A SERIES OF NO ISTHMUS (NOI) ALLELES OF THE ZEBRAFISH PAX2.1 GENE REVEALS MULTIPLE SIGNALING EVENTS IN DEVELOPMENT OF THE MIDBRAIN-HINDBRAIN BOUNDARY

Generation of cell diversity in the vertebrate central nervus system s tarts during gastrulation stages in the ectodermal germ layer and invo lves specialized cell groups, such as the organizer located at the mid brain-hindbrain boundary (MHB). Mutations in the zebrafish no isthmus (noi) gene alter development of the MHB, and affect the pax2.1 gene (f ormerly pax(zf-b)). Analysis of the structure of pax2.1 reveals at lea st 12 normal splice variants, The noi alleles can be arranged, by mole cular and phenotypic criteria, into a series of five alleles of differ ing strength, ranging from a null allele to weak alleles, In keeping w ith a role in development of the MHB organizer, gene expression is alr eady affected in the MHB primordium of the gastrula neural ectoderm in noi mutants. eng3 activation is completely and eng2 activation is str ongly dependent on noi function. In contrast, onset of wnt1,fgf8 and h er5 expression occurs normally in the null mutants, but is eliminated later on. Our observations suggest that three signaling pathways, invo lving pax2.1, wntl and fgf8, are activated independently in early ante rior-posterior patterning of this area. In addition, analysis of the a llelic series unexpectedly suggests that noi activity is also required during dorsal-ventral patterning of the MHB in somitogenesis stages, and possibly in a later eng expression phase. We propose that noi/pax2 .1 participates in sequential signaling processes as a key integrator of midbrain-hindbrain boundary development.