Cellular, biochemical, and molecular mechanisms of insulin action (see also Chapter 12) Insulin receptor/insulin receptor tyrosine kinase
The insulin receptor is a glycoprotein consisting of two α subunits and two β subunits linked by disulfide bonds and expresses insulin-stimulated kinase activity directed towards its own tyrosine residues [12,84–87]. Insulin receptor phosphorylation of the α subunit, with subsequent activation of insulin receptor tyrosine kinase, represents the first step in the action of insulin on glucose metabolism. Insulin receptors devoid of tyrosine activity are completely ineffective in mediating insulin stimulation of cellular metabolism. Mutagenesis of any of the three major phosphorylation sites (at residue 1158, 1163, and 1162) impairs the insulin receptor kinase activity, and this is associated with a marked decreased in the acute metabolic and growth-promoting effects of insulin [12,86–88].
Insulin receptor signal transduction
After activation, insulin receptor tyrosine kinase phosphorylates specific intracellular proteins, of which at least nine have been identified (Figure 14.14). Six of these belong to the family of insulin-receptor substrate (IRS) proteins; IRS-1, IRS-2, IRS-3, IRS-4, IRS-5, IRS-6 (the others include Shc, Cb1, Gab-1, p60 [dok], and APS). In muscle, IRS-1 serves as the major docking protein that interacts with the insulin receptor tyrosine kinase and undergoes tyrosine phosphorylation in regions containing amino acid sequence motifs (YXXM or YMXM) that, when phosphorylated, serve as recognition sites for proteins containing src-homology 2 (SH2) domains. Mutation of these specific tyrosines severely impairs the ability of insulin to stimulate glycogen synthesis, establishing the important role of IRS-1 in insulin signal transduction [12,84 – 88]. In liver, IRS-2 serves as the primary docking protein that undergoes tyrosine phosphorylation and mediates the effect of insulin on hepatic glucose production, gluconeogenesis, and glycogen formation. In adipocytes, Cb1 is phosphorylated after its interaction with the insulin receptor tyrosine kinase and is required for stimulation of GLUT4 translocation .
In muscle, the phosphorylated tyrosine residues on IRS-1 mediate an association between the SH2 domain of the 85-kDa regulatory subunit of PI3K, leading to activation of the enzyme (Figure 14.14). PI3K is a heterodimeric enzyme composed of an 85-kDa regulatory subunit and a 110-kDa catalytic subunit. The latter catalyzes the 3′-phosphorylation of phosphatidylinositol (PI) in the plasma membrane glycolipids, thereby converting PI4,5-bisphosphate to PI3,4,5-triphosphate and PI4-phosphate to PI3,4 biphosphate. PI(3,4,5)P3 and PI(3,4)P2 lead to the stimulation of glucose transport [12,84–88]. Activation of PI3K by phosphorylated IRS-1 also stimulates glycogen synthase, via a process that involves activation of protein kinases B/Akt and subsequent inhibition of kinase such as GSK3 and activation of protein phosphatase-1 . Insulin stimulates glycogen synthesis by simultaneously activating glycogen synthase and inhibiting glycogen phosphorylase [89,90] (Figure 14.15). The effect of insulin is mediated via the PI3K pathway, which inactivates phosphatases, particularly PP-1. It is believed that PP-1 is the primary regulator of glycogen metabolism [84,89 – 91]. In skeletal muscle, PP-1 associates with a specific glycogen binding regulatory subunit, causing dephosphorylation (activation) of glycogen synthase. PP-1 also phosphorylates (inactivates) glycogen phosphorylase. The precise steps that link insulin receptor tyrosine kinase/PI3K activation to stimulation of PP-1 have yet to be defined. Akt has been shown to phosphorylate and thus inactivate GSK3. This decreases glycogen synthase phosphorylation, leading to activation of the enzyme. From the physiologic standpoint, it makes sense that activation of glucose transport and glycogen synthase should be linked to the same signaling mechanisms to provide a coordinated and efficient stimulation of intracellular glucose metabolism.
Activation of the insulin signal transduction system in insulin target tissues leads to the stimulation of glucose transport. The effect of insulin is brought about by the translocation of a large intracellular pool of GLUTs (associated with low density microsomes) to the plasma membrane [1,2,84,92]. As discussed earlier, there are five major, different facilitative GLUTs with distinctive tissue distributions. GLUT4, the insulin regulatable transporter, is found in insulin-sensitive tissues (muscle and adipocytes), has a Km of ∼5 mmol L−1, which is close to that of the fasting plasma glucose concentration and is associated with HK-II [19,93]. In adipocytes and muscle, its concentration in the plasma membrane increases markedly after exposure to insulin, and this increase is associated with a reciprocal decline in the intracellular GLUT4 pool. Acute physiologic hyperinsulinemia does not increase the total number of GLUT4 in muscle, even though several studies have demonstrated an increase in muscle GLUT4 mRNA. Using a novel isotopic dilution technique, the in vivo dose – response curve for the action of insulin on glucose transport in human forearm skeletal muscle has been described [94,95] (Figure 14.16). GLUT1 represents the predominant GLUT in the insulin-independent tissue (brain and erythrocytes). It is located primarily in the plasma membrane, where its concentration changes little after the addition of insulin. It has a low Km (∼1 mmol L−1 ) and is well suited for its function, which is to mediate basal glucose uptake. It is found in association with HK-I [19,93,96]. GLUT2 predominates in the liver and pancreatic β cells, where it is found in association with a specific HK, HK-IV. In the β cell, HK-IV is referred to as glucokinase . GLUT2 has a high Km (∼15 – 20 mmol L−1 ) and, as a consequence, the glucose concentration in cells expressing this transporter increases in direct proportion to the increase in plasma glucose concentration. This characteristic allows these cells to behave as glucose sensors.