A multifaceted module of BRI1 ETHYLMETHANE SULFONATE SUPRESSOR1 (BES1)-MYB88 in growth and stress tolerance of apple.

The R2R3 transcription factor MdMYB88 has previously been reported to function in biotic and abiotic stress responses. Here, we identify BRI1 ETHYLMETHANE SULFONATE SUPRESSOR1 (MdBES1), a vital component of brassinosteroid (BR) signaling in apple (Malus × domestica) that directly binds to the MdMYB88 promoter, regulating the expression of MdMYB88 in a dynamic and multifaceted mode. MdBES1 positively regulated expression of MdMYB88 under cold stress and pathogen attack, but negatively regulated its expression under control and drought conditions. Consistently, MdBES1 was a positive regulator for cold tolerance and disease resistance in apple, but a negative regulator for drought tolerance. In addition, MdMYB88 participated in BR biosynthesis by directly regulating the BR biosynthetic genes DE ETIOLATED 2 (MdDET2), DWARF 4 (MdDWF4), and BRASSINOSTEROID 6 OXIDASE 2 (MdBR6OX2). Applying exogenous BR partially rescued the erect leaf and dwarf phenotypes, as well as defects in stress tolerance in MdMYB88/124 RNAi plants. Moreover, knockdown of MdMYB88 in MdBES1 overexpression (OE) plants decreased resistance to a pathogen and C-REPEAT BINDING FACTOR1 expression, whereas overexpressing MdMYB88 in MdBES1 OE plants increased expression of SQUAMOSA PROMOTER BINDING PROTEIN-LIKE 3 （MdSPL3） and BR biosynthetic genes, suggesting that MdMYB88 contributes to MdBES1 function during BR biosynthesis and the stress response. Taken together, our results reveal multifaceted regulation of MdBES1 on MdMYB88 in BR biosynthesis and stress tolerance.

Apple TIME FOR COFFEE contributes to freezing tolerance by promoting unsaturation of fatty acids.

Freezing stress is a major environmental factor that threatens the growth and development of fruit trees. MdMYB88 and its paralogue MdMYB124 have been identified as pivotal regulators in apple (Malus × domestica) freezing stress tolerance. Here, we demonstrated that a target of MdMYB88 and MdMYB124, TIME FOR COFFEE (TIC), contributes to freezing tolerance in apple. MdMYB88 and MdMYB124 directly bound the MdTIC promoter and positively regulated its expression under cold conditions. MdTIC RNAi plants displayed reduced freezing tolerance when MdTIC expression was repressed. Moreover, MdTIC RNAi plants lowered antioxidant enzyme activity. Transcriptome profiling revealed altered expression of cold-responsive genes in MdTIC RNAi plants under cold conditions, including MdPLC2, MdMKK2, and MdICE1. We also discovered that disordered MdTIC expression changed the saturation of fatty acids. Taken together, our data suggest that MdTIC is required for apple to tolerate freezing by mediating the expression of cold-responsive genes and fatty acid composition.

Genome-wide identification and expression analysis of brassinosteroid biosynthesis and metabolism genes regulating apple tree shoot and lateral root growth.

In plants, brassinosteroid biosynthesis and metabolism genes affect endogenous brassinosteroid metabolic processes as well as stem and root growth. However, there is little information available regarding these genes in apple. In this study, 22 brassinosteroid biosynthesis and metabolism genes were identified in apple (Malus domestica). These genes were named according to their chromosomal locations and the Arabidopsis thaliana homologs. Their conserved characteristic domains, evolutionary relationships, syntenic relationships, chemical characteristics, gene/protein structures, interactions among the encoded proteins, promoter sequences, and functions were investigated. These 22 genes were clustered with their A. thaliana homologs based on bioinformatics analyses, which suggested they are functionally similar in apple and A. thaliana. Tissue-specific expression levels revealed that most of these genes are important for stem growth and development, while several of these genes affect lateral root formation. The transcription patterns of these genes in shoot tips were investigated following diverse treatments [brassinosteroid (shoot tips and roots), brassinazole, auxin, and temperature]. Gene expression levels were also examined in different grafting combinations ('Nagafu No. 2'/Malling 9 and 'Nagafu No. 2'/'Nagafu No. 2') and shoot varieties ('Yanfu No. 6' and 'Nagafu No. 2'). The results indicated that these genes may be involved in apple stem and root growth. The comprehensive genome-wide analysis of brassinosteroid biosynthesis and metabolism genes presented herein may be useful for breeding new apple cultivars with increased vigor. The data also represent a rich genetic resource for future apple gene functional investigations that may have implications for the genetic improvement of apple tree species.