Melatonin enhances KCl salinity tolerance by maintaining K+ homeostasis in Malus hupehensis.

Large amounts of potash fertilizer are often applied to apple (Malus domestica) orchards to enhance fruit quality and yields, but this treatment aggravates KCl-based salinity stress. Melatonin (MT) is involved in a variety of abiotic stress responses in plants. However, its role in KCl stress tolerance is still unknown. In the present study, we determined that an appropriate concentration (100 µm) of MT significantly alleviated KCl stress in Malus hupehensis by enhancing K+ efflux out of cells and compartmentalizing K+ in vacuoles. Transcriptome deep-sequencing analysis identified the core transcription factor gene MdWRKY53, whose expression responded to both KCl and MT treatment. Overexpressing MdWRKY53 enhanced KCl tolerance in transgenic apple plants by increasing K+ efflux and K+ compartmentalization. Subsequently, we characterized the transporter genes MdGORK1 and MdNHX2 as downstream targets of MdWRKY53 by ChIP-seq. MdGORK1 localized to the plasma membrane and enhanced K+ efflux to increase KCl tolerance in transgenic apple plants. Moreover, overexpressing MdNHX2 enhanced the KCl tolerance of transgenic apple plants/callus by compartmentalizing K+ into the vacuole. RT-qPCR and LUC activity analyses indicated that MdWRKY53 binds to the promoters of MdGORK1 and MdNHX2 and induces their transcription. Taken together, our findings reveal that the MT-WRKY53-GORK1/NHX2-K+ module regulates K+ homeostasis to enhance KCl stress tolerance in apple. These findings shed light on the molecular mechanism of apple response to KCl-based salinity stress and lay the foundation for the practical application of MT in salt stress.

Melatonin promotes Al3+ compartmentalization via H+ transport and ion gradients in Malus hupehensis.

Soil acidification in apple (Malus domestica) orchards results in the release of rhizotoxic aluminum ions (Al3+) into soil. Melatonin (MT) participates in plant responses to abiotic stress; however, its role in AlCl3 stress in apple remains unknown. In this study, root application of MT (1 µM) substantially alleviated AlCl3 stress (300 µM) in Pingyi Tiancha (Malus hupehensis), which was reflected by higher fresh and dry weight, increased photosynthetic capacity, and longer and more roots compared with plants that did not receive MT treatment. MT functioned mainly by regulating vacuolar H+/Al3+ exchange and maintaining H+ homeostasis in the cytoplasm under AlCl3 stress. Transcriptome deep sequencing analysis identified the transcription factor gene SENSITIVE TO PROTON RHIZOTOXICITY 1 (MdSTOP1) was induced by both AlCl3 and MT treatments. Overexpressing MdSTOP1 in apple increased AlCl3 tolerance by enhancing vacuolar H+/Al3+ exchange and H+ efflux to the apoplast. We identified 2 transporter genes, ALUMINUM SENSITIVE 3 (MdALS3) and SODIUM HYDROGEN EXCHANGER 2 (MdNHX2), as downstream targets of MdSTOP1. MdSTOP1 interacted with the transcription factor NAM ATAF and CUC 2 (MdNAC2) to induce MdALS3 expression, which reduced Al toxicity by transferring Al3+ from the cytoplasm to the vacuole. Furthermore, MdSTOP1 and MdNAC2 coregulated MdNHX2 expression to increase H+ efflux from the vacuole to the cytoplasm to promote Al3+ compartmentalization and maintain cation balance in the vacuole. Taken together, our findings reveal an MT-STOP1 + NAC2-NHX2/ALS3-vacuolar H+/Al3+ exchange model for the alleviation of AlCl3 stress in apple, laying a foundation for practical applications of MT in agriculture.

Strigolactone alleviates the salinity-alkalinity stress of Malus hupehensis seedlings.

Salinity-alkalinity stress can remarkably affect the growth and yield of apple. Strigolactone (SL) is a class of carotenoid-derived compounds that functions in stress tolerance. However, the effects and mechanism of exogenous SL on the salinity-alkalinity tolerance of apple seedlings remain unclear. Here, we assessed the effect of SL on the salinity-alkalinity stress response of Malus hupehensis seedlings. Results showed that treatment with 100 µM exogenous SL analog (GR24) could effectively alleviate salinity-alkalinity stress with higher chlorophyll content and photosynthetic rate than the apple seedlings without GR24 treatment. The mechanism was also explored: First, exogenous GR24 regulated the expression of Na+/K+ transporter genes and decreased the ratio of Na+/K+ in the cytoplasm to maintain ion homeostasis. Second, exogenous GR24 increased the enzyme activities of superoxide, peroxidase and catalase, thereby eliminating reactive oxygen species production. Third, exogenous GR24 alleviated the high pH stress by regulating the expression of H+-ATPase genes and inducing the production of organic acid. Last, exogenous GR24 application increased endogenous acetic acid, abscisic acid, zeatin riboside, and GA3 contents for co-responding to salinity-alkalinity stress indirectly. This study will provide important theoretical basis for analyzing the mechanism of exogenous GR24 in improving salinity-alkalinity tolerance of apple.

Brassinolide alleviates Fe deficiency-induced stress by regulating the Fe absorption mechanism in Malus hupehensis Rehd.

KEY MESSAGE: Exogenous brassinolide promotes Fe absorption through mechanism I strategy, thus improving the tolerance of Malus hupehensis seedlings to Fe deficiency stress. Iron (Fe) deficiency is a common nutritional disorder that results in decreased yield and poor fruit quality in apple production. As a highly active synthetic analog of brassinosteroids, brassinolide (BL) plays numerous roles in plant responses to abiotic stresses. However, its role in Fe deficiency stress in apple plants has never been reported. Herein, we found that the exogenous application of 0.2 mg L-1 BL could significantly enhance the tolerance of apple seedlings to Fe deficiency stress and result in a low etiolation rate and a high photosynthetic rate. The functional mechanisms of this effect were also explored. We found that first, exogenous BL could improve Fe absorption through the mechanism I strategy. BL induced the activity of H+-ATPase and the expression of MhAHA family genes, resulting in rhizosphere acidification. Moreover, BL could enhance the activity of Fe chelate reductase and absorb Fe through direct binding with the E-box of the MhIRT1 or MhFRO2 promoter via the transcription factors MhBZR1 and MhBZR2. Second, exogenous BL alleviated osmotic stress by increasing the contents of osmolytes (proline, solution proteins, and solution sugar) and scavenged reactive oxygen species by improving the activities of antioxidant enzymes. Lastly, exogenous BL could cooperate with other endogenous plant hormones, such as indole-3-acetic acid, isopentenyl adenosine, and gibberellic acid 4, that respond to Fe deficiency stress indirectly. This work provided a theoretical basis for the application of exogenous BL to alleviate Fe deficiency stress in apple plants.