A novel mutation within the MIR96 gene causes non-syndromic inherited hearing loss in an Italian family by altering pre-miRNA processing.
The miR-96, miR-182 and miR-183 microRNA (miRNA) family is essential for differentiation and function of the vertebrate inner ear. Recently, point mutations within the seed region of miR-96 were reported in two Spanish families with autosomal dominant non-syndromic sensorineural hearing loss (NSHL) and in a mouse model of NSHL. We screened 882 NSHL patients and 836 normal-hearing Italian controls and identified one putative novel mutation within the miR-96 gene in a family with autosomal dominant NSHL. Although located outside the mature miR-96 sequence, the detected variant replaces a highly conserved nucleotide within the companion miR-96*, and is predicted to reduce the stability of the pre-miRNA hairpin. To evaluate the effect of the detected mutation on miR-96/mir-96* biogenesis, we investigated the maturation of miR-96 by transient expression in mammalian cells, followed by real-time reverse-transcription polymerase chain reaction (PCR). We found that both miR-96 and miR-96* levels were significantly reduced in the mutant, whereas the precursor levels were unaffected. Moreover, miR-96 and miR-96* expression levels could be restored by a compensatory mutation that reconstitutes the secondary structure of the pre-miR-96 hairpin, demonstrating that the mutation hinders precursor processing, probably interfering with Dicer cleavage. Finally, even though the mature miR-96 sequence is not altered, we demonstrated that the identified mutation significantly impacts on miR-96 regulation of selected targets. In conclusion, we provide further evidence of the involvement of miR-96 mutations in human deafness and demonstrate that a quantitative defect of this miRNA may contribute to NSHL.
Dipartimento di Biologia e Genetica per Scienze Mediche, Università degli Studi di Milano, Milan, Italy. giulia.solda@unimi.it
Giulia
G
Soldà
Michela
M
Robusto
Paola
P
Primignani
Pierangela
P
Castorina
Elena
E
Benzoni
Antonio
A
Cesarani
Umberto
U
Ambrosetti
Rosanna
R
Asselta
Stefano
S
Duga
D005786:Gene Expression Regulation; D006367:HeLa Cells; D006319:Hearing Loss, Sensorineural / Q000235:genetics*; D006801:Humans; D007558:Italy; D035683:MicroRNAs / Q000235:genetics* / Q000378:metabolism; D009154:Mutation; D009690:Nucleic Acid Conformation; D012322:RNA Precursors / Q000737:chemistry / Q000378:metabolism; D012323:RNA Processing, Post-Transcriptional
C541492:MIRN96 microRNA, human; D035683:MicroRNAs; D012322:RNA Precursors