Drought stress caused by water deficit reduces plant productivity in many regions of the world. In plants, basic helix–loop–helix (bHLH) transcription factors regulate a wide range of cellular activities related to growth, development and stress response; however, the role of tomato SlbHLHs in drought stress responses remains elusive. Here, we used reverse genetics approaches to reveal the function of SlbHLH96, which is induced by drought and abscisic acid (ABA) treatment. We found that SlbHLH96 functions as a positive regulator of drought tolerance in tomato. Overexpression of SlbHLH96 in tomato improves drought tolerance by stimulating the expression of genes encoding antioxidants, ABA signaling molecules and stress-related proteins. In contrast, silencing of SlbHLH96 in tomato reduces drought tolerance. SlbHLH96 physically interacts with an ethylene-responsive factor, SlERF4, and silencing of SlERF4 in tomato also decreases drought tolerance. Furthermore, SlbHLH96 can repress the expression of the ABA catabolic gene, SlCYP707A2, through direct binding to its promoter. Our results uncover a novel mechanism of SlbHLH96-mediated drought tolerance in tomato plants, which can be exploited for breeding drought-resilient crops.

The R2R3-MYB is a large gene family involved in various plant functions, including carotenoid biosynthesis. However, this gene family lacks a comprehensive analysis in wolfberry (Lycium barbarum L.) and other Solanaceae species. The recent sequencing of the wolfberry genome provides an opportunity for investigating the organization and evolutionary characteristics of R2R3-MYB genes in wolfberry and other Solanaceae species. A total of 610 R2R3-MYB genes were identified in five Solanaceae species, including 137 in wolfberry. The LbaR2R3-MYB genes were grouped into 31 subgroups based on phylogenetic analysis, conserved gene structures, and motif composition. Five groups only of Solanaceae R2R3-MYB genes were functionally divergent during evolution. Dispersed and whole duplication events are critical for expanding the R2R3-MYB gene family. There were 287 orthologous gene pairs between wolfberry and the other four selected Solanaceae species. RNA-seq analysis identified the expression level of LbaR2R3-MYB differential gene expression (DEGs) and carotenoid biosynthesis genes (CBGs) in fruit development stages. The highly expressed LbaR2R3-MYB genes are co-expressed with CBGs during fruit development. A quantitative Real-Time (qRT)-PCR verified seven selected candidate genes. Thus, Lba11g0183 and Lba02g01219 are candidate genes regulating carotenoid biosynthesis in wolfberry. This study elucidates the evolution and function of R2R3-MYB genes in wolfberry and the four Solanaceae species.

Light-harvesting chlorophyll a/b-binding (LHC) proteins, as the antenna complex, collect and transfer light energy to the reaction centers of PSII. They are crucial for abiotic stress responses, especially in the photoprotection under cold stress. However, members of the LHC gene family in tomato (Solanum lycopersicum L.) have not yet been identified. In this study, a total of 39 SlLHC proteins containing the chlorophyll a/b binding domain or light-harvesting-like domain were identified, and classified into four subfamilies: Lhc, Lil, PsbS, and FCII. Further qRT-PCR analysis showed SlLhcb1.11 was inhibited and SlELIP1 was induced at low temperature (4 °C). Subsequently, the result of VIGS experiment showed that silencing SlLhcb1.11 or SlELIP1 genes resulted in lighter leaf color, reduced chlorophyll content, compromised photosynthesis, and decreased cold tolerance in tomato plants. These findings offer novel insights into the structure and function of SlLHC genes, thereby contributing to genetic resources for the development of cold-tolerant tomato germplasm.