The retina represents an accessible, well-studied area of the central nervous system (CNS). It has served as a model of the CNS and is thus fairly well described through the applications of classical techniques, such as histology, physiology, and biochemistry. We feel that it is also an ideal tissue for application of molecular biological techniques for examination of the molecular mechanisms that govern development and disease. Using retrovirus vectors, we have devised a technique that allows us to introduce exogenous genes into the developing tissue in situ. We can score the effects of these genes by harvesting the retina at virtually any time. In this proposal, we plan to introduce and misexpress genes into the developing retina to look for effects in overall patterning, cell commitment, differentiation and/or morphology, as well as disease. We plan to use genes that have been shown to be important in other systems for these processes, such as homeobox containing genes, receptor-tyrosine kinase genes and growth factor genes. In addition, we will test the hypothesis that neuropeptides and transmitters have a developmental role by misexpressing several neuropeptide genes, and the synthetic enzyme for dopamine. We further plan to investigate the endogenous expression of genes implicated in other systems to have a role in CNS development. The techniques of in situ hybridization and immunohistochemistry will be applied to characterize endogenous expression. Positive results in any of these assays will be followed by the examination of RNA structure using Northern blots and protein structure using Western blots. cDNA libraries prepared from neonatal rat and embryonic chick retinas will be screened for the presence of clones encoding any genes that are identified by the above methods, or from other systems, such as Drosophila. Any cDNA clones will be sequenced and used for further characterization of the gene structure. In addition, open reading frames from such cDNAs will be cloned into our retroviral vector and introduced back into the retina at different times in development for assessment of the gene function.

Agency
National Institute of Health (NIH)
Institute
National Eye Institute (NEI)
Type
Research Project (R01)
Project #
5R01EY008064-02
Application #
3265190
Study Section
Visual Sciences A Study Section (VISA)
Project Start
1989-04-01
Project End
1994-03-31
Budget Start
1990-04-01
Budget End
1991-03-31
Support Year
2
Fiscal Year
1990
Total Cost
Indirect Cost
Name
Harvard University
Department
Type
Schools of Medicine
DUNS #
082359691
City
Boston
State
MA
Country
United States
Zip Code
02115
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Rowan, Sheldon; Cepko, Constance L (2005) A POU factor binding site upstream of the Chx10 homeobox gene is required for Chx10 expression in subsets of retinal progenitor cells and bipolar cells. Dev Biol 281:240-55

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