| 2. |
- Göransson, Olga, et al.
(author)
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Mechanism of action of A-769662, a valuable tool for activation of AMP-activated protein kinase
- 2007
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In: Journal of Biological Chemistry. - 1083-351X. ; 282:45, s. 32549-32560
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Journal article (peer-reviewed)abstract
- We have studied the mechanism of A- 769662, a new activator of AMP- activated protein kinase ( AMPK). Unlike other pharmacological activators, it directly activates native rat AMPK by mimicking both effects of AMP, i. e. allosteric activation and inhibition of dephosphorylation. We found that it has no effect on the isolated alpha subunit kinase domain, with or without the associated autoinhibitory domain, or on interaction of glycogen with the beta subunit glycogen- binding domain. Although it mimics actions of AMP, it has no effect on binding of AMP to the isolated Bateman domains of the gamma subunit. The addition of A- 769662 to mouse embryonic fibroblasts or primary mouse hepatocytes stimulates phosphorylation of acetyl- CoA carboxylase ( ACC), effects that are completely abolished in AMPK- alpha 1(-/-) alpha 2(-/-) cells but not in TAK1(-/-) mouse embryonic fibroblasts. Phosphorylation of AMPK and ACC in response to A- 769662 is also abolished in isolated mouse skeletal muscle lacking LKB1, a major upstream kinase for AMPK in this tissue. However, in HeLa cells, which lack LKB1 but express the alternate upstream kinase calmodulin- dependent protein kinase kinase-beta, phosphorylation of AMPK and ACC in response to A- 769662 still occurs. These results show that in intact cells, the effects of A- 769662 are independent of the upstream kinase utilized. We propose that this direct and specific AMPK activator will be a valuable experimental tool to understand the physiological roles of AMPK.
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| 3. |
- Lizcano, Jose M, et al.
(author)
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LKB1 is a master kinase that activates 13 kinases of the AMPK subfamily, including MARK/PAR-1
- 2004
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In: EMBO Journal. - : Wiley. - 1460-2075 .- 0261-4189. ; 23:4, s. 833-843
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Journal article (peer-reviewed)abstract
- We recently demonstrated that the LKB1 tumour suppressor kinase, in complex with the pseudokinase STRAD and the scaffolding protein MO25, phosphorylates and activates AMP-activated protein kinase (AMPK). A total of 12 human kinases (NUAK1, NUAK2, BRSK1, BRSK2, QIK, QSK, SIK, MARK1, MARK2, MARK3, MARK4 and MELK) are related to AMPK. Here we demonstrate that LKB1 can phosphorylate the T-loop of all the members of this subfamily, apart from MELK, increasing their activity >50-fold. LKB1 catalytic activity and the presence of MO25 and STRAD are required for activation. Mutation of the T-loop Thr phosphorylated by LKB1 to Ala prevented activation, while mutation to glutamate produced active forms of many of the AMPK-related kinases. Activities of endogenous NUAK2, QIK, QSK, SIK, MARK1, MARK2/3 and MARK4 were markedly reduced in LKB1-deficient cells. Neither LKB1 activity nor that of AMPK-related kinases was stimulated by phenformin or AICAR, which activate AMPK. Our results show that LKB1 functions as a master upstream protein kinase, regulating AMPK-related kinases as well as AMPK. Between them, these kinases may mediate the physiological effects of LKB1, including its tumour suppressor function.
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