A complex suite of cell behaviours drives morphogenesis of the mammalian limb. While various transcription factors that regulate limb pattern formation have been identified, the link between transcriptional regulation and morphogenetic cell behaviour remains incompletely understood. Sevan Hopyan, Chi-chung Hui and colleagues now provide evidence that the Iroquois transcription factors, well known as regulators of pattern formation, control early limb bud morphogenesis via the cell cycle. Early limb buds in Irx3/5 mutants are short and bulbous, and this shape deformation is associated with changes in mesodermal cell behaviour: cell movements are disoriented and reduced, intercalation is diminished, cell cycle time is increased, and a high proportion of mitotic cells exhibit chromatin bridges. The appearance of these chromatin bridges is often followed by chromatin fragmentation, and even where mitosis is completed, planes of division are disoriented. Using BioID in cell culture, the authors find that IRX3/5 bind to various cell cycle regulators, including the cohesin subunits SMC1 and SMC3, and the transcription factor CUX1; physical proximity was also demonstrated in limb buds with co-immunoprecipitation, STED imaging and proximity ligation assays. Irx3/5 mutants show a reduced abundance of cohesin subunits and CUX1, and, finally, early Cux1/2−/− mutant limb buds phenocopy Irx3/5 mutants (albeit showing a broader DV axis), and also show chromatin bridges. Thus, IRX3/5 shape the limb bud via chromatid segregation.
