Affinity purification of microRNA-133a with the cardiac transcription factor, Hand2.

Predictions of microRNA-mRNA interactions typically rely on bioinformatic algorithms, but these algorithms only suggest the possibility of microRNA binding and may miss important interactions as well as falsely predict others. We developed an affinity purification approach to empirically identify microRNAs associated with the 3'UTR of the mRNA encoding Hand2, a transcription factor essential for cardiac development. In addition to miR-1, a known regulator of Hand2 expression, we determined that the Hand2 3'UTR also associated with miR-133a, a microRNA cotranscribed with miR-1 in cardiac and muscle cells. Using a sequential binding assay, we showed that miR-1 and miR-133a could occupy the Hand2 3'UTR concurrently. miR-133a inhibited Hand2 expression in tissue culture models, and miR-133a double knockout mice had elevated levels of Hand2 mRNA and protein. We conclude that Hand2 is regulated by miR-133a in addition to miR-1. The affinity purification assay should be generally applicable for identifying other microRNA-mRNA interactions.

MicroRNA pathways in neural development and plasticity.

MicroRNAs contribute significantly to the development, survival, function, and plasticity of neurons. They silence expression of target genes by reducing mRNA stability and translation. Production of microRNAs is controlled via developmental and environmental cues and these small molecules, in concert with classical transcriptional regulators, amplify changes in neuronal maturation, dendrite morphogenesis, and synaptogenesis. Neurons compartmentalize mRNAs and microRNAs within specific subcellular domains to facilitate control of local protein synthesis in response to neuronal stimuli and to modulate synaptic plasticity. This review addresses issues relevant to microRNA function in neurons, in particular, their ability to reinforce developmental decisions and promote synaptic plasticity.