Coloring in apple fruit due to anthocyanin accumulation is inhibited by high temperature; however, the underlying mechanism remains unclear. In the present study, total anthocyanin and cyanidin 3-galactoside contents were determined and compared between cv. ‘Redchief Delicious’ apple fruits at 25 °C and 35 °C treatments. The high temperature (35 °C) treatment substantially decreased total anthocyanin and cyanidin 3-galactoside contents. The transcriptomes of 25 °C- and 35 °C-treated apples were analyzed by high-throughput RNA sequencing. A total of 8354 differentially expressed genes (DEGs) were detected at four time points corresponding to the two temperature treatments. The up-regulated DEGs were annotated using GO as well as KEGG databases. A network module of 528 genes (including 21 transcription factors) most associated with the total anthocyanin and cyanidin 3-galactoside contents was constructed by weighted correlation network analysis (WGCNA). In the WGCNA module, we unearthed a LOB domain-containing gene designated as MdLBD37. The expression of MdLBD37 was sharply up-regulated by high temperature and negatively correlated with the total anthocyanin and cyanidin 3-galactoside contents. Overexpression of MdLBD37 in apple fruit and calli decreased the expression of anthocyanin biosynthetic genes, such as MdCHI, MdCHS, MdF3H, MdANS, MdDFR, and MdUFGT, along with anthocyanin accumulation. Our results suggested that MdLBD37 significantly influenced the high-temperature inhibition of anthocyanin accumulation in apples. The findings shed more light on the mechanism of anthocyanin inhibition during high-temperature stress in apples.

Apples (Malus domestica Borkh) are important fruits in China; however, salt stress is severe in northern regions, and the key to plant resistance to salt stress lies in the rootstock. Therefore, it is necessary to explore rootstocks with strong salt resistance for the development of the apple industry. This study used tissue culture seedlings of six apple rootstocks, namely, ‘71-3-150’, ‘54-118’, ‘M9T337’, ‘GM256’, ‘ML176’, and ‘ML2’, as experimental materials. The seedlings were treated with a medium containing 150 mM NaCl, and the physiological indicators and related gene expression responses of several rootstocks were studied after salt stress. The results showed that salt stress affects the growth of both the aboveground and underground parts of plants. Through physiological indicators and the related gene expression responses of rootstocks, it was observed that salt stress significantly increased Na⁺ contents in different rootstocks. Simultaneously, the activity of various antioxidant enzymes and the expression levels of related genes also increased. In summary, by analyzing the parameters of various physiological indicators, it can be concluded that among the studied rootstocks, the ‘71-3-150’ and ‘54-118’ rootstocks have the strongest resistance to salt stress, while the ‘M9T337’ and ‘GM256’ rootstocks exhibit moderate resistance, and the ‘ML176’ and ‘ML2’ rootstocks have the weakest resistance.

Apple ring rot, caused by Botryosphaeria dothidea, represents a major challenge for apple cultivation worldwide. However, the genetic factors that confer resistance in apples are not well understood. This study utilized time-series transcriptome profiling, combined with weighted gene co-expression network analysis and previously identified quantitative trait loci, leading to the identification of MdNHL8 as a candidate gene linked to resistance. A genome-wide examination of the apple NDR1/HIN1-like gene family revealed 57 members with diverse conserved domains, confirming that MdNHL8 features a distinct RBR-type E3 ligase domain. Subcellular localization experiments showed that MdNHL8 is localized in the nucleus and plasma membrane. Functional assays indicated that overexpression of MdNHL8 enhances resistance to apple ring rot and significantly elevates the expression of MdPR1, whereas silencing this gene does not enhance resistance and exhibits a trend toward increased susceptibility. Furthermore, additional transcriptome sequencing analysis of MdNHL8 transgenic calli highlighted a turquoise module closely linked to MdNHL8-mediated resistance, identifying MdCYCB3, MdEIX2, and MdABC4 as candidate downstream genes with expression patterns tightly correlated with MdNHL8. This research identifies MdNHL8 as a positive regulator in apple defense against B. dothidea and provides a basis for further exploring the regulatory network centered on MdNHL8 in disease resistance, offering insights for future breeding efforts aimed at enhancing resistance to apple ring rot.