据表明，成熟苹果组织会针对不同的低氧胁迫条件精细且特异性地调节感知和调控机制。
