April 2011
Volume 52, Issue 14
Free
ARVO Annual Meeting Abstract  |   April 2011
Heterochromatin Is Retained During Lens Fiber Cell Differentiation
Author Affiliations & Notes
  • Melinda K. Duncan
    Biological Sciences, University of Delaware, Newark, Delaware
  • Robert P. Sheehan
    Biological Sciences, University of Delaware, Newark, Delaware
Investigative Ophthalmology & Visual Science April 2011, Vol.52, 2779. doi:
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      Melinda K. Duncan, Robert P. Sheehan; Heterochromatin Is Retained During Lens Fiber Cell Differentiation. Invest. Ophthalmol. Vis. Sci. 2011;52(14):2779.

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      © ARVO (1962-2015); The Authors (2016-present)

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Abstract

Purpose: : The nuclei of lens fiber cells are lost as these cells undergo terminal differentiation. While the vast majority of the nuclear DNA disappears from these cells, confocal imaging of lens sections stained with highly specific and sensitive fluorescent DNA dyes revealed a punctate pattern of small particles in terminally differentiated fiber cells. This work investigates the distribution and molecular identity of these particles.

Methods: : Mouse, rat, human and cow lenses were sectioned and stained with both the specific fluorescent DNA dye Draq5 and antibodies specific for histone modifications associated with either heterochromatin or euchromatin. The cellular distribution and size of these particles was assessed by confocal histomorphometry.

Results: : The punctate spots detected in terminally differentiated lens fibers by upon staining with Draq5 also intensely stain with antibodies against histone H3, trimethylated on Lysine 9, and 5-Methylcytosine, which are both molecular markers of compact heterochromatin. In contrast, only intact or partially degraded fiber nuclei stain with antibodies against histone H3 Lysine 9 acetylation or histone H4 lysine 8 acetylation, which are predominant in the more accessible euchromatin. In mouse lenses, these presumptive particles of heterochromatin have a mean cross-sectional area of 1.1 microns squared but have a very wide size distribution down from that of an intact nucleus. Further, they are distributed broadly within lens fiber cells suggesting that they are able to diffuse away from the site of their parent nuclei.

Conclusions: : These data suggest that the DNA fragments persisting in mature lens fiber cells are predominately composed of heterochromatin, likely because their highly compacted state prevented access of the nucleases responsible for DNA degradation during lens fiber cell differentiation. We are currently testing the hypothesis that these fragments preferentially contain genes that are transcriptionally silent in the lens.

Keywords: transcription • differentiation • immunohistochemistry 
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