Seeing is believing: strategies for studying microRNA expression

Introduction Studies during the early 1990s uncovered a novel mechanism by which lin-4 inhibits the nuclear factor encoded by lin-14 to promote the transition between the first and second larval stages of C. elegans development. In particular, lin-4 encodes a small RNA that binds to multiple sites in the 3′ untranslated region (3′-UTR) of the lin-14 transcript, thereby negatively regulating lin-14 at a post-transcriptional level (Lee et al ., 1993; Wightman et al ., 1993). Nearly a decade would pass before it became fully evident that lin-4 was actually the prototype of a novel and extensive class of regulatory RNA, now collectively referred to as the microRNA (miRNA) family (Lagos-Quintana et al ., 2001; Lau et al ., 2001; Lee and Ambros, 2001; Reinhart et al ., 2000). These miRNAs are ∼21–24 nucleotide RNAs that are processed from precursor transcripts containing a characteristic hairpin structure, and have been identified in diverse animals, plants and even viruses (Bartel, 2004; Griffiths-Jones et al ., 2006; Lai, 2003). MiRNAs now constitute one of the largest gene families known, with hundreds to perhaps a thousand or more genes in individual species. Knowledge of temporal and spatial elements of gene expression is essential for a comprehensive understanding of gene function, whether in the context of normal physiology or pathology. With whole genome sequences and extensive databases of expressed sequences in hand, the systematic analysis of mRNA expression patterns using microarrays, in situ hybridization, and even promoter fusions is well underway.