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Insulin, Diabetes, and Obesity

Insulin, Diabetes, and Obesity. Insulin signalling mechanisms: The insulin receptor is a tyrosine kinase (RTK) Signalling through phosphoinositols and MAPK’s Mechanism of insulin resistance and Type II diabetes

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Insulin, Diabetes, and Obesity

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  1. Insulin, Diabetes, and Obesity • Insulin signalling mechanisms: • The insulin receptor is a tyrosine kinase (RTK) • Signalling through phosphoinositols and MAPK’s • Mechanism of insulin resistance and Type II diabetes • Control of glucose levels through both energy homeostasis and hormonal/neuronal mechanisms • Connections to obesity Jason Kahn: Diabetes and Obesity

  2. Consequences of Obesity • The chart shows the estimated impact of obesity on life expectancy at birth in the USA. • This reduction in life expectancy is comparable to the effect of all accidental deaths combined. • Obesity is second only to smoking as a preventable cause of premature death. • Is this really how we want to save Social Security? • Source: Ohlansky et al. (2005) New England Journal of Medicine, vol. 352, p. 11 (and see dissenting editorial) Jason Kahn: Diabetes and Obesity

  3. The Insulin Receptor is an RTK • Receptor tyrosine kinases are transmembrane glycoproteins with external ligand-bindng domains and internal tyrosine kinase domains. • They are activated by ligand-induced dimerization or else changes in the orientation of monomers ina pre-existing dimer. Typically they respond to protein growth factors. • The insulin receptor (InsR) subfamily members are disulfide-linked 22 heterotetramers processed from a single precursor polypeptide. V+V 3/e, Chapter 19 Jason Kahn: Diabetes and Obesity

  4. Insulin Receptor Structure • RTK monomers within a dimer cross-phosphorylate each others’ Tyr residues on the “activation loop” of the kinase domain • The RTK also phosphorylates cytoplasmic target proteins • Tyr phosphorylation causes a conformational change in the activation loop which makes it part of the active site rather than blocking ATP binding • Subunit rotation closes the active site around ATP N-term domain Unphosphorylated P-Tyr Phosphorylated Activation Loop Substrate peptide C-term domain V+V 3/e, Chapter 19 Jason Kahn: Diabetes and Obesity

  5. Signalling by RTKs • Phosphotyrosine residues in peptides are specially recognized by SH2 domains (named for Src Homology; SH3 domains bind proline-rich peptides) and also by PTB (phosphotyrosine-binding) domains • Thus RTK auto-cross-phosphorylation outside of the activation loops recruits SH2-domain containing proteins like the Src tyrosine kinase or PTB-containing domains • The Insulin receptor acts through phosphorylating Tyr residues on the PTB-containing Insulin-Responsive Substrate proteins IRS1, IRS2, IRS3, and IRS4, which then recruit SH2-domain proteins • Many other adapter domains help assemblesignalling complexes (WW, PDZ, PH =pleckstrinhomology) V+V 3/e, Chapter 19 SH2 Domain P-Tyr binding pocket Figure 19-32 The NMR structure of the PTB domain of Shc in complex with a 12-residue polypeptide from the Shc binding site of a nerve growth factor (NGF) receptor. Jason Kahn: Diabetes and Obesity

  6. Signalling through Phosphoinositides • Because there is such a diversity of adapter modules, RTK’s can assemble a wide variety of downstream target proteins • The insulin receptor acts through the four IRS proteins. The PH domains of the IRS proteins interact with membrane polyphosphoinositides, localizing them to the plasma membrane. The PTB domain of the IRS targets it to the insulin receptor, which then phosphorylates tyrosine residues that then become docking sites for SH2 domains. • There are 8 possible phosphoinositides with PO3-at positions 3, 4, and/or 5. They are synthesizedby phosphoinositide kinases. The productsact as second messengers by recruiting targetsto the membrane. • The phosphorylated IRS-1 protein recruits a class IAphosphoinositide-3-kinase (PI-3K, , or ) throughthe SH2 domains of its p85regulatory subunit. Phospatidyl inositides: Jason Kahn: Diabetes and Obesity

  7. Phosphoinositides II • Sugars are information-rich,in an analog rather than the digital way of nucleic acids (…glyomics). The sterecochemistry of hydroxyls, modifications, identity of sugars in a polymer, branching, etc. are important in signaling, cell-cell interaction, protein trafficking, etc. • Membranes are good places to organize signalling complexes -- it’s a 2-D rather than 3-D search, and rafts and caveolae can create additional levels of organization Jason Kahn: Diabetes and Obesity

  8. Control of Glycogen Metabolism • (See Gomperts et al., esp. chapters 13 and 17) • The p110 catalytic subunit of PI3K phosphorylates PI(4,5)P2 (inositol phosphorylated at positions 4 and 5) to give the second messenger PI(3,4,5)P3 • PI(3,4,5)P3 recruits both Protein Kinase B (PKB, a.k.a. Akt) and PDK1 (PI(3)-dependent protein kinase) to the membrane, via their PH domains. Phosphorylation of PKB by PDK1 (and PDK2) releases it from the membrane in an activated form. • PKB inactivates glycogen synthase kinase-3 = GSK-3 = of course, in Greek), which thus reduces phosphorylation of glycogen synthase and promotes glycogen synthesis. • PI3K also phosphorylates FRAP/mTOR. This relieves inhibition of translation initiation factor eIF-4E,and FRAP/mTOR also activates the S6 protein of the 40S ribosomal subunit. This promotes protein biosynthesis. • IRS-1 also activates the ISPK(insulin-stimulated protein kinase),which we recall phosphorylatesthe GM subunit of proteinphosphatase 1 (PP1),activating PP1c andstimulating glycogensynthesis. Reset by phosphatases PP2A and PP2B (calcineurin) Jason Kahn: Diabetes and Obesity

  9. Signalling through PI3 Kinase • PI3K binds the IRS, catalyzes PIP3 formation, hence recruits PKB Control of kinases by membrane localization and release Jason Kahn: Diabetes and Obesity

  10. Review and Overview • Figures from Saltiel and Kahn (2001), Nature vol. 414. Special Insight section on diabetes. • We don’t need to go through every pathway in detail. Jason Kahn: Diabetes and Obesity

  11. Branching of Pathways • Recruitment of other proteins to the RTK or IRS’s activates MAP kinase and other pathways that control gene expression and glucose transport. Jason Kahn: Diabetes and Obesity

  12. Branching of Pathways II RTK’s can recruit many different downstream targets …leading to activation of many different pathways • Images from Gomperts Specificity comes from colocalizationvia protein-protein interactions and protein-membrane interactions. Jason Kahn: Diabetes and Obesity

  13. Effects on Gene Expression • Insulin signalling helps control the levels of the proteins involved in glycolysis vs. gluconeogenesis and fatty acid synthesis. Jason Kahn: Diabetes and Obesity

  14. Shutting Off the Signal…What Can Go Wrong? • Normal homeostatic mechanisms: • Protein tyrosine phosphatases (PTP’s) that dephosphorylate the Insuline receptor and IRS’s • The related PTEN and SHIP2 phosphatases dephosphorylate PI(3,4,5)P3 to shut off the PI3K signalling (possible drug targets…). • Ser/Thr phosphorylation of IRS proteins by the PI3K pathway reduces their suitability as substrates for tyrosine phosphorylation. This is a normal negative feedback mechansim, but… • Mechanisms contributing to Type II Diabetes: • Ser/Thr phosphorylation of the IRS proteins is also carried out by protein kinase C isoforms that are upregulated by the metabolites resulting from fat metabolism. • So…excessive concentrations of circulating free fatty acids leads to a decrease in the body’s ability to respond to insulin = insulin resistance. This often precedes diabetes: the b cells have to make more insulin in order for it to take effect, and eventually they are unable to keep up. • Excessive FFA inhibits glucose uptake: the body thinks it has enough stored energy. • Adipocytes (fat cells) also secrete hormones (TNF-) that lead to IRS phosphorylation. • Adipocytes also secrete leptin, a hormone which regulates appetite. Absence of leptin or failure to respond to leptin is associated with obesity and insulin resistance. • The connection to obesity is clear, although there are many factors at work. • Consequences: • Starvation in the midst of plenty. If insulin signalling does not work, then muscle cells fail to take up glucose and circulating levels increase. Gluconeogenesis is activated. Protein is metabolized to make more glucose. Fat stores are mobilized to make ketone bodies. • Effects: Wasting away. Ketoacidosis. A decreased blood pH leads to excessive excretion of ions and water by the kidneys. Glycalation of proteins. Jason Kahn: Diabetes and Obesity

  15. IRS Phosphorylation • V+V 3/e chapter 27 • Eventually IRS phosphorylation leads to apoptosis of the  cells, and when the  cell mass declines too far, diabetes results. • Inflammation, stress, hyperglycemia, hyperlipidemia etc. can lead to IRS-2 degradation. • See Rhodes, 2005, Science, 307, p. 380. Jason Kahn: Diabetes and Obesity

  16. Current Directions • See special section in Science, January 21, 2005. • Genetics (O’Rahilly et al., p. 370): • Alleles that predispose people to diabetes are being identified through linkage studies in families and by comparison to knock-out mice. • Diabetes is heterogeneous and polygenic, with some dramatic defects but many mutations that give only modest “odds ratios.” • Unfortunately knock out mice often do not display the expected phenotypes, e.g. hyperglycemia. • Will require a lot of work to sort out life-style effects from differential genetic sensitivity effects. • There are also epigenetic effects, as suggested by correlations between poor fetal nutrition and subsequent diabetes (a “thrifty phenotype”). Jason Kahn: Diabetes and Obesity

  17. Evolutionary Aspects • Lazar, p. 373 • Why are we so poorly adapted to modern life? • We were not selected under conditions of unlimited food and no need for exercise. • “Thrifty genes” that cause the aggressive storage of energy could have been selected for by famine and malnutrition. They are good at listening to glucagon, haven’t had much need to deal with long-term hyperglycemia or hyperlipidemia. (Populations that have historically lived under harsh conditions are particularly vulnerable to MacDonalds, e.g. the Pima tribe of native Americans.) • Inflammation, infection, and stress reduce responsiveness to insulin, presumably originally in order to conserve energy needed to recover or fight infection. Jason Kahn: Diabetes and Obesity

  18. Brain-body Connections • Schwartz and Porte, p. 375. • The brain’s use of glucose is not regulated by insulin • However, the brainresponds to leptinand insulin to control appetite and energy balance. • Hypothalamic responses to free fatty acids may help control glucose homeostasis by the liver, but in general the mechanism by which the brain helps control metabolism are not clear. Jason Kahn: Diabetes and Obesity

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