| We have isolated a zinc finger transcription factor Ru49 from a murine cerebellar cDNA library. Ru49 has a novel protein-protein interaction domain, the LeR domain, which is capable of both homo-dimerization and hetero-dimerization. We also showed that zinc fingers of Ru49 can bind to a novel bipartite DNA binding sequence in vitro. By in situ hybridizations, Ru49 was shown to be expressed in the granule cell lineages of the cerebellum, the dentate gyrus and the olfactory bulb. To further study the expression and function of Ru49 in vivo, I took a transgenic approach to identify an Ru49 genomic fragment that can direct proper expression of a marker gene in vivo. Since small genomic constructs failed to do so, I developed a novel method to modify Bacterial Artificial Chromosomes (BACs) and used modified BACs to generate transgenic mice. Four independent transgenic lines were generated carrying the 131kb Ru49 BAC containing either a lacZ or an EGFP1 marker gene. All four lines exhibited integration site independent and proper expression of the marker genes in vivo. In addition, they revealed novel quantitative aspects of Ru49 expression pattern in cerebellar granule cells. To study the function of Ru49 in cerebellar development, I performed phenotypic analyses on BAC.EGFP1 transgenic mouse lines which also overexpressed Ru49 in the endogenous expression pattern in vivo. In two independent lines, such overexpression resulted in an increase in cerebellar granule cell production, which is not due to a decrease in apoptosis, but likely due to an increase in cell proliferation. Furthermore, Ru49 overexperssion also resulted in an increase in formation of the intraculminate fissure (ICF) in the cerebellum. In conclusion, Ru49 is a cerebellar granule cell lineage specific transcription factor that plays a critical role in controlling cerebellar granule cell production and cerebellar morphogenesis. |
