Fuente: PubMed "pollen" Plant Sci. 2026 Apr 17:113148. doi: 10.1016/j.plantsci.2026.113148. Online ahead of print.ABSTRACTBlueberry (Vaccinium ashei) characterizes in extremely rich anthocyanins. It has been found that some bHLHs play a crucial function in anthocyanin biosynthesis, however, the available information on the anthocyanin accumulation as regulated by VdbHLH genes has been very limited in blueberry fruits. In the present study, 68 VdbHLH genes were assessed in rabbiteye blueberry by transcriptome analysis, and classified into 23 subfamilies, among which VdbHLH60, a member from the IIIf subfamily, was synchronically up-regulated with the accumulation of anthocyanins, Subsequently, the VdbHLH60-overexpressing tomato lines were obtained, and fruit color of the trangenic lines were significantly deepened, with obviously higher anthocyanin contents in comparison with this of the wild-type (WT). Conversely, after silencing VdbHLH60 in blueberry fruits, the expression of VdMYB55 and anthocyanin-related function genes were significantly reduced, and the total anthocyanin content (TAC) was remarkably decreased; the silenced fruits remained green. With GUS staining assays, it was found that pro-VdbHLH60 appeared blue in the stem meristems and pollen sites of transgenic Arabidopsis thaliana, indicating that pro-VdbHLH60 functioned to activate the expression of downstream genes. Yeast one-hybrid (Y1H) and dual-Luciferase assays demonstrated that VdbHLH60 could bind to the promoter of VdF3'5'H to positively regulated the expression of this gene. Meanwhile, pro-VdF3'5'H appeared blue in the stem meristematic tissue, pollen and pod of transgenic A. thaliana, revealing that the pro-VdF3'5'H exhibited strong initiating activity in anthocyanin biosynthesis. Further, yeast two-hybrid (Y2H), bimolecular fluorescence complementation (BiFC), and dual-Luciferase assays substantially verified the interaction between VdbHLH60 and VdMYB55, which considerably up-regulated VdF3'5'H. Therefore, VdbHLH60-VdMYB55 -VdF3'5'H module positively regulates the anthocyanin biosynthesis of rabbiteye blueberry fruits, which may better understand the regulatory network of anthocyanin accumulation in this fruit species.PMID:42001965 | DOI:10.1016/j.plantsci.2026.113148