The heart is the first organ to form in mammalian embryos. In mice, the heart starts to pump blood shortly after embryonic day 8 (reviewed in Chen and Wang, 2012), and this primitive circulatory system is vital for ensuring the proper development of the embryo. Many different cell types coexist within the heart, such as muscle cells, vascular cells and pacemaker cells (reviewed in Chien et al., 2008). These different types of cell arise from a common pool of progenitor cells, but the details of this process and how these cells are maintained within the mature heart have puzzled cardiovascular scientists. Now, in eLife, Kathryn Ivey, Deepak Srivastava and co-workers at the Gladstone Institute of Cardiovascular Disease—including Amy Heidersbach as first author—have offered some important clues about the underlying mechanisms (Heidersbach et al., 2013).
The development of the heart is regulated at both the transcriptional and post-transcriptional level; regulation at the post-transcriptional level involves small non-coding RNA molecules called microRNAs (miRs). These molecules, which are evolutionarily conserved, regulate gene expression by binding to specific sequences within the messenger RNAs and reducing their stability or preventing them from being translated into proteins. Precise miR activity is required for the heart to develop normally, and for it to be able to respond to challenges such as insufficient blood supply, and pressure overload (the increased stress that develops in the left ventricular wall when the heart pumps blood). Currently known cardiac enriched miRs include miR-1, 133, 206, 208 and 499 (reviewed in Chen and Wang, 2012), with miR-1 being the most abundant in the adult mouse heart.
MiR-1 is co-transcribed with miR-133a, and has two copies in the mouse genome, miR-1-1 on chromosome 2 and miR-1-2 on chromosome 18. Srivastava and co-workers have previously investigated the role of miR-1-2 during the development and maintenance of the heart (Zhao et al., 2007). In the current study, they have also involved its close relative, miR-1-1, in order to investigate the consequences of complete loss of miR-1 in the mammalian heart, and to identify any functions of miR-1 that are dependent on the amount of miR present.
Both miR-1-1 and miR-1-2 give rise to identical mature miR-1 species. Accordingly, targeted deletion of miR-1-1 results in a phenotype similar to that described for miR-1-2-null mice. However, the genetic background of the knock-out mice seems to affect the phenotype, as indicated by results from Heidersbach et al. and also those of another group who observed no phenotypic changes in miR-1-133 knock-out mice (Wystub et al., 2013). Nevertheless, both groups show that mice lacking all miR-1 copies (miR-1 null) die before weaning due to developmental defects. Heidersbach et al. observed abnormalities in mitochondria (the cell’s energy-producing organelles) as well as dysfunctional sarcomeres—the basic functional units of muscle fibres, which consist of thick filaments formed from the protein myosin and thin filaments made up of the protein actin.
Heidersbach et al. identified a gene called myosin light chain kinase (mlck) as being a direct target of miR-1. The mlck gene encodes an enzyme that adds phosphate groups to myosin molecules, and levels of this enzyme were increased in miR-1 null mice. However, this increased level of mlck was accompanied by a reduction, rather than an increase, in the phosphorylation of its substrate, the myosin light chain. This apparent contradiction was resolved by the discovery that miR-1 deletion leads to increased expression of an alternative form of MLCK, known as telokin, which lacks catalytic activity. Telokin is normally found only in smooth muscle—the type of weakly contractile muscle that lines the gut and the blood vessels—and its increased expression prevented MLCK kinase from phosphorylating myosin.
However, the fact that telokin alone was primarily induced—and not other forms of mlck, which all have regulatory sequences that are recognized by miR-1—implies that this might not be the whole story. Likewise, the dramatic increase in mlck expression (roughly fourfold) also suggests involvement of a more complex regulatory mechanism, since miRs generally change the expression level of their target messenger RNA by only 20–50% (reviewed in van Rooij and Olson, 2012).
When Heidersbach et al. analyzed the genes that were up-regulated in the hearts of miR-1 null mice, they found that levels of a protein called smMYOCD were dramatically increased. This protein, known as ‘smooth muscle version of Myocardin’, interacts with a transcription factor called SRF, which regulates the expression of genes that determine the characteristics of cardiac and smooth muscle. It turns out that smMYOCD is a direct target of miR-1. Moreover, the smMYOCD-SRF complex is more effective at promoting transcription of telokin than the cardiac version of Myocardin is. This completes a negative feedback loop that ensures that heart cells have a cardiac rather than smooth muscle phenotype: miR-1 suppresses smMYOCD, promoting the expression of the cardiac enzyme mlck, rather than smooth muscle genes such as telokin, resulting in phosphorylation of myosin and the formation of sarcomeres (Figure 1).
The formation of highly contractile cardiac muscle, as opposed to weakly contractile smooth muscle, depends on the inhibition of a protein called telokin by a microRNA called miR-1. In addition to …
This impressive feat of molecular sleuthing by Heidersbach et al. raises a number of important points that go beyond this specific story. Theoretically, miRs can target hundreds of genes across the genome due to the short target sequences (6–8 nucleotides) they recognize. The working model built by Heidersbach et al. illustrates how networks of genes regulated by miRs can indirectly control the expression of a critical gene in any tissue (Figure 1). Direct miR-mediated repression of a transcriptional activator for a target gene amplifies the influence of that miR on its target. In the example described by Heidersbach et al., the feedback loop comprised of miR-1, SRF and smMYOCD guarantees the sensitive and powerful regulation of telokin in response to environmental and pathological stimuli, and probably makes the heart more robust.
Heidersbach et al. also reveal the importance of computational methods to unwind the complex web woven by miRs in organ development and homeostasis. Future progress in this area will require creative computational approaches to help discover new mechanisms and principles of miR-regulated gene expression.
© 2013, Tao and Martin
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Hair follicle development is initiated by reciprocal molecular interactions between the placode-forming epithelium and the underlying mesenchyme. Cell fate transformation in dermal fibroblasts generates a cell niche for placode induction by activation of signaling pathways WNT, EDA, and FGF in the epithelium. These successive paracrine epithelial signals initiate dermal condensation in the underlying mesenchyme. Although epithelial signaling from the placode to mesenchyme is better described, little is known about primary mesenchymal signals resulting in placode induction. Using genetic approach in mice, we show that Meis2 expression in cells derived from the neural crest is critical for whisker formation and also for branching of trigeminal nerves. While whisker formation is independent of the trigeminal sensory innervation, MEIS2 in mesenchymal dermal cells orchestrates the initial steps of epithelial placode formation and subsequent dermal condensation. MEIS2 regulates the expression of transcription factor Foxd1, which is typical of pre-dermal condensation. However, deletion of Foxd1 does not affect whisker development. Overall, our data suggest an early role of mesenchymal MEIS2 during whisker formation and provide evidence that whiskers can normally develop in the absence of sensory innervation or Foxd1 expression.
Wing dimorphism is a common phenomenon that plays key roles in the environmental adaptation of aphid; however, the signal transduction in response to environmental cues and the regulation mechanism related to this event remain unknown. Adenosine (A) to inosine (I) RNA editing is a post-transcriptional modification that extends transcriptome variety without altering the genome, playing essential roles in numerous biological and physiological processes. Here, we present a chromosome-level genome assembly of the rose-grain aphid Metopolophium dirhodum by using PacBio long HiFi reads and Hi-C technology. The final genome assembly for M. dirhodum is 447.8 Mb, with 98.50% of the assembled sequences anchored to nine chromosomes. The contig and scaffold N50 values are 7.82 and 37.54 Mb, respectively. A total of 18,003 protein-coding genes were predicted, of which 92.05% were functionally annotated. In addition, 11,678 A-to-I RNA-editing sites were systematically identified based on this assembled M. dirhodum genome, and two synonymous A-to-I RNA-editing sites on CYP18A1 were closely associated with transgenerational wing dimorphism induced by crowding. One of these A-to-I RNA-editing sites may prevent the binding of miR-3036-5p to CYP18A1, thus elevating CYP18A1 expression, decreasing 20E titer, and finally regulating the wing dimorphism of offspring. Meanwhile, crowding can also inhibit miR-3036-5p expression and further increase CYP18A1 abundance, resulting in winged offspring. These findings support that A-to-I RNA editing is a dynamic mechanism in the regulation of transgenerational wing dimorphism in aphids and would advance our understanding of the roles of RNA editing in environmental adaptability and phenotypic plasticity.