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International Textbook of Diabetes Mellitus, 4th Ed., Excerpt #69: Regulation of Glucose Metabolism in Liver Part 5 of 11

Transcriptional regulation of gluconeogenesis

The gradual shift from the fed-to-fasted state is followed closely by hormonal regulation of genes encoding PEPCK, fructose-1,6-bisphosphatase, and the glucose-6-phosphatase catalytic subunit. The transcriptional regulation of these genes remains a developing research area and its many emerging nuances cannot be completely covered here. However, this regulation is accomplished largely through two transcription factors, CREB and FOXO, the nuclear receptors GR, HNF-4α, and the coactivator PGC-1α [21,22] (Figure 13.4). Classic glucoregulatory hormones such as insulin, glucagon, and glucocorticoids mediate gluconeogenic gene expression by promoting or inhibiting the interaction of these proteins with the promoter regions of these genes [23]. More recently described hormones such as adiponectin, and posttranslational mediators of hepatocyte energy status like AMPK [24] and sirtuins also act on these transcriptional regulators to influence glucose metabolism in liver, and are discussed later.


Induction of gluconeogenic gene expression

Fasting causes increased levels and stability of PEPCK mRNA [25,26]. The PEPCK gene promoter contains binding sites for more than a dozen transcription factors, nuclear receptors and co-regulators. This allows the promoter to be exquisitely responsive to nutrition and stress. The best-studied factors regulating PEPCK expression are its activation by glucagon and glucocorticoids, and its inhibition by insulin [25,26] (Figure 13.4). Glucagon activates its G-protein coupled receptor on the cell surface of hepatocytes and stimulates cyclic AMP (cAMP) production. cAMP binds protein kinase A (PKA) and activates its kinase domain. After diffusion into the nucleus, PKA phosphorylates Ser133 of the cAMP-response element binding protein (CREB). CREB then binds to the cyclic AMP response element (CRE) in the PEPCK and G6Pase gene promoters [27]. Binding of phosphorylated CREB to its CRE site nucleates the assembly of its co-activator, the CREB binding protein (CBP), and initiates transcription. The stability of this complex is modulated through phosphorylation of the interacting domains by several kinases, including Ca2+ /calmodulin-dependent kinases [21].

Transcription by CREB is also robustly potentiated by a sec- ond co-activator, cAMP-regulated transcriptional co-activator (CRTC2, or TORC2) (Figure 13.4). CRTC2 is maintained inactive by salt inducible kinases (SIK) which phosphorylate and sequester CRTC2 in the cytoplasm [21]. In addition to activating CREB, PKA also inhibits SIK, while other phosphatases, including calcineurin (a Ca2+/calmodulin-dependent phosphatase) dephosphorylate CRTC2. Dephosphorylated CRTC2 is then translocated into the nuclease and promotes transcription of CREB targets [21]. In addition to gluconeogenic genes, CREB also initiates the expression of other positive regulators of gluconeogenesis such as PGC-1a and NR4A, a nuclear receptor that also induces gluconeogenic gene expression. Thus, glucagon is a powerful impetus for regulating gluconeogenic gene expression in liver, but its complexity also allows for its actions to be moderated or magnified by other factors, as we will see later. Employing a similar paradigm, glucocorticoids bind to the cytosolic glucocorticoid receptor (GR) causing its dissociation from a chaperone protein and its translocation to the nucleus where it binds to a glucocorticoid response element (GRE) in the PEPCK promoter (Figure 13.4). Formation of the GR/GRE complex leads to recruitment of other proteins (albeit it different ones from those bound to the CREB/CRE complex), again leading to stimulation of PEPCK gene transcription.

Repression of gluconeogenic gene expression

Insulin represses glucagon and glucocorticoid induction of gluconeogenic gene expression through incompletely understood mechanisms that may be partially permissive in nature [28] (Figure 13.4). In contrast to glucagon action, which facilitates the binding of CREB to its promoters, insulin signaling excludes the forkhead transcription factor (FOXO1) from the nucleus and prohibits its binding to the promoters of PEPCK and G6pase [23]. Mouse studies in which insulin signaling and FOXO1 are acutely deleted have demonstrated that the upreg- ulation of gluconeogenic genes during loss of insulin action requires FOXO1 [28,29]. Binding of insulin to its receptor initiates phosphorylation of the insulin receptor substrates and PI3Kinase. These events trigger a broader phosphorylation cascade which mediates the metabolic effect of insulin. Akt1/2, also known as protein kinase B, is activated by PI3Kinase and is required for most of insulin’s actions on lipid and glucose metabolism [22]. Akt1/2 (mainly Akt2 in the normal mouse liver) acts on gluconeogenesis by phosphorylating FOXO1, which is then excluded from the nucleus, causing gluconeogenic genes to be repressed [22]. Notably, Akt1/2 also phosphorylate and deactivate proteins required for glucagon action such as SIK and PGC-1α (see later), and activates proteins required for lipogenesis [23].

Role of PGC-1-1alpha

PGC-1alpha is a co-activator that binds to multiple transcription factors and nuclear receptors during fasting and/or the energy challenged state [30]. Like other coactivators, PGC-1alpha acts on nuclear chromatin to facilitate the binding transcription factors to target genes. Although very low in liver of fed mice, PGC-1alpha increases dramatically in liver during fasting [31]. The full induction of gluconeogenesis requires functioning PGC-1alpha and several models of type 1 and type 2 diabetes have increased expression or activity of hepatic PGC-1alpha [32]. PGC-1alpha is activated in response to cAMP and CREB and is thus an important component of counterregulatory hormone signaling. Once expressed, PGC-1alpha activity is modified posttranslationally by phosphorylation and acetylation under the regulation of AMPK and sirtuins (see later). Active PGC-1alpha induces the gluconeogenic enzymes, PEPCK, fructose-1,6-bisphosphatase, and the catalytic subunit of the G6Pase complex by co-activating hepatocyte nuclear factor 4-alpha and FOXO1 [31] (Figure 13.4).

PGC-1alpha also mediates fat oxidation in liver during fasting by co-activating PPARalpha. The induction of energy production by PGC-1α is essential to support the energetic expense of gluconeogenesis during fasting and is an additional route of gluconeogenic control by PGC-1alpha [32].

In summary, the regulation of gluconeogenic gene expression is carried out by a rise in cAMP levels caused by counterregulatory hormones during fasting or exercise. These hormones activate CREB, increase PGC-1alpha expression and stimulate expression of gluconeogenic genes via FOXO1 and HNF4alpha. Their effects are antagonized by insulin in the fed state via Akt1-mediated phosphorylation of FOXO1, resulting in suppression of gluconeogenic gene expression.

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