Gunaratne et al. BMC Genomics 2011, 12:277 http://www.biomedcentral.com/1471-2164/12/277 R E S E A R C H A R T I C L E Open Access Song exposure regulates known and novel microRNAs in the zebra finch auditory forebrain Preethi H Gunaratne1,2,3†, Ya-Chi Lin4†, Ashley L Benham1, Jenny Drnevich5, Cristian Coarfa11, Jayantha B Tennakoon1, Chad J Creighton6, Jong H Kim1, Aleksandar Milosavljevic11, Michael Watson7, Sam Griffiths-Jones8 and David F Clayton4,9,10* Abstract Background: In an important model for neuroscience, songbirds learn to discriminate songs they hear during tape-recorded playbacks, as demonstrated by song-specific habituation of both behavioral and neurogenomic responses in the auditory forebrain. We hypothesized that microRNAs (miRNAs or miRs) may participate in the changing pattern of gene expression induced by song exposure. To test this, we used massively parallel Illumina sequencing to analyse small RNAs from auditory forebrain of adult zebra finches exposed to tape-recorded birdsong or silence. Results: In the auditory forebrain, we identified 121 known miRNAs conserved in other vertebrates. We also identified 34 novel miRNAs that do not align to human or chicken genomes. Five conserved miRNAs showed significant and consistent changes in copy number after song exposure across three biological replications of the song-silence comparison, with two increasing (tgu-miR-25, tgu-miR-192) and three decreasing (tgu-miR-92, tgu- miR-124, tgu-miR-129-5p). We also detected a locus on the Z sex chromosome that produces three different novel miRNAs, with supporting evidence from Northern blot and TaqMan qPCR assays for differential expression in males and females and in response to song playbacks. One of these, tgu-miR-2954-3p, is predicted (by TargetScan) to regulate eight song-responsive mRNAs that all have functions in cellular proliferation and neuronal differentiation. Conclusions: The experience of hearing another bird singing alters the profile of miRNAs in the auditory forebrain of zebra finches. The response involves both known conserved miRNAs and novel miRNAs described so far only in the zebra finch, including a novel sex-linked, song-responsive miRNA. These results indicate that miRNAs are likely to contribute to the unique behavioural biology of learned song communication in songbirds. Background Songbirds are important models for exploring the neural and genomic mechanisms underlying vocal communica- tion, social experience and learning (reviewed in [1]). Songbirds communicate using both innate calls and learned vocalizations (songs), and unique specializations of the brain evolved to support this behavior (reviewed in [2]). In the zebra finch, only the male produces songs, although both sexes process and discriminate specific songs [3-6]. The genome is actively engaged by song communication, as first shown in an early * Correspondence: dclayton@uiuc.edu † Contributed equally 4Department of Cell and Developmental Biology, University of Illinois, Urbana-Champaign, IL 61801, USA Full list of author information is available at the end of the article demonstration of how gene responses in the brain dis- criminate among different song stimuli [7]. The genomic response is not a simple correlate of neural activity and it can vary significantly according to the salience and behavioral context of the experience [8-13]. Recent stu- dies using microarray technology have now shown that song exposure affects the expression of thousands of genes in the auditory forebrain [14,15]. Repeated expo- sure to one song leads to an altered gene expression profile, correlated with habituation of both the beha- vioral and immediate genomic responses to that specific song. These observations suggest the involvement of large and dynamic transcriptional network in the recog- nition and memory of complex vocal signals [14]. MicroRNAs (miRNAs or miRs) are emerging as potential control points in transcriptional networks, and © 2011 Gunaratne et al; licensee BioMed Central Ltd. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.