A common Greenlandic TBC1D4 variant confers muscle insulin resistance and type 2 diabetes.

The Greenlandic population, a small and historically isolated founder population comprising about 57,000 inhabitants, has experienced a dramatic increase in type 2 diabetes (T2D) prevalence during the past 25 years. Motivated by this, we performed association mapping of T2D-related quantitative traits in up to 2,575 Greenlandic individuals without known diabetes. Using array-based genotyping and exome sequencing, we discovered a nonsense p.Arg684Ter variant (in which arginine is replaced by a termination codon) in the gene TBC1D4 with an allele frequency of 17%. Here we show that homozygous carriers of this variant have markedly higher concentrations of plasma glucose (ß = 3.8 mmol l(-1), P = 2.5 × 10(-35)) and serum insulin (ß = 165 pmol l(-1), P = 1.5 × 10(-20)) 2 hours after an oral glucose load compared with individuals with other genotypes (both non-carriers and heterozygous carriers). Furthermore, homozygous carriers have marginally lower concentrations of fasting plasma glucose (ß = -0.18 mmol l(-1), P = 1.1 × 10(-6)) and fasting serum insulin (ß = -8.3 pmol l(-1), P = 0.0014), and their T2D risk is markedly increased (odds ratio (OR) = 10.3, P = 1.6 × 10(-24)). Heterozygous carriers have a moderately higher plasma glucose concentration 2 hours after an oral glucose load than non-carriers (ß = 0.43 mmol l(-1), P = 5.3 × 10(-5)). Analyses of skeletal muscle biopsies showed lower messenger RNA and protein levels of the long isoform of TBC1D4, and lower muscle protein levels of the glucose transporter GLUT4, with increasing number of p.Arg684Ter alleles. These findings are concomitant with a severely decreased insulin-stimulated glucose uptake in muscle, leading to postprandial hyperglycaemia, impaired glucose tolerance and T2D. The observed effect sizes are several times larger than any previous findings in large-scale genome-wide association studies of these traits and constitute further proof of the value of conducting genetic association studies outside the traditional setting of large homogeneous populations.

A novel domain cassette identifies Plasmodium falciparum PfEMP1 proteins binding ICAM-1 and is a target of cross-reactive, adhesion-inhibitory antibodies.

Cerebral Plasmodium falciparum malaria is characterized by adhesion of infected erythrocytes (IEs) to the cerebral microvasculature. This has been linked to parasites expressing the structurally related group A subset of the P. falciparum erythrocyte membrane protein 1 (PfEMP1) family of IE adhesion ligands and to IEs with affinity for ICAM-1. However, recent evidence has cast doubt on both these associations, tempering hopes of the feasibility of developing a vaccine based on ICAM-1-binding PfEMP1. In this study, we report the identification of a domain cassette (DC) present in group A var genes from six genetically distinct P. falciparum parasites. The three domains in the cassette, which we call DC4, had a high level of sequence identity and cluster together phylogenetically. Erythrocytes infected by these parasites and selected in vitro for expression of DC4 adhered specifically to ICAM-1. The ICAM-1-binding capacity of DC4 was mapped to the C-terminal third of its Duffy-binding-like ß3 domain. DC4 was the target of broadly cross-reactive and adhesion-inhibitory IgG Abs, and levels of DC4-specific and adhesion-inhibitory IgG increased with age among P. falciparum-exposed children. Our study challenges earlier conclusions that group A PfEMP1 proteins are not central to ICAM-1-specific IE adhesion and support the feasibility of developing a vaccine preventing cerebral malaria by inhibiting cerebral IE sequestration.

AMP-activated protein kinase regulates nicotinamide phosphoribosyl transferase expression in skeletal muscle.

Deacetylases such as sirtuins (SIRTs) convert NAD to nicotinamide (NAM). Nicotinamide phosphoribosyl transferase (Nampt) is the rate-limiting enzyme in the NAD salvage pathway responsible for converting NAM to NAD to maintain cellular redox state. Activation of AMP-activated protein kinase (AMPK) increases SIRT activity by elevating NAD levels. As NAM directly inhibits SIRTs, increased Nampt activation or expression could be a metabolic stress response. Evidence suggests that AMPK regulates Nampt mRNA content, but whether repeated AMPK activation is necessary for increasing Nampt protein levels is unknown. To this end, we assessed whether exercise training- or 5-amino-1-ß-D-ribofuranosyl-imidazole-4-carboxamide (AICAR)-mediated increases in skeletal muscle Nampt abundance are AMPK dependent. One-legged knee-extensor exercise training in humans increased Nampt protein by 16% (P < 0.05) in the trained, but not the untrained leg. Moreover, increases in Nampt mRNA following acute exercise or AICAR treatment (P < 0.05 for both) were maintained in mouse skeletal muscle lacking a functional AMPK α2 subunit. Nampt protein was reduced in skeletal muscle of sedentary AMPK α2 kinase dead (KD), but 6.5 weeks of endurance exercise training increased skeletal muscle Nampt protein to a similar extent in both wild-type (WT) (24%) and AMPK α2 KD (18%) mice. In contrast, 4 weeks of daily AICAR treatment increased Nampt protein in skeletal muscle in WT mice (27%), but this effect did not occur in AMPK α2 KD mice. In conclusion, functional α2-containing AMPK heterotrimers are required for elevation of skeletal muscle Nampt protein, but not mRNA induction. These findings suggest AMPK plays a post-translational role in the regulation of skeletal muscle Nampt protein abundance, and further indicate that the regulation of cellular energy charge and nutrient sensing is mechanistically related.

AMP-activated protein kinase controls exercise training- and AICAR-induced increases in SIRT3 and MnSOD.

The mitochondrial protein deacetylase sirtuin (SIRT) 3 may mediate exercise training-induced increases in mitochondrial biogenesis and improvements in reactive oxygen species (ROS) handling. We determined the requirement of AMP-activated protein kinase (AMPK) for exercise training-induced increases in skeletal muscle abundance of SIRT3 and other mitochondrial proteins. Exercise training for 6.5 weeks increased SIRT3 (p < 0.01) and superoxide dismutase 2 (MnSOD; p < 0.05) protein abundance in quadriceps muscle of wild-type (WT; n = 13-15), but not AMPK α2 kinase dead (KD; n = 12-13) mice. We also observed a strong trend for increased MnSOD abundance in exercise-trained skeletal muscle of healthy humans (p = 0.051; n = 6). To further elucidate a role for AMPK in mediating these effects, we treated WT (n = 7-8) and AMPK α2 KD (n = 7-9) mice with 5-amino-1-ß-D-ribofuranosyl-imidazole-4-carboxamide (AICAR). Four weeks of daily AICAR injections (500 mg/kg) resulted in AMPK-dependent increases in SIRT3 (p < 0.05) and MnSOD (p < 0.01) in WT, but not AMPK α2 KD mice. We also tested the effect of repeated AICAR treatment on mitochondrial protein levels in mice lacking the transcriptional coactivator peroxisome proliferator-activated receptor γ-coactivator 1α (PGC-1α KO; n = 9-10). Skeletal muscle SIRT3 and MnSOD protein abundance was reduced in sedentary PGC-1α KO mice (p < 0.01) and AICAR-induced increases in SIRT3 and MnSOD protein abundance was only observed in WT mice (p < 0.05). Finally, the acetylation status of SIRT3 target lysine residues on MnSOD (K122) or oligomycin-sensitivity conferring protein (OSCP; K139) was not altered in either mouse or human skeletal muscle in response to acute exercise. We propose an important role for AMPK in regulating mitochondrial function and ROS handling in skeletal muscle in response to exercise training.

Biotin starvation causes mitochondrial protein hyperacetylation and partial rescue by the SIRT3-like deacetylase Hst4p.

The essential vitamin biotin is a covalent and tenaciously attached prosthetic group in several carboxylases that play important roles in the regulation of energy metabolism. Here we describe increased acetyl-CoA levels and mitochondrial hyperacetylation as downstream metabolic effects of biotin deficiency. Upregulated mitochondrial acetylation sites correlate with the cellular deficiency of the Hst4p deacetylase, and a biotin-starvation-induced accumulation of Hst4p in mitochondria supports a role for Hst4p in lowering mitochondrial acetylation. We show that biotin starvation and knockout of Hst4p cause alterations in cellular respiration and an increase in reactive oxygen species (ROS). These results suggest that Hst4p plays a pivotal role in biotin metabolism and cellular energy homeostasis, and supports that Hst4p is a functional yeast homologue of the sirtuin deacetylase SIRT3. With biotin deficiency being involved in various metabolic disorders, this study provides valuable insight into the metabolic effects biotin exerts on eukaryotic cells.