Temporal and spatial regulation of microRNA activity with photo-activatable cantimirs

MicroRNAs (miRNAs) are small noncoding RNAs which play numerous important roles in physiology and human diseases. During animal development, many miRNAs are expressed continuously from early embryos throughout adults, yet it is unclear whether these miRNAs are actually required at all the stages of development. Current techniques of manipulating microRNA function lack the required spatial and temporal resolution to adequately address the functionality of a given microRNA at a specific time or at single-cell resolution. To examine stage- or cell-specific function of miRNA during development and to achieve precise control of miRNA activity, we have developed photo-activatable antisense oligonucleotides against miRNAs. These caged oligonucleotides can be activated with 365 nm light with extraordinarily high efficiency to release potent antisense reagents to inhibit miRNAs. Initial application of these caged antimirs in a model organism ( C. elegans ) revealed that the activity of a miRNA ( lsy-6 ) is required specifically around the comma stage during embryonic development to control a left/right asymmetric differentiation program in the C.elegans nervous system. This suggests that a transient input of lsy-6 during development is sufficient to specify the neuronal cell fate. The modular design and the facile assembly of these caged antisense oligonucleotides should facilitate their applications in detailed functional analyses of miRNAs and their target genes.