Role of differential cell proliferation in the tail bud in aberrant mouse neurulation

In the mouse mutant curly tail, the phenotypes spina bifida and curled tail result from a delay in closure of the posterior neuropore (PNP). At the developmental stage when this delay can first be recognized, the caudal region of the embryo demonstrates a transiently enhanced curvature of the body axis which likely inhibits elevation, convergence, and fusion of the neural folds. The enhanced curvature is thought to be the result of a decreased proliferation in the ventrally located gut endoderm and notochord, together with a normal proliferation of the overlying neuroepithelium of the PNP. However, the proliferation defect and the enhanced curvature were originally demonstrated at the same developmental stage, while it is expected that reduced proliferation should precede enhanced curvature and delayed PNP closure. The caudal region originates from the tail bud and we therefore propose that the enhanced curvature is induced by a disturbed dorso-ventral proliferation pattern in the tail bud. Using flow cytometry, proliferation patterns were determined separately for the dorsal and ventral halves of the tail bud of curly tail and of control embryos as well as of recombinant embryos having the curly tail phenotype with a genetic background which is matched to the BALB/c control strain. In general, it appeared that about half of the cell cycle duration in tail bud cells was occupied by S phase, about 40% by G0/G1 and the rest by G2/M. For the control embryos, no dorso-ventral differences in relative phase duration were demonstrated. However, curly tail and recombinant embryos at the 21-25 somite stage, prior to the onset of enhanced curvature, exhibited ventrally a higher proportion of G0/G1 phase cells than dorsally, and a complementary relationship for S phase cells. We interpret these observations as indicating a prolonged G1 phase at the ventral side of the tail bud, resulting in a prolongation of the cell cycle and thus a decreased proliferation. In 26-30 somite stage embryos, prior to the normalization of curvature in curly tail embryos, the dorso-ventral proliferation balance was re-established. We conclude that a reduced proliferation in the ventral part of the tail bud of the curly tail embryo precedes both the onset of enhanced curvature and the previously observed reduction in proliferation of the hindgut and notochord, and is a likely candidate for an early event in the pathogenetic sequence leading to the curly tail phenotype.