Neurons in the inferior olivary nuclei (IO neurons) send climbing fibers to Purkinje cells to elicit functions of the cerebellum. IO neurons and Purkinje cells are derived from neural progenitors expressing the proneural gene ptf1a. In this study, we found that the homeobox gene gsx2 was co-expressed with ptf1a in IO progenitors in zebrafish. Both gsx2 and ptf1a zebrafish mutants showed a strong reduction or loss of IO neurons. The expression of ptf1a was not affected in gsx2 mutants and vice versa. In IO progenitors, the ptf1a mutation increased apoptosis whereas the gsx2 mutation did not, suggesting that ptf1a and gsx2 are regulated independently of each other and have distinct roles. The fibroblast growth factors (Fgf) 3/8a and retinoic acid signals negatively and positively, respectively, regulated gsx2 expression and thereby the development of IO neurons. mafba and hox genes are at least partly involved in the Fgf- and retinoic acid-dependent regulation of IO neuronal development. Our results indicate that gsx2 mediates the rostro-caudal positional signals to specify the identity of IO neurons from ptf1a-expressing neural progenitors.

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Background: Although the cell cycle and cell differentiation should be coordinately regulated to generate a variety of neurons in the brain, the molecules that are involved in this coordination still remain largely unknown. In this study, we analyzed the roles of a nuclear protein Cfdp1, which is thought to be involved in chromatin remodeling, in zebrafish neurogenesis. Results: Zebrafish cfdp1 mutants maintained the progenitors of granule cells (GCs) in the cerebellum, but showed defects in their differentiation to GCs. cfdp1 mutants showed an increase in phospho-histone 3 (pH 3)-positive cells and apoptotic cells, as well as a delayed cell cycle transition from the G2 to the M phase in the cerebellum. The inhibition of tp53 prevented apoptosis but not GC differentiation in the cfdp1 mutant cerebellum. A similar increase in apoptotic cells and pH 3-positive cells, and defective cell differentiation, were observed in the cfdp1 mutant retina. Although mitotic spindles formed, mitosis was blocked before anaphase in both the cerebellum and retina of cfdp1 mutant larvae. Furthermore, expression of the G2/mitotic-specific cyclin B1 gene increased in the cfdp1 mutant cerebellum.Conclusions: Our findings suggest that Cfdp1 regulates the cell cycle of neural progenitors, thereby promoting neural differentiation in the brain.

Cerebellar neurons such as Purkinje cells (PCs) and granule cells (GCs) are differentiated from neural progenitors expressing proneural genes. Zebrafish mutants of proneural genesptf1aandneurogenin1showed a reduction or loss of PCs, GABAergic interneurons (INs), and reduced expression of GC progenitor genesatoh1a/b/c. Lineage tracing revealed that theptf1a-expressing progenitors gave rise to PCs, INs, and a part of GCs in zebrafish. These data indicate that theptf1a/neurognin1-expressing neural progenitors can generate a variety of cerebellar neurons. In this study, we found that genes encoding transcriptional regulators Foxp1b and Foxp4, as well as Skor1b and Skor2, which are reportedly expressed in PCs, were not expressed inptf1a;neurogenin1 mutants.foxp1b;foxp4mutants showed a strong reduction in PCs, whileskor1b;skor2mutants completely lacked PCs but instead displayed an increase in immature GCs. Misexpression ofskor2in GC progenitors expressingatoh1csuppressed GC fate. These data indicate that Foxp1b/4 and Skor1b/2 function as key transcriptional regulators in the initial step of PC differentiation fromptf1a/neurogenin1-expressing neural progenitors, while Skor1b and Skor2 control PC differentiation by suppressing their differentiation into GCs.

The homeobox gene gsx2 mediates rostro-caudal positional signaling to specify the identify of neurons in the inferior olivary nuclei from neural progenitors expressing the proneural gene ptf1a. 2 ABSTRACTNeurons in the inferior olivary nuclei (IO neurons) send climbing fibers to Purkinje cells to elicit functions of the cerebellum. IO neurons and Purkinje cells are derived from neural progenitors expressing the proneural gene ptf1a. In this study, we found that the homeobox gene gsx2 was co-expressed with ptf1a in IO progenitors in zebrafish.Both gsx2 and ptf1a zebrafish mutants showed a strong reduction or loss of IO neurons.The expression of ptf1a was not affected in gsx2 mutants and vice versa. In IO progenitors, the ptf1a mutation increased apoptosis whereas the gsx2 mutation did not, suggesting that ptf1a and gsx2 are independently regulated and have distinct roles. The fibroblast growth factors (Fgf) 3/8a and retinoic acid signals negatively and positively, respectively, regulated gsx2 expression and thereby the development of IO neurons.mafba and hox genes are at least partly involved in the Fgf-and retinoic acid-dependent regulation of IO neuronal development. Our results indicate that gsx2 mediates the rostro-caudal positional signals to specify the identity of IO neurons from ptf1a-expressing neural progenitors. K., Suster, M. L., Mizusawa, K., Nagayoshi, S., Kotani, T., Urasaki, A., Kishimoto, Y., Hibi, M. and Kawakami, K. (2008). Genetic dissection of neural circuits by Tol2 transposon-mediated Gal4 gene and enhancer trapping in zebrafish. . Anatomy of zebrafish cerebellum and screen for mutations affecting its development. Dev . Biol. 330, 406-426. Begemann, G., Marx, M., Mebus, K., Meyer, A. and Bastmeyer, M. (2004). Beyond the neckless phenotype: influence of reduced retinoic acid signaling on motor neuron development in the zebrafish hindbrain. Dev. Biol. 271, 119-129. Asakawa,