Metformin is a widely-used drug that results in clear benefits in relation to glucose metabolism and diabetes-related complications. The mechanisms underlying these benefits are complex and still not fully understood. Physiologically, metformin has been shown to reduce hepatic glucose production, yet not all of its effects can be explained by this mechanism and there is increasing evidence of a key role for the gut. At the molecular level the findings vary depending on the doses of metformin used and duration of treatment, with clear differences between acute and chronic administration. Metformin has been shown to act via both AMP-activated protein kinase (AMPK)-dependent and AMPK-independent mechanisms; by inhibition of mitochondrial respiration but also perhaps by inhibition of mitochondrial glycerophosphate dehydrogenase, and a mechanism involving the lysosome. In the last 10 years, we have moved from a simple picture, that metformin improves glycaemia by acting on the liver via AMPK activation, to a much more complex picture reflecting its multiple modes of action. More work is required to truly understand how this drug works in its target population: individuals with type 2 diabetes.Electronic supplementary materialThe online version of this article (doi:10.1007/s00125-017-4342-z) contains a slideset of the figures for download, which is available to authorised users.

Protein kinase B lies "downstream" of phosphatidylinositide (PtdIns) 3-kinase and is thought to mediate many of the intracellular actions of insulin and other growth factors. Here we show that FKHR, a human homologue of the DAF16 transcription factor in Caenorhabditis elegans, is rapidly phosphorylated by human protein kinase B␣ (PKB␣) at Thr-24, Ser-256, and Ser-319 in vitro and at a much faster rate than BAD, which is thought to be a physiological substrate for PKB. The same three sites, which all lie in the canonical PKB consensus sequences (Arg-Xaa-Arg-Xaa-Xaa-(Ser/Thr)), became phosphorylated when FKHR was cotransfected with either PKB or PDK1 (an upstream activator of PKB). All three residues became phosphorylated when 293 cells were stimulated with insulin-like growth factor 1 (IGF-1). The IGF-1-induced phosphorylation was abolished by the PtdIns 3-kinase inhibitor wortmannin but not by PD 98059 (an inhibitor of the mitogen-activated protein kinase cascade) or by rapamycin. These results indicate that FKHR is a physiological substrate of PKB and that it may mediate some of the physiological effects of PKB on gene expression. DAF16 is known to be a component of a signaling pathway that has been partially dissected genetically and includes homologues of the insulin/IGF-1 receptor, PtdIns 3-kinase and PKB. The conservation of Thr-24, Ser-256, and Ser-319 and the sequences surrounding them in DAF16 therefore suggests that DAF16 is also a direct substrate for PKB in C. elegans.In recent years evidence has accumulated that many of the metabolic actions of insulin may be mediated by a protein kinase cascade that lies "downstream" of phosphatidylinositide (PtdIns) 1 3-kinase and the second messengers PtdIns(3,4,5)P 3 and PtdIns(3,4)P 2 (reviewed in Refs. 1 and 2). A central player in this cascade is protein kinase B (PKB, also called c-Akt).This enzyme is activated when it becomes phosphorylated at Thr-308 and Ser-473 (3) by 3-phosphoinositide-dependent protein kinases 1 and 2 (PDK1, PDK2), respectively (4 -7). The activation of PKB by PDK1 in vitro has an absolute requirement for PtdIns(3,4,5)P 3 or PtdIns(3,4)P 2 (4), and these mediators facilitate activation by binding to the pleckstrin homology domains of both PKB (5, 7) and PDK1 (8). Consistent with these observations, the phosphorylation of PKB at Thr-308, induced by either insulin or insulin-like growth factor 1 (IGF-1) is prevented by inhibitors of PtdIns 3-kinase (3). PDK2 has not yet been characterized although, like the phosphorylation of Thr-308, the insulin or IGF-1-induced phosphorylation of Ser-473 is prevented by inhibitors of PtdIns 3-kinase (3).PKB mediates the metabolic actions of insulin by phosphorylating regulatory proteins at serine or threonine residues that lie in Arg-Xaa-Arg-Xaa-Xaa-(Ser/Thr) motifs (9), of which the best characterized are the cardiac isoform of 6-phosphofructo-2-kinase (PFK2) (2, 10), the protein kinase glycogen synthase kinase 3 (GSK3) (11,12), and the mammalian target of rapamycin (mTOR) (13), as well as the proapo...

Insulin inhibits the expression of multiple genes in the liver containing an insulin response sequence (IRS) (CAAAA(C/T)AA), and we have reported that protein kinase B (PKB) mediates this effect of insulin. Genetic studies in Caenorhabditis elegans indicate that daf-16, a forkhead/winged-helix transcription factor, is a major target of the insulin receptor-PKB signaling pathway. FKHR, a human homologue of daf-16, contains three PKB sites and is expressed in the liver. Reporter gene studies in HepG2 hepatoma cells show that FKHR stimulates insulin-like growth factor-binding protein-1 promoter activity through an IRS, and introduction of IRSs confers this effect on a heterologous promoter. Insulin disrupts IRS-dependent transactivation by FKHR, and phosphorylation of Ser-256 by PKB is necessary and sufficient to mediate this effect. Antisense studies indicate that FKHR contributes to basal promoter function and is required to mediate effects of insulin and PKB on promoter activity via an IRS. To our knowledge, these results provide the first report that FKHR stimulates promoter activity through an IRS and that phosphorylation of FKHR by PKB mediates effects of insulin on gene expression. Signaling to FKHR-related forkhead proteins via PKB may provide an evolutionarily conserved mechanism by which insulin and related factors regulate gene expression.Insulin exerts important effects on gene expression in multiple tissues (1). In the liver, insulin suppresses the expression of a number of genes that contain a conserved insulin response sequence (IRS) 1 (CAAAA(C/T)AA), including insulin-like growth factor-binding protein-1 (IGFBP-1), apolipoprotein CIII (apoCIII), phosphoenolpyruvate carboxykinase (PEPCK), and glucose-6-phosphatase (2-6). This observation suggests that insulin may regulate the expression of multiple hepatic genes through a common mechanism. Insulin rapidly suppresses the expression of IGFBP-1 and PEPCK at the transcriptional level, and this effect is not disrupted by pretreatment with cycloheximide (7, 8), indicating that it is mediated by post-translational modification of pre-existing factors, perhaps by their phosphorylation. Specific factors that mediate the inhibitory effects of insulin on hepatic gene expression through a conserved IRS remain to be identified.Recent studies indicate that protein kinase B (PKB) functions downstream from phosphatidylinositol 3Ј-kinase (PI3K) in the insulin signaling pathway (9, 10) and that it plays an important role in mediating effects of insulin and related growth factors on glucose and amino acid transport, glycogen and protein synthesis, and cell survival (11)(12)(13)(14)(15)(16)(17)(18)(19). Following its activation, PKB is translocated to the nucleus where it may exert effects on gene expression (20,21). Activated PKB increases the expression of leptin and fatty acid synthase in adipocytes (22, 23) and suppresses PEPCK mRNA levels in liver-derived cells stimulated by cAMP and glucocorticoids (24), mimicking the effects of insulin. Based on studies using pha...