News and Publications

Cerebellar Development in the Mouse: Genetics, Cell Biology, and Gene Expression

Our research goal is to understand the cell-cell interactions that lead to neuronal differentiation and synaptogenesis. We focus on the interrelated processes of development of afferent fibers, dendrites, and spines in the CNS. The cerebellum is an attractive model system because it has a limited number of cell types and a regular architecture and wiring pattern. The cerebellum undergoes extensive development postnatally in rodents, providing an accessible experimental setting for studying neuronal migration, differentiation, and synaptogenesis. In addition, these processes are perturbed in a number of mutant mouse strains.

Genetic and Genomics Approaches

In collaboration with scientists at the Genomics Institute of the Novartis Research Foundation, San Diego, California, we are using a genetic screen to detect new mouse mutants with abnormal cerebellar development. To date, 11 such mutants have been identified. The genes of 2 strains have been mapped, and gene identification is under way for the rest.

We are also using gene chips to obtain expression profiles. We are characterizing gene expression in the developing cerebellum in normal mice and comparing the results with gene expression in strains of ataxic mice with mutations in known genes. In this way, we will gain insight into the signaling pathways necessary for normal cerebellar development and also identify targets for intervention in cerebellar ataxia.

Cell Biology Approaches

We are using primary neuron cultures to explore the cell-cell interactions necessary for the development of granule and Purkinje cells. Purified Purkinje cells survive and produce axons but no mature dendrites. Adding back the cerebellar granule cells, which form synapses with the Purkinje cells, triggers the development of dendrites and spines on the Purkinje cells, recapitulating the steps of differentiation that occur in vivo. Using this culture system, we can characterize the roles of specific signal transduction pathways in the interaction of granule and Purkinje cells. Preliminary data indicated that BDNF/TrkB signaling regulates the number and morphology of spines on Purkinje cells. We will use the same in vitro system to test other signaling systems identified in the genetic and genomic approaches described.

The combination of these approaches provides new and exciting ways to identify the signal transduction pathways important for the survival, development, and synaptogenesis of neurons in the CNS.

Publications

Carlson, C.M., Dupuy, A.J., Fritz, S., Roberg-Perez, K.J., Fletcher, C.F., Largaespada, D.A. Transposon mutagenesis of the mouse germline. Genetics, in press.

FANTOM Consortium and RIKEN Genome Exploration Research Group Phase II Team. Analysis of the mouse transcriptome based on functional annotation of 60,770 full-length cDNAs. Nature 420:563, 2002.

Fletcher, C.F., Lennon, V.A. Do calcium channel autoantibodies cause cerebellar ataxia with Lambert-Eaton syndrome? Ann. Neurol. 53:5, 2003.

Gustincich, S., Arakawa, Y., Batalov, S., Beisel, K.W., Bono, H., Carninci, P., Fletcher, C.F., Grimmond, S., Hirokawa, N., Jarvis, E.D., Jegla, T., Kawasaka, Y., Miki, H., Raviola, E., Teasdale, R.D., Waki, K., Zimmer, A., Kawai, J., Hayashizaki, Y., Okazaki, Y. Analysis of the mouse transcriptome for genes involved in the function of the nervous system. Genome Res., in press.

Liu, L., Zwingman, T.A., Fletcher, C.F. In vivo analysis of voltage-dependent calcium channels. J. Bioenerg. Biomembr., in press.

Tottene, A., Fellin, T., Pagnutti, S., Luvisetto, S., Striessnig, J., Fletcher, C., Pietrobon, D. Familial hemiplegic migraine mutations increase Ca²⁺ influx through single human CaV2.1 channels and decrease maximal CaV2.1 current density in neurons. Proc. Natl. Acad. Sci. U. S. A. 99:13284, 2002.

Wilson, S.M., Bhattacharyya, B., Rachel, R.A., Coppolo, V., Tessarollo, L., Householder, D.B., Fletcher, C.F., Miller, R.J., Copeland, N.G., Jenkins, N.A. Synaptic defects in ataxia mice result from a mutation in Usp14, a ubiquitin-specific protease. Nat. Genet. 32:420, 2002.

Wiltshire, T., Pletcher, M.T., Batalov, S., Barnes, S.W., Tarantino, L.M., Cooke, M.P., Wu, H., Smylie, K., Santrosyan, A., Copeland, N.G., Jenkins, N.A., Kalush, F., Mural, R.J., Glynne, R.J., Kay, S.A., Adams, M.D., Fletcher, C.F. Genome-wide single-nucleotide polymorphism analysis defines haplotype patterns in mouse. Proc. Natl. Acad. Sci. U. S. A. 100:3380, 2003.