Obesity-linked insulin resistance is a major precursor to the development of type 2 diabetes. Previous work has shown that phosphorylation of PPARγ (peroxisome proliferator-activated receptor γ) at serine 273 by cyclin-dependent kinase 5 (Cdk5) stimulates diabetogenic gene expression in adipose tissues1. Inhibition of this modification is a key therapeutic mechanism for anti-diabetic drugs that bind PPARγ, such as the thiazolidinediones and PPARγ partial agonists or non-agonists2. For a better understanding of the importance of this obesity-linked PPARγ phosphorylation, we created mice that ablated Cdk5 specifically in adipose tissues. These mice have both a paradoxical increase in PPARγ phosphorylation at serine 273 and worsened insulin resistance. Unbiased proteomic studies show that extracellular signal-regulated kinase (ERK) kinases are activated in these knockout animals. Here we show that ERK directly phosphorylates serine 273 of PPARγ in a robust manner and that Cdk5 suppresses ERKs through direct action on a novel site in MAP kinase/ERK kinase (MEK). Importantly, pharmacological inhibition of MEK and ERK markedly improves insulin resistance in both obese wild-type and ob/ob mice, and also completely reverses the deleterious effects of the Cdk5 ablation. These data show that an ERK/Cdk5 axis controls PPARγ function and suggest that MEK/ERK inhibitors may hold promise for the treatment of type 2 diabetes.
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Choi, J. H. et al. Anti-diabetic drugs inhibit obesity-linked phosphorylation of PPARγ by Cdk5. Nature 466, 451–456 (2010)
Choi, J. H. et al. Antidiabetic actions of a non-agonist PPARγ ligand blocking Cdk5-mediated phosphorylation. Nature 477, 477–481 (2011)
Yu, J. G. et al. The effect of thiazolidinediones on plasma adiponectin levels in normal, obese, and type 2 diabetic subjects. Diabetes 51, 2968–2974 (2002)
Tontonoz, P., Hu, E. & Spiegelman, B. M. Stimulation of adipogenesis in fibroblasts by PPARγ2, a lipid-activated transcription factor. Cell 79, 1147–1156 (1994)
Chawla, A., Schwarz, E. J., Dimaculangan, D. D. & Lazar, M. A. Peroxisome proliferator-activated receptor (PPAR) gamma: adipose-predominant expression and induction early in adipocyte differentiation. Endocrinology 135, 798–800 (1994)
Saltiel, A. R. & Kahn, C. R. Insulin signalling and the regulation of glucose and lipid metabolism. Nature 414, 799–806 (2001)
Krüger, M. et al. Dissection of the insulin signaling pathway via quantitative phosphoproteomics. Proc. Natl Acad. Sci. USA 105, 2451–2456 (2008)
De Fea, K. & Roth, R. A. Modulation of insulin receptor substrate-1 tyrosine phosphorylation and function by mitogen-activated protein kinase. J. Biol. Chem. 272, 31400–31406 (1997)
Zheng, Y. et al. Improved insulin sensitivity by calorie restriction is associated with reduction of ERK and p70S6K activities in the liver of obese Zucker rats. J. Endocrinol. 203, 337–347 (2009)
Lazar, D. F. et al. Mitogen-activated protein kinase kinase inhibition does not block the stimulation of glucose utilization by insulin. J. Biol. Chem. 270, 20801–20807 (1995)
Jiang, Z. Y. et al. Characterization of selective resistance to insulin signaling in the vasculature of obese Zucker (fa/fa) rats. J. Clin. Invest. 104, 447–457 (1999)
Jager, J. et al. Deficiency in the extracellular signal-regulated kinase 1 (ERK1) protects leptin-deficient mice from insulin resistance without affecting obesity. Diabetologia 54, 180–189 (2011)
Hawasli, A. H. et al. Cyclin-dependent kinase 5 governs learning and synaptic plasticity via control of NMDAR degradation. Nature Neurosci. 10, 880–886 (2007)
Eguchi, J. et al. Transcriptional control of adipose lipid handling by IRF4. Cell Metab. 13, 249–259 (2011)
Ohshima, T. et al. Targeted disruption of the cyclin-dependent kinase 5 gene results in abnormal corticogenesis, neuronal pathology and perinatal death. Proc. Natl Acad. Sci. USA 93, 11173–11178 (1996)
Hawasli, A. H. et al. Regulation of hippocampal and behavioral excitability by cyclin-dependent kinase 5. PLoS ONE 4, e5808 (2009)
Meijer, L. et al. Biochemical and cellular effects of roscovitine, a potent and selective inhibitor of the cyclin-dependent kinases cdc2, cdk2 and cdk5. Eur. J. Biochem. 243, 527–536 (1997)
Bach, S. et al. Roscovitine targets, protein kinases and pyridoxal kinase. J. Biol. Chem. 280, 31208–31219 (2005)
Shah, K., Liu, Y., Deirmengian, C. & Shokat, K. M. Engineering unnatural nucleotide specificity for Rous sarcoma virus tyrosine kinase to uniquely label its direct substrates. Proc. Natl Acad. Sci. USA 94, 3565–3570 (1997)
Sun, K.-H., De Pablo, Y., Vincent, F. & Shah, K. Deregulated Cdk5 promotes oxidative stress and mitochondrial dysfunction. J. Neurochem. 107, 265–278 (2008)
Tarricone, C. et al. Structure and regulation of the CDK5-p25(nck5a) complex. Mol. Cell 8, 657–669 (2001)
Hu, E., Kim, J. B., Sarraf, P. & Spiegelman, B. M. Inhibition of adipogenesis through MAP kinase-mediated phosphorylation of PPARγ. Science 274, 2100–2103 (1996)
Shao, D. et al. Interdomain communication regulating ligand binding by PPAR-γ. Nature 396, 377–380 (1998)
Rangwala, S. M. et al. Genetic modulation of PPARγ phosphorylation regulates insulin sensitivity. Dev. Cell 5, 657–663 (2003)
Sharma, P. et al. Phosphorylation of MEK1 by cdk5/p35 down-regulates the mitogen-activated protein kinase pathway. J. Biol. Chem. 277, 528–534 (2002)
Hornbeck, P. V. et al. PhosphoSitePlus: a comprehensive resource for investigating the structure and function of experimentally determined post-translational modifications in man and mouse. Nucleic Acids Res. 40, D261–D270 (2012)
Puigserver, P. et al. A cold-inducible coactivator of nuclear receptors linked to adaptive thermogenesis. Cell 92, 829–839 (1998)
Ohno, H., Shinoda, K., Spiegelman, B. M. & Kajimura, S. PPARγ agonists induce a white-to-brown fat conversion through stabilization of PRDM16 protein. Cell Metab. 15, 395–404 (2012)
Flaherty, K. T. et al. Combined BRAF and MEK inhibition in melanoma with BRAF V600 mutations. N. Engl. J. Med. 367, 1694–1703 (2012)
Solit, D. B. et al. BRAF mutation predicts sensitivity to MEK inhibition. Nature 439, 358–362 (2006)
Banks, A. S. et al. Dissociation of the glucose and lipid regulatory functions of FoxO1 by targeted knockin of acetylation-defective alleles in mice. Cell Metab. 14, 587–597 (2011)
Barrett, S. D. et al. The discovery of the benzhydroxamate MEK inhibitors CI-1040 and PD 0325901. Bioorg. Med. Chem. Lett. 18, 6501–6504 (2008)
Albeck, J. G., Mills, G. B. & Brugge, J. S. Frequency-modulated pulses of ERK activity transmit quantitative proliferation signals. Mol. Cell 49, 249–261 (2013)
Lau, K. S. et al. In vivo systems analysis identifies spatial and temporal aspects of the modulation of TNF-α-induced apoptosis and proliferation by MAPKs. Sci. Signal. 4, ra16 (2011)
Robinson, M. J., Stippec, S. A., Goldsmith, E., White, M. A. & Cobb, M. H. A constitutively active and nuclear form of the MAP kinase ERK2 is sufficient for neurite outgrowth and cell transformation. Curr. Biol. 8, 1141–1150 (1998)
Patricelli, M. P. et al. In situ kinase profiling reveals functionally relevant properties of native kinases. Chem. Biol. 18, 699–710 (2011)
McAllister, F. E. et al. Mass spectrometry based method to increase throughput for kinome analyses using ATP probes. Anal. Chem. 85, 4666–4674 (2013)
Rappsilber, J., Mann, M. & Ishihama, Y. Protocol for micro-purification, enrichment, pre-fractionation and storage of peptides for proteomics using StageTips. Nature Protocols 2, 1896–1906 (2007)
Kettenbach, A. N. & Gerber, S. A. Rapid and reproducible single-stage phosphopeptide enrichment of complex peptide mixtures: application to general and phosphotyrosine-specific phosphoproteomics experiments. Anal. Chem. 83, 7635–7644 (2011)
Ting, L., Rad, R., Gygi, S. P. & Haas, W. MS3 eliminates ratio distortion in isobaric multiplexed quantitative proteomics. Nature Methods 8, 937–940 (2011)
McAlister, G. C. et al. Increasing the multiplexing capacity of TMTs using reporter ion isotopologues with isobaric masses. Anal. Chem. 84, 7469–7478 (2012)
Huttlin, E. L. et al. A tissue-specific atlas of mouse protein phosphorylation and expression. Cell 143, 1174–1189 (2010)
Eng, J. K., McCormack, A. L. & Yates, J. R. An approach to correlate tandem mass spectral data of peptides with amino acid sequences in a protein database. J. Am. Soc. Mass Spectrom. 5, 976–989 (1994)
Elias, J. E. & Gygi, S. P. Target-decoy search strategy for increased confidence in large-scale protein identifications by mass spectrometry. Nature Methods 4, 207–214 (2007)
We thank E. Rosen for providing us with the adiponectin Cre mice before their initial publication; members of the Spiegelman laboratory (Dana-Farber Cancer Institute) and D. Cohen (Brigham and Women's Hospital) for discussions; and C. Palmer and K. LeClair for reading the manuscript. B.M.S. acknowledges National Institutes of Health (NIH) grant DK31405. A.B. acknowledges NIH grant DK93638, the Harvard University Milton Fund, and the Harvard Digestive Disease Center, Core D.
B.M.S. is a consultant to and shareholder in Ember Therapeutics. The remaining authors declare no competing interests.
Extended data figures and tables
Extended Data Figure 1 Metabolic profiling of adipose-specific Cdk5-KO mice on a standard chow diet.
a–e, Fasting plasma levels of glucose (a), insulin (b), total triacylglycerols (c), free fatty acids (FFA) (d) and total cholesterol (e) (n = 16 (control) and 17 (KO)). f, g, Body weights (f) and intraperitoneal glucose tolerance test (g). Mice were 12 weeks of age (n = 14 (control) and 11 (KO)). No significant differences were observed. Error bars indicate s.e.m.
Extended Data Figure 2 Energy homeostasis of adipose-specific Cdk5-KO mice maintained on a high-fat diet.
a–f, After a 48-h acclimatization period, singly housed mice were monitored for oxygen consumption (VO2) (a), carbon dioxide production (VCO2) (b), respiratory exchange ratio (RER) (c), ambulatory locomotor activity (d), cumulative food intake (e) and body weights (f) (n = 8 per group). Shaded areas signify the dark phase of the light cycle. No significant differences were observed. Error bars indicate s.e.m.
Brown adipose tissue protein lysates from mice maintained on a high-fat diet for 12 weeks. Blotting for phospho-p38, phospho-JNK and phospho-S473 and pT308 AKT was performed before loading for total protein amounts.
Mouse MEK2 T395/T397 corresponds to human MEK2 T394/T396. These sites share identity with MEK1 T386/T388 in both humans and mouse. Cdk5 has previously been shown to phosphorylate MEK1 at T286, a site not shared with MEK2. ERK has been shown to phosphorylate MEK1 T386 and contribute to regulation of kinase activity31. Homo, Homo sapiens; trog, Pan troglodytes; mus, Mus musculus; rat, Rattus norvegicus; bos, Bos taurus; canis, Canis lupus familiaris.
Extended Data Figure 5 Body weight of control and of adipose-specific Cdk5-KO mice maintained on a high-fat diet after treatment with PD0325901.
Treatment similar to that in Fig. 4a–c. The body weights are not significantly different by ANOVA. Error bars indicate s.e.m.
a–c, Glucose tolerance test (a), adiponectin levels (b) and body weights (c) of ob/ob mice treated with PD0325901 (n = 7 (vehicle) and 8 (PD)). *P ≤ 0.05 by Student’s t-test. Error bars indicate s.e.m.
Extended Data Figure 7 Inflammatory markers in epididymal white adipose tissue from ob/ob mice treated with MEK inhibitors.
Gene expression analysis was performed on M1 macrophage markers Nos2 and tumour necrosis factor-α (TNF-α); M2 macrophage markers Arg1, Chil3, Il10, Itgax, Clec10a/Mgl1 and Mgl2; chemotactic ligand Ccl2 and receptor Ccr2; and macrophage surface markers Emr1, Cd68 and Csf1r (n = 7 or 8 mice per group as in Fig. 5f, h). Gene expression was analysed by ANOVA. Error bars indicate s.e.m. *P < 0.05; **P < 0.01; ***P < 0.001.
a, In the lean state, PPARγ is not phosphorylated. b, In the obese state, S273 phosphorylation is driven by both Cdk5 and ERK with CDK5 repressing MEK and ERK activity. c, Cdk5-KO results in derepression of MEK and ERK kinases and increased phosphorylation of S273 PPARγ. d, MEK inhibition markedly decreases S273 PPARγ phosphorylation. e, PPARγ ligands, including the thiazolidinediones, block the accessibility of S273 PPARγ by either ERK or CDK5 kinases.
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Banks, A., McAllister, F., Camporez, J. et al. An ERK/Cdk5 axis controls the diabetogenic actions of PPARγ. Nature 517, 391–395 (2015). https://doi.org/10.1038/nature13887
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