Development and maintenance of rod and cone photoreceptors require coordinately regulated expression of photoreceptor genes. This regulation occurs at multiple levels and requires the action of a network of transcription regulatory proteins. This regulatory network is centered on the cone-rod homeobox protein CRX and incorporates both photoreceptor-specific transcription factors (TFs) and general chromatin modulators. The ultimate goal of our research is to determine how this regulatory protein network interacts with the chromatin of co-expressed target genes to achieve coordinately regulated spatial and temporal expression, providing a better understanding of photoreceptor development and disease. During the last grant period, we discovered that CRX regulates target gene chromatin configurations by recruiting co-activators capable of catalyzing acetylation of histones and organizing intra-gene interactions between regulatory regions. In the new grant period, we will extend our vision of transcriptional regulation to three dimensions: We hypothesize that rod and cone genes are co-regulated as clusters, requiring appropriate higher-order chromosome organization in the nucleus that is mediated by specific TFs and general epigenetic modulators. To test this hypothesis, Aim 1 will determine the role of histone H3 lysine 4 methylation, a positive mark that occurs downstream of acetylation, by assessing the effects of deleting three catalytic enzymes, MLL1, MLL2 and MLL3 in rods, cones and their precursors;
Aim 2 will reveal specific intra- and inter-chromosome interactions of co-regulated photoreceptor genes and their co-localization in rod vs. cone nuclei using state-of-the-art epigenetic technologies. To determine if these chromosome organizations contribute to transcription co- regulation during subtype specification and maintenance, profiles of chromosome interactions, TF binding, histone marks and transcription will be studied at various developmental ages. We will also determine if the transcription regulatory network mediates the specific chromosome interactions by analyzing mutant retinas lacking specific TFs or epigenetic modulators. This study will provide the first targeted assessment of transcriptional co-regulation of photoreceptor genes and determine the underlying molecular and cellular mechanisms. The new knowledge gained will have implications in developing stem cell-based cell replacement therapy as well as providing new targets for other therapeutic interventions.

Public Health Relevance

The expression of rod and cone genes is coordinately regulated by an intrinsic program established by two types of transcription regulators: photoreceptor-specific transcription factors and general epigenetic modulators. This study investigates the mechanism of action of this program using multi-disciplinary genomics technologies, addressing the importance of each type of transcription regulator in establishing transcription patterns and yielding a three-dimensional view of transcriptional regulation within photoreceptor nuclei. Together, these will provide insights into the intrinsic program governing photoreceptor development and maintenance. We expect these novel approaches to advance our understanding of photoreceptor diseases and lead to new treatment strategies.

Agency
National Institute of Health (NIH)
Institute
National Eye Institute (NEI)
Type
Research Project (R01)
Project #
2R01EY012543-14
Application #
8577320
Study Section
Special Emphasis Panel (BVS)
Program Officer
Neuhold, Lisa
Project Start
2000-03-01
Project End
2018-07-31
Budget Start
2013-08-01
Budget End
2014-07-31
Support Year
14
Fiscal Year
2013
Total Cost
$456,000
Indirect Cost
$156,000
Name
Washington University
Department
Ophthalmology
Type
Schools of Medicine
DUNS #
068552207
City
Saint Louis
State
MO
Country
United States
Zip Code
63130
Brightman, Diana S; Razafsky, David; Potter, Chloe et al. (2016) Nrl-Cre transgenic mouse mediates loxP recombination in developing rod photoreceptors. Genesis 54:129-35
Occelli, Laurence M; Tran, Nicholas M; Narfström, Kristina et al. (2016) CrxRdy Cat: A Large Animal Model for CRX-Associated Leber Congenital Amaurosis. Invest Ophthalmol Vis Sci 57:3780-92
Ruzycki, Philip A; Tran, Nicholas M; Kefalov, Vladimir J et al. (2015) Graded gene expression changes determine phenotype severity in mouse models of CRX-associated retinopathies. Genome Biol 16:171
Tran, Nicholas M; Zhang, Alan; Zhang, Xiaodong et al. (2014) Mechanistically distinct mouse models for CRX-associated retinopathy. PLoS Genet 10:e1004111
Tran, Nicholas M; Chen, Shiming (2014) Mechanisms of blindness: animal models provide insight into distinct CRX-associated retinopathies. Dev Dyn 243:1153-66
Masuda, Tomohiro; Zhang, Xiaodong; Berlinicke, Cindy et al. (2014) The transcription factor GTF2IRD1 regulates the topology and function of photoreceptors by modulating photoreceptor gene expression across the retina. J Neurosci 34:15356-68
Hennig, Anne K; Peng, Guang-Hua; Chen, Shiming (2013) Transcription coactivators p300 and CBP are necessary for photoreceptor-specific chromatin organization and gene expression. PLoS One 8:e69721
Peng, Guang-Hua; Chen, Shiming (2013) Double chromatin immunoprecipitation: analysis of target co-occupancy of retinal transcription factors. Methods Mol Biol 935:311-28
Peng, Guang-Hua; Chen, Shiming (2012) Revealing looping organization of mammalian photoreceptor genes using chromosome conformation capture (3C) assays. Methods Mol Biol 884:305-18
Rao, Rajesh C; Hennig, Anne K; Malik, Muhammad T A et al. (2011) Epigenetic regulation of retinal development and disease. J Ocul Biol Dis Infor 4:121-36

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