June 2017
Volume 58, Issue 8
Open Access
ARVO Annual Meeting Abstract  |   June 2017
Investigation of mRNA and protein targets of the RNA-binding protein Celf1 in mouse lens development
Author Affiliations & Notes
  • Sandeep Aryal
    Department of Biological Sciences, University of Delaware, Newark, Delaware, United States
  • Archana D Siddam
    Department of Biological Sciences, University of Delaware, Newark, Delaware, United States
  • Carole Gautier-Courteille
    Institut de Genetique et Development de Rennes, Universite de Rennes, Rennes, France
  • Luc Paillard
    Institut de Genetique et Development de Rennes, Universite de Rennes, Rennes, France
  • Salil Lachke
    Department of Biological Sciences, University of Delaware, Newark, Delaware, United States
    Center for Bioinformatics and Computational Biology, University of Delaware, Newark, Delaware, United States
  • Footnotes
    Commercial Relationships   Sandeep Aryal, None; Archana Siddam, None; Carole Gautier-Courteille, None; Luc Paillard, None; Salil Lachke, None
  • Footnotes
    Support  NIH/NEI R01 EY021505
Investigative Ophthalmology & Visual Science June 2017, Vol.58, 1720. doi:
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      Sandeep Aryal, Archana D Siddam, Carole Gautier-Courteille, Luc Paillard, Salil Lachke; Investigation of mRNA and protein targets of the RNA-binding protein Celf1 in mouse lens development. Invest. Ophthalmol. Vis. Sci. 2017;58(8):1720.

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

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Abstract

Purpose : Deletion of a novel RNA-binding protein Celf1 causes lens defects and cataracts in mouse and fish. In non-ocular tissues, Celf1 is known to regulate gene expression by distinct post-transcriptional control mechanisms. Yet, the nature of the factors resulting from Celf1 deficiency that contribute to lens pathology remains unclear. To gain insights into Celf1 function in the lens, we have identified Celf1-direct binding transcripts. These data, combined with detailed phenotypic and molecular characterization of Celf1 mouse mutant lenses, allow the development of a comprehensive model wherein Celf1 mediates post-transcriptional control of several critical regulators of lens fiber differentiation.

Methods : Celf1 conditional knockout (Celf1cKO) mice were generated using the Pax6GFPCre mouse line. RNA-sequencing was performed using Illumina HiSeq platform to identify differentially expressed transcripts in Celf1cKO lenses. RNA immunoprecipitation (RIP) and cross-linking immunoprecipitation followed by RT-qPCR were performed on wild-type mouse lenses to identify Celf1-associated transcripts.

Results : RIP analysis identifies several Celf1-binding transcripts that encode factors important for lens biology, including Prox1 and Actn2. Interestingly, while Prox1 transcript levels remained unchanged, its protein levels were found to be up-regulated in the anterior epithelium of Celf1cKO lenses. In contrast, Actn2 was found to be down-regulated on both transcripts and protein levels in Celf1cKO lenses. Insights into the phenotypic consequence of mis-regulation of these factors were obtained from the observations that the anterior epithelium exhibited defective expression of several epithelial-marker genes, while fiber cells exhibited abnormalities in the F-actin staining pattern in Celf1-cKO lenses.

Conclusions : We report new Celf1-target transcripts in the lens, namely those encoding the transcription factor Prox1 and the actin-binding protein Actn2. Extending our previous findings of Celf1-based regulation of the cyclin-dependent kinase inhibitor p27 (Cdkn1b) and the nuclease Dnase2b, the present data indicate that Celf1 is involved in the negative regulation of Prox1 expression in the lens epithelium and is required for Actn2 expression in lens fiber cells. Together, these data serve to further define the mechanisms of Celf1-mediated control of the transcriptome and proteome in the developing lens.

This is an abstract that was submitted for the 2017 ARVO Annual Meeting, held in Baltimore, MD, May 7-11, 2017.

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