Abstract
Germline mutations in Mir96, one of three co-expressed polycistronic miRNA genes (Mir96, Mir182, Mir183), cause hereditary hearing loss in humans and mice. Transgenic FVB/NCrl- Tg(GFAP-Mir183,Mir96,Mir182)MDW1 mice (Tg1MDW), which overexpress this neurosensory-specific miRNA cluster in the inner ear, were developed as a model system to identify, in the aggregate, target genes and biologic processes regulated by the miR-183 cluster. Histological assessments demonstrate Tg1MDW/1MDW homozygotes have a modest increase in cochlear inner hair cells (IHCs). Affymetrix mRNA microarray data analysis revealed that downregulated genes in P5 Tg1MDW/1MDW cochlea are statistically enriched for evolutionarily conserved predicted miR-96, miR-182 or miR-183 target sites. ABR and DPOAE tests from 18 days to 3 months of age revealed that Tg1MDW/1MDW homozygotes develop progressive neurosensory hearing loss that correlates with histologic assessments showing massive losses of both IHCs and outer hair cells (OHCs). This mammalian miRNA misexpression model demonstrates a potency and specificity of cochlear homeostasis for one of the dozens of endogenously co-expressed, evolutionally conserved, small non-protein coding miRNA families. It should be a valuable tool to predict and elucidate miRNA-regulated genes and integrated functional gene expression networks that significantly influence neurosensory cell differentiation, maturation and homeostasis.
Highlights
MicroRNAs are short (~20–24nt) endogenous non-coding RNAs that posttranscriptionally repress the expression of protein-coding genes
Of 425 implanted embryos, 3 of the 16 live born pups were identified as founders and three independent inbred lines were established in FVB/NClr: Tg(GFAP-Mir183,Mir96,Mir182)1MDW, Tg(GFAP-Mir183,Mir96,Mir182)2MDW, Tg(GFAP-Mir183,Mir96,Mir182)3MDW (Tg1MDW, Tg2MDW, Tg3MDW)
The mutual exclusion hypothesis of miRNA function[16] predicts that in Tg1MDW/1MDW, miR-183 cluster misexpression, driven by the glial fibrillary acidic protein (GFAP) promoter in adjacent and lineage-related SCs should repress evolutionarily conserved target genes that are simultaneously essential to SC function and incompatible for attaining and/or maintaining the hair cells (HCs) fate
Summary
MicroRNAs are short (~20–24nt) endogenous non-coding RNAs that posttranscriptionally repress the expression of protein-coding genes. The spatiotemporal expression pattern of Mir[96], Mir[182] and Mir[183] in the developing vertebrate inner ear and the effects induced by modulating levels of these miRNAs on HC fate determination in zebrafish and chicken argue that these miRNAs collectively function, to some degree, in the transition from inner ear prosensory cells towards a HC fate[8,9,11,12]. Genome-wide studies demonstrate that miRNAs can fine-tune mRNA levels in the context of miRNA/target mRNA co-expression and can act as developmental on-off switches, as in the context of mutually exclusive miRNA/target mRNA expression[15,16,17,18,19] For those effects attributable to the miR-183 cluster in the vertebrate inner ear, these functional distinctions in miRNA regulation might be segregated temporally: in a switch-like manner www.nature.com/scientificreports/. RasGEF domain family, member 1 A periaxin a disintegrin-like and metallopeptidase (reprolysin type) with thrombospondin type 1 motif, 20 223838
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