This proposal s long term goal is to determine the molecular mechanisms governing early mammalian eye formation. A key gene involved in oculogenesis is Pax6: human and mouse Pax6 mutations result in aniridia and Small eye, while mutations in a Drosophila Pax6 homolog result in the eyeless phenotype. Strikingly, Pax6 misexpression in Drosophila produces ectopic eyes, implying that vertebrate and invertebrate eye formation utilize similar, evolutionarily conserved Pax6-dependent genetic hierarchies. During the last grant period, we defined the Pax6 gene product as a transcriptional activator, elucidated the DNA binding properties of its paired domain and identified the Msx and Eya gene families as important downstream components of the eye forming regulatory hierarchy. We also identified two murine Pax6 promoters and several Pax6 regulatory elements which direct expression in different parts of the developing eye. In addition, we have found that Drosophila eyeless and mouse Pax6 regulatory elements appear to function to some extent interchangeably in transgenic mice and flies, suggesting that the upstream regulation of Pax6 is also evolutionarily conserved. In this renewal application, we seek to further elucidate the Pax6- dependent genetic pathway(s) controlling early mammalian eye development. In the first specific aim, we will go upstream by further analyzing regulatory elements controlling Pax6 ocular expression and by identifying the transcription factors that interact with them. In the second specific aim, we will focus on Pax6 downstream targets and clarify the relationship between them in the developing lens placode. In the third aim, we will determine if Msx and Eya are directly regulated by Pax6 and clarify the details of a strong protein-protein interaction we have detected between the highly conserved Eya domain and the Msx and Six homeoproteins. Finally, in the fourth aim, we will use try to complement Pax6 function in prospective lens ectoderm of Pax6-deficient Sey/Sey mice with various Pax6 transgenes and downstream target genes, thus reconstituting early steps of lens induction. This latter goal, although ambitious, has already been accomplished by others in a limited, empiric fashion in insects, fish and amphibians. The proposed work will identify factors controlling Pax6 ocular expression and downstream Pax6 regulatory targets, clarify key details of the emerging Pax6- hierarchy at the molecular level and test this regulatory hierarchy in vivo.

Agency
National Institute of Health (NIH)
Institute
National Eye Institute (NEI)
Type
Research Project (R01)
Project #
5R01EY010123-09
Application #
6518501
Study Section
Visual Sciences A Study Section (VISA)
Program Officer
Liberman, Ellen S
Project Start
1993-07-01
Project End
2004-04-30
Budget Start
2002-05-01
Budget End
2003-04-30
Support Year
9
Fiscal Year
2002
Total Cost
$477,245
Indirect Cost
Name
Brigham and Women's Hospital
Department
Type
DUNS #
071723621
City
Boston
State
MA
Country
United States
Zip Code
02115
Lin, Haotian; Ouyang, Hong; Zhu, Jie et al. (2016) Lens regeneration using endogenous stem cells with gain of visual function. Nature 531:323-8
Zhang, Ying; Fan, Jieqing; Ho, Joshua W K et al. (2016) Crim1 regulates integrin signaling in murine lens development. Development 143:356-66
Li, Gen; Xu, Fan; Zhu, Jie et al. (2015) Transcription Factor PAX6 (Paired Box 6) Controls Limbal Stem Cell Lineage in Development and Disease. J Biol Chem 290:20448-54
Anchan, Raymond M; Lachke, Salil A; Gerami-Naini, Behzad et al. (2014) Pax6- and Six3-mediated induction of lens cell fate in mouse and human ES cells. PLoS One 9:e115106
Lachke, Salil A; Higgins, Anne W; Inagaki, Maiko et al. (2012) The cell adhesion gene PVRL3 is associated with congenital ocular defects. Hum Genet 131:235-50
Lachke, Salil A; Ho, Joshua W K; Kryukov, Gregory V et al. (2012) iSyTE: integrated Systems Tool for Eye gene discovery. Invest Ophthalmol Vis Sci 53:1617-27
Lachke, Salil A; Maas, Richard L (2011) RNA Granules and Cataract. Expert Rev Ophthalmol 6:497-500
Lachke, Salil A; Alkuraya, Fowzan S; Kneeland, Stephen C et al. (2011) Mutations in the RNA granule component TDRD7 cause cataract and glaucoma. Science 331:1571-6
Lachke, Salil A; Maas, Richard L (2010) Building the developmental oculome: systems biology in vertebrate eye development and disease. Wiley Interdiscip Rev Syst Biol Med 2:305-323
Rowan, Sheldon; Siggers, Trevor; Lachke, Salil A et al. (2010) Precise temporal control of the eye regulatory gene Pax6 via enhancer-binding site affinity. Genes Dev 24:980-5

Showing the most recent 10 out of 25 publications