Carotenoids are important pigments in pepper fruits. The colors of each pepper are mainly determined by the composition and content of carotenoid. The ‘ZY’ variety, which has yellow fruit, is a natural mutant derived from a branch mutant of ‘ZR’ with different colors. ZY and ZR exhibit obvious differences in fruit color, but no other obvious differences in other traits. To investigate the main reasons for the formation of different colored pepper fruits, transcriptome and metabolome analyses were performed in three developmental stages (S1–S3) in two cultivars. The results revealed that these structural genes (PSY1, CRTISO, CCD1, CYP97C1, VDE1, CCS, NCED1 and NCED2) related to carotenoid biosynthesis were expressed differentially in the two cultivars. Capsanthin and capsorubin mainly accumulated in ZR and were almost non-existent in ZY. S2 is the fruit color-changing stage; this may be a critical period for the development of different color formation of ZY and ZR. A combination of transcriptome and metabolome analyses indicated that CCS, NCED2, AAO4, VDE1 and CYP97C1 genes were key to the differences in the total carotenoid content. These new insights into pepper fruit coloration may help to improve fruit breeding strategies.

Plant homeodomain (PHD) transcription factor genes are involved in plant development and in a plant’s response to stress. However, there are few reports about this gene family in peppers (Capsicum annuum L.). In this study, the pepper inbred line “Zunla-1” was used as the reference genome, and a total of 43 PHD genes were identified, and systematic analysis was performed to study the chromosomal location, evolutionary relationship, gene structure, domains, and upstream cis-regulatory elements of the CaPHD genes. The fewest CaPHD genes were located on chromosome 4, while the most were on chromosome 3. Genes with similar gene structures and domains were clustered together. Expression analysis showed that the expression of CaPHD genes was quite different in different tissues and in response to various stress treatments. The expression of CaPHD17 was different in the early stage of flower bud development in the near-isogenic cytoplasmic male-sterile inbred and the maintainer inbred lines. It is speculated that this gene is involved in the development of male sterility in pepper. CaPHD37 was significantly upregulated in leaves and roots after heat stress, and it is speculated that CaPHD37 plays an important role in tolerating heat stress in pepper; in addition, CaPHD9, CaPHD10, CaPHD11, CaPHD17, CaPHD19, CaPHD20, and CaPHD43 were not sensitive to abiotic stress or hormonal factors. This study will provide the basis for further research into the function of CaPHD genes in plant development and responses to abiotic stresses and hormones.

Fruit color serves as a crucial visual indicator in chili peppers and is closely linked to the bioactive components that determine their economic and nutritional value. However, the specific components and potential molecular mechanisms that impact fruits’ development and color changes are less thoroughly understood. Here, we utilized three chili pepper varieties (CS03, CS29, and L816) at different developmental stages (young fruit stage, turning color stage, and mature stage) as research materials and integrated transcriptome and non-targeted metabolome analyses to explore the variation in bioactive components and color to explain the molecular regulatory mechanisms underlying different colors of chili peppers during the young fruit stage. Our results showed that flavonoids were the most enriched differential metabolites; aromadendrin 4′-glucoside, diospyrin, precarthamin, kaempferol-3-O-rutinoside, and kaempferol-3-O-Glucoside were significantly enriched in the young fruit stage of pepper CS03; and cyanidin, delphinidin, and cyanidin 3-glucoside were major contributors to the color formation. The upregulation of anthocyanin was related to the structural genes CaC4H, Ca4CL, CaCHS, CaF3H, CaANS, and CaUFGT, and key transcription factors such as CaMYBs and CabHLHs may have contributed to the differential accumulation of anthocyanins in CS03; in addition, RT-qPCR validation was correlated with anthocyanins, but also with flavonoids. This article mainly focuses on the changes in chili pigments, particularly anthocyanins, and explores the molecular mechanisms involved. This provides a reference for research on color in solanaceae vegetables and lays a theoretical foundation for further research on the bioactive components of chili peppers, as well as for optimizing harvesting practices and dietary recommendations.

Anthocyanins are important flavonoid compounds in plants that are associated with the color formation and antioxidant activity of flowers, fruits, and other organs. Ultraviolet B radiation (UV-B) is one of the key environmental factors that influence anthocyanin accumulation in plants and HY5 is involved in plant photomorphogenesis. However, the molecular mechanism of the signal network of UV-B regulating anthocyanin biosynthesis in capsicum via HY5 remains unclear. In this study, we identified the transcription factor CaHY5, which mediates UV-B signaling, and demonstrated its regulatory role in anthocyanin biosynthesis in purple pepper (Capsicum annuum L.). The results showed that there were photoresponsive and hormone-responsive elements on the CaHY5 promoter that responded to UV-B, indoleacetic acid, salicylic acid, 6-benzyladenine, abscisic acid, and melatonin treatments. UV-B treatment induced the expression of CaHY5 and anthocyanin structural genes. CaHY5 gene-silenced positive plants showed different degrees of the yellowing phenomenon, which affected the expression of the anthocyanin biosynthesis structural gene. The expression levels of anthocyanin biosynthesis-related genes in CaHY5-silenced positive plants increased considerably. This study provides insights into the role of CaHY5 in UV-B-induced anthocyanin biosynthesis in purple pepper.

The bHLH transcription factors play a crucial regulatory role in plant growth and development. In this study, the CcbHLH68 gene was cloned from the pepper cultivar ‘Shuanla’. Subsequent bioinformatics analysis, subcellular localization, expression pattern profiling, along with yeast one-hybrid and dual-luciferase reporter assays, were conducted to preliminarily elucidate its regulatory mechanism in capsaicinoid biosynthesis. The results revealed that the visualization of upstream cis-elements of CcbHLH68 suggests its potential regulation by hormones. Furthermore, subcellular localization experiments confirmed that the CcbHLH68 protein is localized in the nucleus. Expression analysis of CcbHLH68 across different tissues by qRT-PCR identified its predominant expression in the placenta at 30 days post-anthesis. Further experimental evidence from both gene silencing and transient overexpression assays demonstrated a positive correlation between CcbHLH68 and the expression of multiple capsaicinoid biosynthetic genes. When it was silenced or transiently overexpressed, the content of capsaicinoids decreased by 40.9% or increased by 113.7%, respectively. Yeast one-hybrid and dual-luciferase reporter assays confirmed that CcbHLH68 can directly bind to the CcCOMT promoter and activate its transcription. In summary, this study preliminarily reveals the molecular mechanism by which CcbHLH68 participates in capsaicinoid biosynthesis through regulating the expression of key genes in the biosynthetic pathway, thereby providing a theoretical foundation for enhancing capsaicinoid content via molecular breeding.