Upstream kinases

The a subunits, for which two isoforms exist in mammals (al, a2), contain conventional protein serine/threonine kinase domains at the N-terminus, with a threonine residue in the activation loop (Thr-172) that must be phosphorylated by upstream kinases (see below) before the kinase is active. The kinase domain is followed by an autoinhibitory domain, whose effect is somehow relieved by interaction with the other subunits. The C-terminal domain of the a subunit is required for the formation of a complex with the C-terminal domain of the (3 subunit, which in turn mediates binding to the y subunit. The al and a2 catalytic subunit isoforms are widely distributed, although a2 is most abundant in muscle and may be absent in cells of the endothelial/hemopoietic lineage. 

AMPK can also be activated by a Ca2+-mediated pathway involving phosphorylation at Thr-172 by the Ca2+/calmodulin-dependent protein kinase, CaMKK 3. CaMKKa and CaMKK 3 were discovered as the upstream kinase for the calmodulin-dependent protein kinases-1 and -IV they both activate AMPK in a Ca2+/ calmodulin-dependent manner in cell-free assays, although CaMKK 3 appears to much more active against AMPK in intact cells. Expression of CaMKKa and CaMKK(3 primarily occurs in neural tissues, but CaMKKp is also expressed in some other cell types. Thus, the Ca2+-mediated pathway for AMPK activation has now been shown to occur in response to depolarization in rat neuronal tissue, in response to thrombin (acting via a Gq-coupled receptor) in endothelial cells, and in response to activation of the T cell receptor in T cells. 

Using chemical inhibitors of upstream kinases that potentially drive trichothecene-induced MAPK activation, two putative kinases have been identified double-stranded RNA- (dsRNA)-activated protein kinase (PKR)46 and hematopoietic cell kinase (Hck).47... 

Protein kinase B (Akt) is a serine/threonine, mitogen-regulated protein kinase involved in the protection of cells from apoptosis as well as the promotion of cell proliferation and diverse metabolic responses. It is activated upon binding of phospholipids and phosphorylation at residues Thr and Ser by upstream kinases such as phosphoinositide-dependent protein kinase 1 and 2 (M15). Activation of... 

Figure 15.3 Resveratrol modulation of pro-inflammatory signaling pathways. Resver-atrol represses transcriptional activation of various pro-inflammatory genes by inhibiting pro-inflammatory stimuli-induced activation of upstream kinases, such as MAP kinases, PI3K/Akt, IKK, PKC, etc., and blocking the DNA binding of eukaryotic transcription factors, such as NF-kB, AP-1, and STAT3.

Hawley, S. A., Boudeau, J., Reid, J. L., Mustard, K. J., Udd, L., Makela, T. P., Alessi, D. R. and Hardie, D. G., 2003, Complexes between the LKB1 tumor suppressor, STRAD alpha/beta and M025 alpha/beta are upstream kinases in the AMP-activated protein kinase cascade, J Biol, 2,... 

Hong, S. P., Feiper, F. C., Woods, A., Carling, D. and Carlson, M., 2003, Activation of yeast Snfl and mammalian AMP-activated protein kinase by upstream kinases, Proc Natl Acad Sci USA, 100, pp 8839-43. 

Fig. 21.1 Regulation of AMPK in cardiomyocyte. External metabolic stresses induce an increase in AMP/ATP ratio. Binding with AMP makes AMPK a better substrate to its upstream kinases, and a poor substrate for protein phosphatases 2C, which is known to dephosphorylate and inactivate AMPK. Moreover, insulin through activation of Akt also decreases AMPK activity in...

Woods, A., Johnstone, S. R., Dickerson, K., Leiper, F. C., Fryer, L. G., Neumann, D.,Schlattner, U., Wallimann, T., Carlson, M., and Carling, D. 2003a. LKB1 is the upstream kinase in the AMP-activated protein kinase cascade. Curr Biol 13 2004-2008. 

Type III inhibitors bind to sites distal to the ATP binding site and are often inactive in simple kinase enzyme assays. This apparent inactivity arises because these compounds can bind to the kinase and render it a poor substrate for an activating upstream kinase, thus disabling its activation. As a consequence, a cascade or cell-based assay may be required. 

Figure 13.13. ARE activation by Nrf2. Nrf2 associates with Keapl, a repressor protein of Nrf2, in the cytoplasm. Upon xenobiotic exposure to cells, upstream kinase pathways are activated and induce dissociation of Nrf2 from Keapl. The hberated Nrf2 translocates into nucleus and dimerize with a small Maf protein and bind to an ARE enhancer sequence identified in various phase II genes.

Many serine/threonine kinases require phosphorylation by an upstream kinase for activation. This requires cloning the upstream kinase as well as additional purification steps after phosphorylation [38, 39, 58]. In some cases, the active... 

Representative of upstream kinases which accept receptor signals are the Raf kinases. They are cytosolic Ser/Thr kinases. Raf kinase is recruited to the membrane and its activity is controlled by Ras. Raf phosphorylates and activates dual specificity, e.g. Ser/Thr-and Tyr-MAPKKs. The MAPKKs have a conserved serine/threonine phosphorylation site which is recognized and phosphorylated by Raf. 

Fig. 4.1 Growth-factor signals are channelled in the Raf/Ras pathway, resulting in cellular proliferation and differentiation, whereas stress and cytokine signals are directed via MAPKKKs and MAPKKs to JNKs 1 and 2 and the p38 MAP kinase. They cause growth inhibition and apoptosis. Second messengers, such as lipid messengers, may also adress the MAP kinase cascade and elicit specific cellular responses. Included in the scheme are upstream kinases, such as Raf, that in turn activate the MAPKKs (not shown), and eventually the p44 MAPKs 1 and 2 (or ERKs). (This scheme is simple and not complete, because some kinases participating in these signalling pathways are not yet defined.)...

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