RNA has emerged as one of the most fascinating molecules in biology. As the Human Genome Project revealed that the number of genes in humans, lower than predicted, is comparable to that from Arabidopsis thaliana and Caenorhabditis elegans, and alternative splicing together with complex combinatorial transcriptional regulation took center stage as fundamental mechanisms to diversify the proteome, significant resources started to focus on the previously so-called “junk DNA”, which encompasses the majority of the human genome and is transcribed into noncoding RNA.
One class of these noncoding RNA molecules, microRNAs, recently emerged as key post-transcriptional regulators in physiological and pathological contexts.
“The work on this topic has been exciting and encouraging, and the field as a whole will be shaped as people understand the relationship between microRNAs and their targets,” says Andrew Z. Fire, Ph.D., professor of pathology and genetics at Stanford University School of Medicine and co-recipient of the 2006 Nobel Prize in Physiology or Medicine.
One of the perceived challenges in studying microRNAs is that each microRNA can apparently regulate up to hundreds of target protein-coding genes, with each target gene potentially regulated by multiple different microRNAs. In this context, the almost 2,000 microRNAs described to date in humans, some of which are present at cellular concentrations that vary by four orders of magnitude, have been viewed as part of an extremely complex and dynamic network.
“The question about the relationship between microRNA molecules and their targets is one that frustrates everyone, as for certain microRNAs it is hard to identify and study their definitive targets, but for others these have been relatively well characterized, in terms of either a broad set of targets that are regulated at modest levels or a few targets that are regulated at substantial levels,” explains Dr. Fire.
Either way, miRNAs are a key family of regulators, and understanding the source of diverse miRNA populations has become a critical part of understanding gene regulation. A recent project in Dr. Fire’s lab tested whether microRNA molecules with no direct genome match could be produced by RNA splicing.
The investigators generated intron-interrupted variants of the Caenorhabditis elegans lin-4 gene, encoding the first microRNA molecule that was discovered and, almost two decades ago, shown to be essential for the temporal control of postembryonic development.
“The intron that we used is a typical intron that a typical mRNA would use,” says Huibin Zhang, a recent Ph.D. graduate in genetics at Stanford University School of Medicine and lead author of the study.
The possibility of processing functional metazoan microRNAs by splicing an intron-interrupted precursor has multiple implications. One of them, the potential requirement for splicing for the in vivo biogenesis of certain microRNAs, would point toward an additional layer in the cellular gene expression regulatory networks.
“This also provides the ability to engineer a microRNA gene and track its expression a little more carefully, as it has to be spliced,” says Dr. Fire. The involvement of splicing in microRNA biogenesis could also help better understand factors that shape splicing in various species.
“The $64,000 question remains the one of target specificity. Once people identify the targets and learn how specific microRNAs interact with them, a lot of things will move forward, and this includes learning why a microRNA could be synthesized with or without introns in terms of the information content involved,” explains Dr. Fire.