Progressive spatial restriction of Sek-1 and Krox-20 gene expression during hindbrain segmentation

After segmentation of the vertebrate hindbrain, expression of the zinc-finger gene Krox-20 and the receptor tyrosine kinase gene Sek-1 is precisely restricted to rhombomeres (r) 3 and 5. This precise segmental expression is likely to reflect a critical requirement for these rhombomeres to acquire a distinct and homogeneous identity and raises the question as to how this relates to the intermingling and restriction of cell movement during segmentation. We have analysed Krox-20 and Sek-1 expression in the mouse and chick hindbrain at single-cell resolution using whole-mount in situ hybridisation and immunocytochemistry. We find that, in the mouse, the presumptive r3 and r5 expression domains each arise as narrow stripes that then broaden, suggestive of a recruitment of cells to an r3/r5 identity and/or a segmental regulation of cell proliferation. In addition, we find that expression of these genes initially occurs in fuzzy domains, and that these are progressively restricted to segmental domains, although occasional "violating" cells are observed even after segmentation. We propose that the establishment and maintenance of these segmental domains may involve both a dynamic regulation of r3/r5 identity and the restriction of cell movement across rhombomere boundaries.