Two B-Box Proteins, GhBBX21 and GhBBX24, Antagonistically Modulate Anthocyanin Biosynthesis in <i>R1</i> Cotton
2025
Shuyan Li | Kunpeng Zhang | Chenxi Fu | Chaofeng Wu | Dongyun Zuo | Hailiang Cheng | Limin Lv | Haiyan Zhao | Jianshe Wang | Cuicui Wu | Xiaoyu Guo | Guoli Song
The red plant phenotype of <i>R1</i> cotton is a genetic marker produced by light-induced anthocyanin accumulation. <i>GhPAP1D</i> controls this trait. There are two 228 bp tandem repeats upstream of <i>GhPAP1D</i> in <i>R1</i> cotton. In this study, GUS staining assays in transgenic <i>Arabidopsis thaliana</i> (L.) Heynh. demonstrated that tandem repeats in the <i>GhPAP1D</i> promoter-enhanced transcriptional activity. <i>GhPAP1D</i> is a homolog of <i>A. thaliana AtPAP1</i>. <i>AtPAP1’s</i> expression is regulated by photomorphogenesis-related transcription factors such as AtHY5 and AtBBXs. We identified the homologs of <i>A. thaliana AtHY5</i>, <i>AtBBX21</i>, and <i>AtBBX24</i> in <i>R1</i> cotton, designated as <i>GhHY5</i>, <i>GhBBX21</i>, and <i>GhBBX24</i>, respectively. Y1H assays confirmed that GhHY5, GhBBX21, and GhBBX24 each bound to the <i>GhPAP1D</i> promoter. Dual-luciferase reporter assays revealed that GhHY5 weakly activated the promoter activity of <i>GhPAP1D</i>. Heterologous expression assays in <i>A. thaliana</i> indicated that <i>GhBBX21</i> promoted anthocyanin accumulation, whereas <i>GhBBX24</i> had the opposite effect. Dual-luciferase assays showed GhBBX21 activated <i>GhPAP1D</i> transcription, while GhBBX24 repressed it. Further study indicated that GhHY5 did not enhance GhBBX21-mediated transcriptional activation of <i>GhPAP1D</i> but alleviates GhBBX24-induced repression. Together, our results demonstrate that GhBBX21 and GhBBX24 antagonistically regulate anthocyanin accumulation in <i>R</i>1 cotton under GhHY5 mediation, providing insights into light-responsive anthocyanin biosynthesis in cotton.
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