Microarrayed Compound Screening (μARCS) to Identify Activators and Inhibitors of AMP-Activated Protein Kinase

Anderson, Steven N.; Cool, Barbara L.; Kifle, Lemma; Chiou, William J.; Egan, David A.; Barrett, Leo W.; Richardson, Paul L.; Frevert, Ernst U.; Warrior, Usha; Kofron, James L.; Burns, David J.

doi:10.1177/1087057103260592

Cited by 39 publications

(33 citation statements)

References 19 publications

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“…High-thoughput procedures have also been developed to screen for AMPK activators or inhibitors (e.g. [41]). We show here that SPR can detect AMPK interactors in complex protein mixtures when different conditions are satisfied: (i) availability of highly pure, active kinase, as we have established by polycistronic bacterial expression of full-length AMPK complex [29]; (ii) reversible, high density immobilization of the kinase on the chip surface for repeated use with fresh protein (e.g.…”

Section: Discussionmentioning

confidence: 99%

A two-dimensional screen for AMPK substrates identifies tumor suppressor fumarate hydratase as a preferential AMPKα2 substrate

Klaus

Polge

Zorman

et al. 2012

Journal of Proteomics

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AMP-activated protein kinase (AMPK) is emerging as a central cellular signaling hub involved in energy homeostasis and proliferation. The kinase is considered as a suitable target for pharmacological intervention in several energy-related pathologies like diabetes type II and cancer, although its signaling network is still incompletely understood. Here we apply an original two-dimensional in vitro screening approach for AMPK substrates that combines biophysical interaction based on surface plasmon resonance with in vitro phosphorylation. By enriching for proteins that interact with a specific AMPK isoform, we aimed to identify substrates that are also preferentially phosphorylated by this specific AMPK isoform. Application of this screen to full-length AMPK α2β2γ1 and soluble rat liver proteins identified the tumor suppressor fumarate hydratase (FH). FH was confirmed to interact with and to be preferentially phosphorylated by the AMPKα2 isoform by using yeast-two-hybrid and in vitro phosphorylation assays. AMPK-mediated phosphorylation of FH significantly increased enzyme activity in vitro and in vivo, suggesting that it is a bona fide AMPK substrate. In vivo, AMPKα2 is supposed to target the cytosolic/nuclear pools of FH, whose tumor suppressor function relies on DNA damage repair and inhibition of HIF-1α-signaling.

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Section: Discussionmentioning

confidence: 99%

A two-dimensional screen for AMPK substrates identifies tumor suppressor fumarate hydratase as a preferential AMPKα2 substrate

Klaus

Polge

Zorman

et al. 2012

Journal of Proteomics

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“…Metformin and the peroxisome proliferator-activated receptor ␥ agonist rosiglitazone have been reported to activate AMPK and are believed to contribute to their insulin-sensitizing actions in diabetic subjects (8). Based on its suggested role in diabetes (8) and obesity (50), there is a keen interest to identify activators and inhibitors of AMPK and their potential benefits in inflammatory diseases.…”

Section: Discussionmentioning

confidence: 99%

5-Aminoimidazole-4-Carboxamide Ribonucleoside: A Novel Immunomodulator with Therapeutic Efficacy in Experimental Autoimmune Encephalomyelitis

Nath

Giri

Prasad

et al. 2005

The Journal of Immunology

129

131

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Experimental autoimmune encephalomyelitis (EAE), an animal model of multiple sclerosis, is a Th1-mediated inflammatory demyelinating disease of the CNS. AMP-activated protein kinase was reported recently to have anti-inflammatory activities by negatively regulating NF-κB signaling. In this study, we investigated the prophylactic and therapeutic efficacy of an AMP-activated protein kinase activator, 5-aminoimidazole-4-carboxamide ribonucleoside (AICAR), in active and passive EAE induced by active immunization with PLP139–151 or MOG35–55 and in adoptive transfer of PLP139–151-sensitized T cells, respectively. In vivo treatment with AICAR exerted both prophylactic and therapeutic effects on EAE, attenuating the severity of clinical disease. The anti-inflammatory effects of AICAR were associated with the inhibition of the Ag-specific recall responses and inhibition of the Th1-type cytokines IFN-γ and TNF-α, whereas it induced the production of Th2 cytokines IL-4 and IL-10. Treatment of PLP139–151-specific T cells in vitro with AICAR decreased their expression of T-bet in response to IL-12, a Th1 transcription factor, whereas in response to IL-4, it induced the expression and phosphorylation of Th2 transcription factors GATA3 and STAT6, respectively. Moreover, treatment of APCs in vitro with AICAR inhibited their capability to present the proteolipid protein peptide to PLP139–151-specific T cells. In an irrelevant Th1-mediated, OT-2 TCR transgenic mouse model, AICAR impaired in vivo Ag-specific expansion of CD4+ T cells. Together, these findings show for the first time that AICAR is a novel immunomodulator with promising beneficial effects for the treatment of multiple sclerosis and other Th1-mediated inflammatory diseases.

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“…To evaluate the roles of AMPK and eEF2K pathways in NH125-induced eEF2 phosphorylation, we used the eEF2K inhibitor A-484954, a widely used AMPK inhibitor compound C (Cpd C) (25), and another proprietary AMPK inhibitor compound A (Cpd A) developed by Abbott Laboratories (26). PC3 cells were treated with the compounds at the indicated concentrations for 6 h before Western blot analysis (Fig.…”

Section: Induction Of Eef2 Phosphorylation Correlates With Growth Inhmentioning

confidence: 99%

1-Benzyl-3-cetyl-2-methylimidazolium iodide (NH125) Induces Phosphorylation of Eukaryotic Elongation Factor-2 (eEF2)

Chen

Gopalakrishnan

Bui

et al. 2011

Journal of Biological Chemistry

Self Cite

136

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Background: NH125 inhibits cancer cell growth through inhibition of eukaryotic elongation factor-2 kinase (eEF2K). Results: NH125 induces eEF2 phosphorylation (peEF2) through multiple pathways in cancer cells. Conclusion: NH125 is not an eEF2K inhibitor in cancer cells. Inhibition of cell growth correlates with induction of peEF2. Significance: NH125-induced peEF2 corrects a misconception and provides an opportunity for a new multipathway approach to anticancer therapies.

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Microarrayed Compound Screening (μARCS) to Identify Activators and Inhibitors of AMP-Activated Protein Kinase

Cited by 39 publications

References 19 publications

A two-dimensional screen for AMPK substrates identifies tumor suppressor fumarate hydratase as a preferential AMPKα2 substrate

A two-dimensional screen for AMPK substrates identifies tumor suppressor fumarate hydratase as a preferential AMPKα2 substrate

5-Aminoimidazole-4-Carboxamide Ribonucleoside: A Novel Immunomodulator with Therapeutic Efficacy in Experimental Autoimmune Encephalomyelitis

1-Benzyl-3-cetyl-2-methylimidazolium iodide (NH125) Induces Phosphorylation of Eukaryotic Elongation Factor-2 (eEF2)

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