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Thanh Hoang, Lin Liu, Kai Wang, Chun Liang, Michael L Robinson; Transcriptome analysis of small RNA expression in newborn mouse lenses. Invest. Ophthalmol. Vis. Sci. 2015;56(7 ):2641.
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© ARVO (1962-2015); The Authors (2016-present)
Lens fiber cell differentiation from epithelial cells involves coordinated changes in gene expression. Despite the wealth of knowledge of transcription factors involved in lens development, very little is known about the epigenetic regulation of lens fiber cell differentiation. Emerging evidence implicate a role for miRNAs in lens development and disease. The aim of this study is to provide a comprehensive comparison of gene expression profiles of small RNAs between lens epithelial and lens fiber cells using RNA-sequencing
Newborn FVB/N strain mouse lenses were dissected into capsules containing adhering epithelial cells and fiber cells. Small RNA, including miRNA and piRNA, was isolated from total RNA by size-selection and prepared for Illumina HiSeq sequencing to yield 5 million reads per sample. Small RNA sequencing reads that passed quality filters were mapped against miRNA reference sequences for detection of all known miRNA. Unmapped reads were subsequently mapped against piRNA database for detection of piRNA. Differential expression of miRNA and piRNA between the two cell types was then analyzed using DESeq. qRT-PCR validation was conducted to confirm small RNA-Seq data
The lens expresses over 350 known miRNAs. Among these, 145 miRNAs were differentially expressed between epithelial and fiber cells (fold change >2, adjusted p value <0.05). The most abundant miRNAs in the lens include: miR-1a, miR-184, miR-26a, miR-204, miR-211 and members of the let-7 family. qPCR assay confirmed that miR-1a was highly expressed in fiber cells, but only weakly expressed in lens epithelial cells and retina. We also detected the expression of over 250 known piRNAs in the lens. This result is interesting because little information exists about the function of piRNA outside the germline
Small RNA-Seq enables a comprehensive view of both the relative abundance and differential expression of small RNA from lens epithelial cells and lens fiber cells, and provides a valuable resource for studying lens development and associated diseases. Current work focuses on investigating the role of miR-1a, miR-184 and miR-26a in the developing lens
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