Rhombomeres are segmental units of the hindbrain that are separated from each other by a specialised zone of boundary cells. Retinoic acid application to a recently segmented hindbrain leads to disappearance of posterior rhombomere boundaries. Boundary loss is preceded by changes in segmental expression of Krox-20 and Cek-8 and followed by alterations in Hox gene expression. The characteristic morphology of boundary cells, their expression of follistatin and the periodic accumulation of axons normally associated with boundaries are all lost. In the absence of boundaries, we detect no change in anteroposterior dispersal of precursor cells and, in most cases, no substantial cell mixing between former rhombomeric units. This is consistent with the idea that lineage restriction can be maintained by processes other than a mechanical barrier composed of boundary cells. Much of the early organisation of the motor nuclei appears normal despite the loss of boundaries and altered Hox expression.

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