Our laboratory studies the molecular mechanisms of insulin signal transduction, in an attempt to discover rational treatments for diabetes and its life-threatening complications. The storage and release of energy during feeding and fasting are essential for development, growth, reproduction, and survival. Insulin and other factors control a common set of intracellular signals that integrate these processes. Insulin resistance disconnects the central nervous system from a reliable estimate of nutrient availability and interferes with the way the body stores food and uses it for energy, growth, and reproduction. Diabetes occurs when insulin secretion fails to compensate for insulin resistance.
When blood glucose levels rise above 5 mM, insulin is secreted by the β cells in the pancreatic islets of Langerhans. This promotes the storage of glucose as glycogen in the liver and muscle, the retention of amino acids in muscle, and the synthesis of fatty acids in liver and their storage as triglycerides in adipose tissue. Diabetes mellitus occurs when insulin fails to perform its physiological function, owing either to an absolute lack of insulin (type 1 diabetes) or to a relative insulin insufficiency exacerbated by peripheral insulin resistance (type 2 diabetes). Although insulin was discovered more than 75 years ago, the molecular mechanisms by which insulin acts are only now being revealed through a multidisciplinary approach including genetics, biochemistry, and cell and molecular biology. We are using our modern view of human genetics to learn how the insulin-signaling system coordinates nutrient storage and utilization among the tissues in the body.
Much of our work on signaling pathways that mediate the insulin response was fueled by our discovery of the insulin receptor substrate (IRS) protein family. Like the receptors for other growth factors and cytokines, the insulin receptor has an extracellular domain that binds insulin. During insulin binding the intracellular tyrosine kinase is activated and mediates phosphorylation of the IRS proteins on multiple tyrosine residues; other receptors, including those for insulin-like growth factor (IGF) and various interleukins, also promote tyrosine phosphorylation of IRS proteins. Tyrosine phosphorylation sites in the IRS proteins interact with the Src-homology 2 (SH2) domains in various signaling proteins. The binding of SH2 proteins to IRS proteins initiates cascades of signals that mediate the insulin response. Enzymes and adaptor proteins regulated by the IRS proteins include phosphatidylinositol 3-kinase (PI 3-kinase), Grb-2, SHP2, and others.
During association with IRS1 or IRS2, PI 3-kinase is activated and its phospholipid products activate various serine kinases and recruit them to the plasma membrane. One of these kinases, PKB/AKT, activates additional kinases that promote multiple biological responses, including glucose transport, protein and glycogen synthesis, and cellular proliferation and survival. In addition to the PI 3-kinase cascade, IRS proteins engage Grb-2 to stimulate the Ras pathway and activate the mitogen-activated protein kinase cascade. The binding of SHP2 generates a complicated response, including feedback inhibition by dephosphorylation of the IRS protein. Finally, the insulin response is fine-tuned by the action of protein-tyrosine phosphatases and various serine kinases that alter the activity of the insulin receptor and the IRS proteins. When the relation between these signaling pathways is disrupted, insulin resistance occurs and contributes to the onset of glucose intolerance, obesity, and diabetes.
Since diabetes is a complicated, multisystem disease, mice provide a valuable experimental model to establish the relation between signal transduction pathways and metabolic regulation. Transgenic mice lacking the genes for Irs1 or Irs2 reveal a surprisingly close relation between the molecular regulation of insulin secretion and that of insulin action. We now understand that IRS1 and IRS2 play unique roles in mediating the effects of insulin and IGF1 on embryonic development, postnatal somatic growth, and glucose homeostasis: No embryos (16.5 days or older) lacking both genes have been detected in our studies, suggesting that signals coordinated by these IRS proteins are essential for embryonic development. However, deletion of Irs1 alone causes insulin resistance and growth retardation, but diabetes never occurs. Insulin signaling is diminished in skeletal muscle and adipose tissues but is surprisingly intact in liver. Without IRS1, mice develop mild glucose intolerance at midlife, but pancreatic β cells grow and increased insulin secretion compensates for the insulin resistance. Additional work is needed to understand how the IRS1 branch of the insulin/IGF-signaling system coordinates mammalian growth, and whether it might play a role in cancer.
By contrast, mice without Irs2 grow into normal-size adults that develop type 2 diabetes. Without IRS2, neonates are insulin resistant but display appropriate compensatory insulin secretion; as they age, however, the peripheral insulin resistance is exacerbated by failure of the pancreatic β cells. Analysis of islet size in 4-week-old Irs2–/– mice reveals a 50-percent reduction in β-cell mass compared to wild-type mice, and insulin content is also low. The β cells are barely detectable in pancreatic islets of young adults between 6 and 10 weeks of age, whereas glucagon-producing α cells are spared. Thus the IRS2 branch of the insulin/IGF-signaling system links peripheral insulin action to β-cell function and compensation.
Common type 2 diabetes in humans does not appear to develop as a consequence of the genetic mutations in IRS1 or IRS2. Inhibition of IRS protein function might, however, be a common path to diabetes. At the physiologic level, obesity, inactivity, and aging are intertwined with insulin resistance. Although moderate compensatory hyperinsulinemia might be well tolerated in the short term, chronic compensatory hyperinsulinemia exacerbates insulin resistance and contributes directly to β-cell failure, infertility, and diabetes. A common mechanism explaining the occurrence of acute and chronic insulin resistance in people is difficult to identify. Mutations in the insulin receptor are an obvious source, but these are rare and usually not accompanied by β-cell failure in humans.
The association between inflammation and insulin resistance has, however, been known for a long time. Activation of stress-induced kinases promote serine phosphorylation of the IRS proteins, which inhibits the tyrosine phosphorylation and signaling capacity. Various cytokines, such as TNFα or interleukin-1β, or metabolites, including circulating free fatty acids, diacylglycerol, ceramides, or glucose, promote serine phosphorylation of the IRS proteins. High doses of salicylates that inhibit stress-induced kinases also reverse hyperglycemia, hyperinsulinemia, and dyslipidemia in obese rodents. In the case of TNFα-induced insulin resistance, the c-Jun NH2-terminal kinase (Jnk) associates with IRS1 and promotes the phosphorylation of Ser307 (Ser312 in human IRS1). Phosphorylation of this residue inhibits the function of the phosphotyrosine-binding (PTB) domain to recruit IRS1 to the activated insulin receptor. Better strategies to inhibit the effects of stress-induced kinases on the IRS proteins might slow the development of diabetes and its complications.
Our mouse models provide unique tools to reveal the common link between various forms of diabetes. Autosomal-dominant forms of early-onset diabetes (maturity-onset diabetes of the young, MODY) are linked to mutations in glucokinase (MODY2) or transcription factors that promote β-cell growth and function, including HNF4α (MODY1), HNF1α (MODY 3), PDX1 (MODY4), HNF1β (MODY5), and NeuroD1/BETA2 (MODY6). PDX1 plays an important role in islet development and β-cell function, as it promotes pancreas formation during development and glucose-sensitive insulin secretion in adults. Consequently, genetic mutations in PDX1 cause glucose intolerance in humans and mice, increasing the risk for type 2 diabetes.
Unexpectedly, the IRS2-branch of the insulin/IGF1-signaling system promotes expression and function of Pdx1, which links directly the insulin/IGF-signaling system to β-cell function. Irs2–/– mice develop diabetes between 8 to 10 weeks of age, but Pdx1 haploinsufficiency caused diabetes in newborn Irs2–/– mice. By contrast, transgenic expression of Pdx1 restored β-cell mass and function in Irs2–/– mice and promoted glucose tolerance throughout life, as these mice survived for at least 20 months without diabetes. Our results suggest that dysregulation of Pdx1 during insulin resistance might be one of the common links between ordinary type 2 diabetes and MODY.
Our understanding of the relation between peripheral insulin signaling and insulin secretion provides important new insight into the pathophysiology of diabetes. Our study of the IRS2 branch of the insulin/IGF-signaling pathway has led us to investigate the female reproductive system, brain development, hypothalamic function, retinal growth and survival, and neovascularization. A full understanding of the molecules that regulate IRS protein function and signaling may reveal new targets for drug design to treat and cure type 2 diabetes.
