Specific mitotic events drive left-right organizer development.

Cell proliferation is crucial for tissue development. Here, we investigate its role in the left-right organizer, which establishes the left-right axis. In zebrafish, we mapped mitotic events in Kupffer's vesicle (KV) and identified an anteriorly enriched, FGF-dependent mitotic pattern. Laser ablation of mitotic cells and pericentrin-null mutants, both reducing mitotic events, resulted in smaller lumens, confirming that cell division is essential for KV development. Pericentrin-null mutants also exhibited defects in leftward cardiac jogging, indicative of KV dysfunction. Using a KV-specific fluorescent microtubule marker, we found that the KV rosette is a transient, centrally organized cluster interconnected by cytokinetic bridges and containing microtubule bundles. This structure emerges after the first four divisions and precedes lumen formation. Mitotic events during KV rounding coincide with rosette formation, spindle rotation and cell extrusion, likely driven by increased packing. Eliminating the first four mitotic events disrupted rosette formation and prevented normal KV rounding. These findings demonstrate that mitotic events are crucial for KV development, with cell division timing shaping KV architecture and function.