miR-132 regulates the differentiation of dopamine neurons by directly targeting Nurr1 expression

• DOI: 10.1242/jcs.086421
• Publication Date: 2012-02-11T04:27:03Z
• Authors: Dehua Yang; Ting Li; Yi Wang; Yuanjia Tang; Huijuan Cui; Yu Tang; Xiaojie Zhang; Degui Chen; Nan Shen; Weidong Le
• Keywords: 0301 basic medicine; Mice; MicroRNAs; 03 medical and health sciences; Dopaminergic Neurons; Gene Expression Profiling; Nuclear Receptor Subfamily 4, Group A, Member 2; Animals; Cell Differentiation; Real-Time Polymerase Chain Reaction; Cells, Cultured; Embryonic Stem Cells

Although it is well established that embryonic stem (ES) cells have the potential to differentiate into dopamine neurons, the molecular basis of this process, particularly the role of microRNAs (miRNAs), remains largely unknown. Here we report that miR-132 plays a key role in the differentiation of dopamine neurons by directly regulating the expression of Nurr1. First, we constructed a mouse ES cell line CGR8, which stably expresses GFP under the tyrosine hydroxylase (TH)-promoter, so the TH-positive neurons could be easily sorted out by a fluorescence-activated cell sorter (FACS). Then, we performed a miRNA array analysis in the purified TH-positive neurons and found that 45 of 585 miRNAs have expression level change >5 fold during dopamine neuron differentiation. Among the 45 miRNAs, we were particularly interested in miR-132 because this miRNA has been reported as being highly expressed in neurons and having a potential role in neurodegenerative diseases. We found that the direct down-regulation of endogenous miR-132 induced by miR-132 antisense oligonucleotide (miR-132-ASO) promoted the differentiation of TH-positive neurons, while ectopic expression of miR-132 in ES cells reduced the number of differentiated TH-positive neurons but did not change the total number of differentiated neurons. Furthermore, we documented that miR-132-ASO could significantly reverse the miR-132-mediated suppression of TH-positive neuron differentiation. Moreover, through bioinformatics assay we identified Nurr1 gene as a potential molecular target of miR-132. Using luciferase-reporter assay and Western blot analysis, we demonstrated that miR-132 could directly regulate the expression of Nurr1. Collectively, our data provide the first evidence that miR-132 is an important molecule regulating ES cell differentiation into dopamine neurons by directly targeting Nurr1 expression.