The addition of the N6-methyladenosine (m6A) mark to primary microRNAs by METTL3 in mammalian cells is found to promote the recognition of these microRNA precursors by DGCR8, a component of the microprocessor complex. While N6-methyl-adenosine (m6A) modification of RNA has been known since the 1970s, it was only recently, with the transcriptome-wide characterization of its pervasiveness, that much attention turned to its role in regulating cellular events. Sohail Tavazoie and colleagues have found that the addition of an m6A mark to primary microRNAs (miRNAs) by methyltransferase-like 3 (METTL3) is required for their recognition by DGCR8, the hairpin-binding component of the microprocessor complex. Consequently, in the absence of METTL3, the biogenesis of mature miRNAs is impeded globally in a cell-type-independent manner. One possible role of such a nuclear m6A mark is to distinguish genuine precursors from random RNAs with secondary structure. Altered METTL3 expression has been reported in various human cancers, suggesting a possible link to aberrant expression of miRNAs in cancerous cells. The first step in the biogenesis of microRNAs is the processing of primary microRNAs (pri-miRNAs) by the microprocessor complex, composed of the RNA-binding protein DGCR8 and the type III RNase DROSHA1,2,3,4. This initial event requires recognition of the junction between the stem and the flanking single-stranded RNA of the pri-miRNA hairpin by DGCR8 followed by recruitment of DROSHA, which cleaves the RNA duplex to yield the pre-miRNA product5. While the mechanisms underlying pri-miRNA processing have been determined, the mechanism by which DGCR8 recognizes and binds pri-miRNAs, as opposed to other secondary structures present in transcripts, is not understood. Here we find in mammalian cells that methyltransferase-like 3 (METTL3) methylates pri-miRNAs, marking them for recognition and processing by DGCR8. Consistent with this, METTL3 depletion reduced the binding of DGCR8 to pri-miRNAs and resulted in the global reduction of mature miRNAs and concomitant accumulation of unprocessed pri-miRNAs. In vitro processing reactions confirmed the sufficiency of the N6-methyladenosine (m6A) mark in promoting pri-miRNA processing. Finally, gain-of-function experiments revealed that METTL3 is sufficient to enhance miRNA maturation in a global and non-cell-type-specific manner. Our findings reveal that the m6A mark acts as a key post-transcriptional modification that promotes the initiation of miRNA biogenesis.

The protein hormone adiponectin is known to have many beneficial systemic effects, including promoting cell survival, anti-inflammation and insulin sensitivity. Phil Scherer and his colleagues have found that these pleiotropic effects are mediated by a ceramidase activity associated with the two known isoforms of the adiponectin receptor. The adipocyte-derived secretory factor adiponectin promotes insulin sensitivity, decreases inflammation and promotes cell survival. No unifying mechanism has yet explained how adiponectin can exert such a variety of beneficial systemic effects. Here, we show that adiponectin potently stimulates a ceramidase activity associated with its two receptors, AdipoR1 and AdipoR2, and enhances ceramide catabolism and formation of its antiapoptotic metabolite—sphingosine-1-phosphate (S1P)—independently of AMP-dependent kinase (AMPK). Using models of inducible apoptosis in pancreatic beta cells and cardiomyocytes, we show that transgenic overproduction of adiponectin decreases caspase-8-mediated death, whereas genetic ablation of adiponectin enhances apoptosis in vivo through a sphingolipid-mediated pathway. Ceramidase activity is impaired in cells lacking both adiponectin receptor isoforms, leading to elevated ceramide levels and enhanced susceptibility to palmitate-induced cell death. Combined, our observations suggest a unifying mechanism of action for the beneficial systemic effects exerted by adiponectin, with sphingolipid metabolism as its core upstream signaling component.

Adipose tissue can undergo rapid expansion during times of excess caloric intake. Like a rapidly expanding tumor mass, obese adipose tissue becomes hypoxic due to the inability of the vasculature to keep pace with tissue growth. Consequently, during the early stages of obesity, hypoxic conditions cause an increase in the level of hypoxia-inducible factor 1alpha (HIF1alpha) expression. Using a transgenic model of overexpression of a constitutively active form of HIF1alpha, we determined that HIF1alpha fails to induce the expected proangiogenic response. In contrast, we observed that HIF1alpha initiates adipose tissue fibrosis, with an associated increase in local inflammation. "Trichrome- and picrosirius red-positive streaks," enriched in fibrillar collagens, are a hallmark of adipose tissue suffering from the early stages of hypoxia-induced fibrosis. Lysyl oxidase (LOX) is a transcriptional target of HIF1alpha and acts by cross-linking collagen I and III to form the fibrillar collagen fibers. Inhibition of LOX activity by beta-aminoproprionitrile treatment results in a significant improvement in several metabolic parameters and further reduces local adipose tissue inflammation. Collectively, our observations are consistent with a model in which adipose tissue hypoxia serves as an early upstream initiator for adipose tissue dysfunction by inducing a local state of fibrosis.

Insulin resistance is currently a major health problem. This may be because of a marked decrease in daily physical activity during recent decades combined with constant food abundance. This lifestyle collides with our genome, which was most likely selected in the late Paleolithic era (50,000–10,000 BC) by criteria that favored survival in an environment characterized by fluctuations between periods of feast and famine. The theory of thrifty genes states that these fluctuations are required for optimal metabolic function. We mimicked the fluctuations in eight healthy young men [25.0 ± 0.1 yr (mean ± SE); body mass index: 25.7 ± 0.4 kg/m2] by subjecting them to intermittent fasting every second day for 20 h for 15 days. Euglycemic hyperinsulinemic (40 mU·min−1·m−2) clamps were performed before and after the intervention period. Subjects maintained body weight (86.4 ± 2.3 kg; coefficient of variation: 0.8 ± 0.1%). Plasma free fatty acid and β-hydroxybutyrate concentrations were 347 ± 18 and 0.06 ± 0.02 mM, respectively, after overnight fast but increased ( P < 0.05) to 423 ± 86 and 0.10 ± 0.04 mM after 20-h fasting, confirming that the subjects were fasting. Insulin-mediated whole body glucose uptake rates increased from 6.3 ± 0.6 to 7.3 ± 0.3 mg·kg−1·min−1 ( P = 0.03), and insulin-induced inhibition of adipose tissue lipolysis was more prominent after than before the intervention ( P = 0.05). After the 20-h fasting periods, plasma adiponectin was increased compared with the basal levels before and after the intervention (5,922 ± 991 vs. 3,860 ± 784 ng/ml, P = 0.02). This experiment is the first in humans to show that intermittent fasting increases insulin-mediated glucose uptake rates, and the findings are compatible with the thrifty gene concept.