Correction: A Positive Feedback Mechanism That Regulates Expression of miR-9 during Neurogenesis

MiR-9, a neuron-specific miRNA, is an important regulator of neurogenesis. In this study we identify how miR-9 is regulated during early differentiation from a neural stem-like cell. We utilized two immortalized rat precursor clones, one committed to neurogenesis (L2.2) and another capable of producing both neurons and non-neuronal cells (L2.3), to reproducibly study early neurogenesis. Exogenous miR-9 is capable of increasing neurogenesis from L2.3 cells. Only one of three genomic loci capable of encoding miR-9 was regulated during neurogenesis and the promoter region of this locus contains sufficient functional elements to drive expression of a luciferase reporter in a developmentally regulated pattern. Furthermore, among a large number of potential regulatory sites encoded in this sequence, Mef2 stood out because of its known pro-neuronal role. Of four Mef2 paralogs, we found only Mef2C mRNA was regulated during neurogenesis. Removal of predicted Mef2 binding sites or knockdown of Mef2C expression reduced miR-9-2 promoter activity. Finally, the mRNA encoding the Mef2C binding partner HDAC4 was shown to be targeted by miR-9. Since HDAC4 protein could be co-immunoprecipitated with Mef2C protein or with genomic Mef2 binding sequences, we conclude that miR-9 regulation is mediated, at least in part, by Mef2C binding but that expressed miR-9 has the capacity to reduce inhibitory HDAC4, stabilizing its own expression in a positive feedback mechanism. Citation: Davila JL, Goff LA, Ricupero CL, Camarillo C, Oni EN, et al. (2014) A Positive Feedback Mechanism That Regulates Expression of miR-9 during Neurogenesis. PLoS ONE 9(4): e94348. doi:10.1371/journal.pone.0094348 Editor: Wenhui Hu, Temple University School of Medicine, United States of America Received December 23, 2013; Accepted March 13, 2014; Published April 8, 2014 Copyright: 2014 Davila et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. Funding: This work was supported by grants from National Institutes of Health (NIH) (R21 MH085088) and the New Jersey Commission for Science and Technology. JD and CC were supported by fellowships from the New Jersey Commission on Spinal Cord Research; CLR was supported by a National Science Foundation (NSF) IGERT training grant; EO was supported by an NIH Pipeline Grant (5R25GM055145). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. Competing Interests: The authors have declared that no competing interests exist. * E-mail: [email protected] . These authors contributed equally to this work. ¤a Current address: Institute for Stem Cell Biology and Regenerative Medicine, Stanford University School of Medicine, Stanford, California, United States of America ¤b Current address: Department of Stem Cells and Regenerative Medicine, Harvard University, Cambridge, Massachusetts, United States of America ¤c Current address: Center for Craniofacial Regeneration, Columbia University Medical Center, New York, New York, United States of America ¤d Current address: Center of Excellence in Neurosciences, Texas Tech University Health Sciences Center, El Paso, Texas, United States of America