Abstract
Abstract: :
Purpose: The Atonal-related bHLH transcription factor Math5 and the POU domain transcription factor Brn-3b are required for the development of retinal ganglion cells (RGCs). Gene disruption of Brn-3b and Math5 resulted in the loss of retinal ganglion cells in mouse retinas. Cell lineage analysis has shown that Brn-3b was not required for the initial commitment of RGC cell fate or for the migration of ganglion cells to the RGC layer. However, Brn-3b was essential for the normal differentiation of retinal ganglion cells; without it, the nascent RGCs failed to project axons properly and underwent enhanced apoptosis. These results suggest that Brn-3b regulate the activity of genes whose products play essential roles in the formation of RGC axons. Unlike that of Brn-3b, the loss of Math5 blocked the initial differentiation of RGCs and math5-null mutation led to the loss of brn-3b expression, suggesting that math5 is essential for RGC differentiation and that math 5 acts upstream to activate brn-3b-dependent differentiation processes in RGCs. The purpose for the present study is to further identify the downstream target genes of math5 and brn-3b regulatory pathway and to establish embryonic retina cDNA microarrays and gene expression profiles as a new approach for gene discovery. Method: Total RNAs from wild type and mutant retinas were used as probes to identify the downstream target genes in the cDNA microarrays experiments. Result: We have generated a mouse E14.5 retinal cDNA expression database and cDNA microarrays. To date, we have archived 15,268 ESTs and have annotated 9,035, which represent 5,288 genes. The fraction of singly occurring ESTs as a function of total EST accrual suggests that the total number of expressed genes in mouse E14.5 retinas could approach 27,000. The annotated 9,035 ESTs were categorized by their known or putative functions. Representation of genes involved in eye development was significantly higher in the retinal clone set compared to the NIA mouse 15,000 cDNA clone set. Screening of the cDNA microarrays using wild type and brn-3b-null retinal cDNA probes revealed a regulatory linkage between the transcription factor Brn-3b and expression of GAP-43, a protein associated with axon growth. Conclusion: The E14.5 retinal cDNA database and the cDNA microarrays will be a valuable platform for gene expression profiling and gene discovery.
Keywords: 564 retinal development • 415 ganglion cells • 316 animal model