Cell Biology, Neurobiology & Anatomy

Brian A. Link, PhD
Professor

Department of Cell Biology, Neurobiology & Anatomy
Medical College of Wisconsin
8701 Watertown Plank Road
Milwaukee, WI 53226-0509

Phone: (414) 955-8072
FAX: (414) 955-6517
email: blink@mcw.edu

Mechanisms of vertebrate retinogenesis

There are several interrelated and ongoing projects for this topic. Emphasis has been placed on identifying regulators of retinogenesis by analyzing zebrafish mutants that show disrupted retinal lamination (Figure 1).

This phenotype is analogous to the "rough-eye" phenotype of the Drosophila ommatidium which has been so critical in understanding the principles of signaling during development. Characterizations of zebrafish mutants have shed light on fundamental processes of retinogenesis including regulation and selection of cell cycle exit, post-mitotic cell migration, and the mechanisms of coordinated cellular differentiation. Additional emphasis in our lab is now being placed on studying the process of interkinetic nuclear migration and its relationships to neurogenesis. Interkinetic nuclear migration is the cellular behavior where neuroepithelia nuclei migrate in an apical-basal manner in phase with the cell cycle. The process is heterogeneous among the progenitors. Confocal time-lapse imaging with cell type specific GFP-based transgenes revealed that particular patterns of interkinetic nuclear migration influence which progenitor cells exit the cell cycle (Baye and Link, 2007; Del Bene et al, 2008) (Figure 2 and Movie 1).

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Movie 1. In vivo confocal time-lapse movie shows heterogeneity of interkinetic nuclear migration in retinal progenitors from 24 hpf. Images are composed of compressed z-stacks of H2A-GFP fluorescence overlaid with single mid-plane bright-field images. Apical and basal surfaces are outlined with dashed white lines in the first and last frame and an arrow indicates a cell undergoing mitosis at the apical surface. The movie plays at 6 frames per second covering 12 hr of development time (Baye and Link, 2007).

Specifically, neuroepithelia with deep basal nuclear migrations and shorter cell cycles are biased to produce neurons, rather than give rise to daughter cells that re-enter the cell cycle. The lab is currently exploring the signals and mechanisms underlying these relationships.

Complex gene interactions underlying glaucoma-phenotypes

The overall goal of these studies is to understand the relationships between genes that contribute to glaucoma-phenotypes. The glaucomas are a heterogeneous group of ocular disorders characterized by retinal ganglion cell death, optic nerve damage and visual field loss. Elevated intraocular pressure is a principal risk factor for this neurodegenerative disease. The majority and perhaps all forms of glaucoma are complex – requiring the interaction of multiple genes. While several genes that contribute to glaucoma have been identified, many additional loci remain unknown. We are using zebrafish to identify and study genes which promote glaucoma-phenotypes (Figure 3).

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• Clark BS, Cui S, Miesfeld JB, Klezovitch O, Vasioukhin V, Link BA. Loss of Llgl1 in retinal neuroepithelia reveals links between apical domain size, Notch activity and neurogenesis. Development. 2012 139(9):1599-610.

• Clark BS, Winter M, Cohen AR, Link BA. Generation of Rab-based transgenic lines for in vivo studies of endosome biology in zebrafish. Dev Dyn. 2011 240(11):2452-65.

• Willardsen MI and BA Link. (2011). Cell biological regulation of division fate in vertebrate neuroepithelial cells. Developmental Dynamics. Dev Dyn. 2011 240(8):1865-79.

• Collery RF and Link BA. Dynamic Smad-mediated BMP signaling revealed through transgenic zebrafish. Developmental Dynamics. Developmental Dynamics. 240(3):712-22.

• Veth KN, Willer JR, Collery RF, Gray MP, Willer GB, Wagner DS, Mullins MC, Udvadia AJ, Smith RS, John SWM, Gregg RG, Link BA. Mutations in zebrafish lrp2 result in adult onset ocular pathogenesis that models myopia and other risk factors for glaucoma. PLoS Genetics. 7(2):e1001310.

• Insinna C, Baye LM, Amsterdam A, Besharse JC, Link BA. (2010). Analysis of a zebrafish dync1h1 mutant reveals multiple functions for cytoplasmic dynein 1 during retinal photoreceptor development. Neural Development. 5:12.

• McMahon C, Gestri G, Wilson SW, Link BA. (2009). Lmx1b is essential for survival of periocular mesenchymal cells and influence of Fgf-mediated retinal patterning in zebrafish. Developmental Biology. 332(2):287-298.

• Norden C, Young S., Link BA, Harris WA. (2009). Actomyosin is the main driver of interkinetic nuclear migration in the retina. Cell. 138(6):1195-1208.

• Skarie JM and Link BA. (2009). FoxC1 is essential for vascular basement membrane integrity and hyaloid vessel morphogenesis. Invest. Ophthalmol Vision Sci. 50(11): 5026-5034.

• Gray MP*, Smith RS*, Soules KA, John SWM, Link BA. (2009). The aqueous humor outflow pathway of zebrafish. Invest. Ophthalmol Vision Sci. 50(4):1515-1521. *(equal contribution)

• Del Bene F, Wehman AM, Link BA*, Baier H*. (2008). Interkinetic nuclear migration cooperates with an apical-basal Notch signaling gradient to regulate neurogenesis. Cell. 134:1055-1065. *(co-corresponding authors).

• Skarie, JM and Link, BA (2008). The primary open-angle glaucoma gene WDR36 functions in ribosomal-RNA processing and interacts with the p53 stress-response pathway. Human Molecular Genetics, 17:2474-2485.

• Baye, LM and Link, BA (2007). Interkinetic nuclear migration and the selection of neurogenic cell divisions during vertebrate retinogenesis. J Neuroscience 27:10143-52, 2007.

• Cui S, C Otten, S Rohr, S Abdelilah-Seyfried and BA Link (2007). Analysis of aPKClambda and aPKCzeta reveals multiple and redundant functions during vertebrate retinogenesis. Molecular Cellular Neuroscience 34(3):431-444, 2007.