Extremely complex populations of small RNAs in the mouse retina and RPE/choroid.

PURPOSE MicroRNAs (miRNAs) are small noncoding RNAs of approximately 18 to 22 nucleotides in length that regulate gene expression. They are widely expressed in the retina, being both required for its normal development and perturbed in disease. The aim of this study was to apply new high-throughput sequencing techniques to more fully characterize the miRNAs and other small RNAs expressed in the retina and retinal pigment epithelium (RPE)/choroid of the mouse. METHODS Retina and RPE/choroid were dissected from eyes of 3-month-old C57BL/6J mice. Small RNA libraries were prepared and deep sequencing performed on a genome analyzer. Reads were annotated by alignment to miRBase, other noncoding RNA databases, and the mouse genome. RESULTS Annotation of 9 million reads to 320 miRNAs in retina and 340 in RPE/choroid provides the most comprehensive profiling of miRNAs to date. Two novel miRNAs were identified in retina. Members of the sensory organ-specific miR-183, -182, -96 cluster were among the most highly expressed, retina-enriched miRNAs. Remarkably, miRNA "isomiRs," which vary slightly in length and are differentially detected by Taqman RT-qPCR assays, existed for all the microRNAs identified in both tissues. More variation occurred at the 3' ends, including nontemplated additions of T and A. Drosha-independent mirtron miRNAs and other small RNAs derived from snoRNAs were also detected. CONCLUSIONS Deep sequencing has revealed the complexity of small RNA expression in the mouse retina and RPE/choroid. This knowledge will improve the design and interpretation of future functional studies of the role of miRNAs and other small RNAs in retinal disease.