Genetic Editing of the zebrafish clock gene reveals previously unexpected molecular and phenotypic effects

The histone acetyltransferase (HAT) activity of the circadian protein CLOCK is thought to couple rhythmic transcription with chromatin remodeling, yet it in vivo function remains poorly understood. Here, we establish innovative zebrafish models to dissect the physiological relevance of CLOCKB acetyltransferase domain. Using CRISPR–Cas9 and base-editing approaches, we generated three complementary clockb mutant lines: a point mutation within the HAT domain (FEN), a complete knockout (CBKO), and in-frame deletion removing the entire HAT domain (INF). Loss of Clockb HAT function impaired histone H3 acetylation and disrupted circadian gene expression, demonstrating that Clockb is required to sustain rhythmic chromatin states. Beyond circadian control, clockb deficiency caused striking developmental alterations affecting eye formation, retinal organization, locomotor behavior, and pigment cell differentiation, revealing that Clockb acetyltransferase activity is essential for developmental homeostasis.At the systemic level, clockb mutants displayed consistent dysregulation of thyroid-axis genes, suggesting that Clockb links circadian chromatin regulation to endocrine signaling pathways controlling growth and pigmentation. Together, these findings identify Clockb as a chromatin-dependent regulator that synchronizes circadian transcription with endocrine and developmental programs. The zebrafish models developed here provide a unique in vivo platform to uncover how intrinsic HAT activity of circadian factors shapes vertebrate physiology.