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J. Y. Zhao, F. Zhuang, K. Kawai, J. Zhang, Y.-H. Liu; Msx2 Regulates Lens Development by Controlling Lens Cell Cycle. Invest. Ophthalmol. Vis. Sci. 2008;49(13):5408.
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Microphthalmia/anophthalmia is a relatively common genetic malformation that causes vision impairment and blindness with a birth prevalence ranging from 1 to 2 per 10,000. Any disruption to the early inductive phase or subsequent proliferation and differentiation phase of eye development will lead to anophthalmia/ microphthalmia. Msx2 presents itself as a new addition to a growing list of genes that function to facilitate ocular growth. As a homeodomain protein, Msx2 is considered as a transcriptional repressor until our recent discovery of its transcriptional activator activity. The purpose of the current investigation is uncovering the mechanism of Msx2 gene regulated lens development.
To demonstrate its role in controlling lens morphogenesis, we performed molecular and histological analysis of lens development in Msx2 transgenic and knockout animals. Cell transfection experiments were performed to identify functional domains in cell cycle regulation.
Inactivation of Msx2 resulted in the delayed separation of lens from the cornea. Lens epithelial cell proliferation persists without exiting cell cycle to allow differentiation to occur, as such primary lens fiber cells failed to elongate. Surprisingly, alpha Crystallin expression is significantly elevated in the Msx2 null mutant lens. To demonstrate the primary role of Msx2 in controlling lens cell cycle, αTN4 lens epithelial cells were transfected with a Msx2-GFP expression plasmid. Transfected cells were then labeled with BrdU to ascertain the status of cell proliferation. Msx2-GFP positive cells failed to incorporate BrdU. To confirm that cells that overexpressed Msx2-GFP were selectively prevented from entering mitosis, anti-phospho-histone 3 (p-H3) immunofluorescence failed to detect cell nuclei that were GFP positive. The cell cycle inhibitory function is mediated by the C-terminal domain of Msx2 protein because removal of the Msx2 C-terminal domain abrogated the ability of Msx2 to inhibit cell cycle.
Msx2 regulates lens development by controlling initial cell cycle exiting events to generate primary lens fiber cells. Elevated Msx2 expression in the lens vesicle may prevent lens epithelial cells from re-entering cell cycle thus allowing the activation of the differentiation program. Msx2 achieves its cell cycle inhibitory activity via its C-terminal domain.
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