Characterization of an Abundant COL9A1 Transcript in the Cochlea with a Novel 3′ UTR: Expression Studies and Detection of miRNA Target Sequence

Abstract

EST N66408 represents one of several large unique clusters expressed in the Morton human fetal cochlear cDNA library. N66408 is 575 bp in size and initial BLAST analysis of this sequence showed no homology to any known genes or expressed sequence tags (ESTs) from other organs or tissues. Sequence of the original cochlear clone from which N66408 was derived revealed that the corresponding cDNA was about 700 bp in size, including 125 bp at its 5 0 end with homology to the 3 0 end of COL9A1 in addition to 575 bp of novel sequence. RT-PCR analysis using primers specific to COL9A1 isoforms 1 and 2 detected expression of both isoforms in human fetal cochlea. Tissue in situ hybridization using the novel 3 0 UTR sequence as probe showed abundant expression in spiral limbus and spiral ligament, and a moderate level of expression in the organ of Corti. dbEST analysis of ESTs specific to the 3 0 UTR of COL9A1 showed 19 ESTs derived from various tissues; three polyadenylation sites were identified and the majority of these ESTs were derived from overlapping polyadenylation signals at the second site (position 749-758). Comparison of the 3 0 UTR of human COL9A1 with its orthologs as well as with dbEST uncovered a highly conserved region around the overlapping polyadenylation signals at position 749-758 in mammals. A search of the microRNA database revealed a highly conserved target sequence for miR-9 immediately preceding the overlapping polyadenylation signals in the novel 3 0 UTR of COL9A1, suggesting its role in posttranscriptional regulation of COL9A1.

Little but loud: Small RNAs have a resounding affect on ear development

“…Although miRNA function in development and maintenance of the inner ear is just beginning to be explored, studies already suggest that their genetic regulatory properties are essential to the proper establishment of sensory neurons and sensory epithelial supporting and hair cells. Moreover, a study of miRNA expression during hair cell regeneration in newt sensory epithelia suggest a role for let-7 miRNAs in the process (Tsonis et al, 2007), and miR-9 function has been suggested in the regulation of a COL9A1 expression in human cochlea (Sivakumaran et al, 2006). Many questions remain regarding the functional contributions and mechanistic actions of individual miRNAs amidst the complexity of genetic and biochemical interactions that contribute to development, differentiation, and maintenance of specific cell types throughout the inner ear.…”

The Role of MicroRNAs in Environmental Risk Factors, Noise-Induced Hearing Loss, and Mental Stress

“…Since 2006 when miRNAs appeared in the mammalian inner ear field, they have exploded as an additional layer of gene regulation in both inner ear development and disease (172). miRNAs show particular expression patterns in the inner ear.…”

A dynamic interplay between alternative polyadenylation and microRNA regulation: Implications for cancer

“…Splicing independent alternative polyadenylation produces transcript isoforms with 3' UTRs of different lengths, containing a constitutive segment and an alternative segment. If a microRNA target is located within the alternative part of the 3' UTR, the shorter transcript should be target free and should escape microRNA-mediated inhibition, while the longer transcript should be inhibited (47)(48)(49). Such variations are very common.…”