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Metabolic signalling

For cells and organisms to survive and grow it is critical that energy supply matches energy demand. The demand for energy is a continuously varying parameter; we are interested in studying the processes involved in synchronizing metabolic pathways (those that produce energy vs those that consume it) to maintain the perfect balance.

Research Overview

A major focus of our research is investigating regulation of an enzyme called AMP-activated protein kinase (AMPK). Analogous to a car’s fuel gauge, AMPK detects when energy in the cell is low and co-ordinates multiple branches of metabolism (e.g. fat burning, protein synthesis, digestion of cellular components) to redress energy imbalance. AMPK also has body-wide effects, being a key regulator of appetite and responsible for adaption to exercise. These roles have elevated AMPK to its current standing as an attractive drug target for diseases such as type 2 diabetes, cardiovascular disease and cancer.

AMPK represents a nexus in a complicated network of signalling pathways that act as ‘detection systems’, allowing the cell to sense environmental stresses and availability of nutrients. Our aim is to uncover how these signalling pathways interact, the components involved and how we can use them to exploit the therapeutic potential of AMPK. Working closely with the Protein Chemistry & Metabolism Unit we use a combination of biochemical and cell-based techniques, protein crystallography, medicinal chemistry, mass spectrometry and animal models to provide insight into the regulatory control of this important enzyme.

Research Themes

Signalling pathways regulating AMPK

AMP-activated protein kinase (AMPK) is a heterotrimeric serine/threonine protein kinase consisting of an α catalytic subunit and regulatory β and γ subunits. Multiple isoforms exist for each subunit (α1/2, β1/2 and γ1/2/3) with each displaying tissue-specific expression profiles. Given the wide range of cellular effects attributed to AMPK it is unsurprising that the enzyme is subject to complex regulation, not only by adenine nucleotides but also other signalling networks. Phosphoproteomic analyses have identified >100 phosphorylation sites on AMPK, however understanding of the biological consequences of these phosphorylation events is limited to just a handful. For example phosphorylation of Thr172 in the activation loop of the α-subunit kinase domain by LKB1 or CaMKKβ activates the enzyme and facilitates signalling in response to increases in AMP/ATP ratio. Alternatively, phosphorylation of Ser485 in the α-subunit C-terminus by Akt (or by autophosphorylation) leads to suppression of Thr172 phosphorylation and down-regulation of AMPK signalling. Other phosphorylation sites are predicted to localize AMPK to specific cellular components or membranes, thereby conferring temporospatial specificity to AMPK signalling. Expanding upon the known signaling networks that communicate cellular state to AMPK is vitally important given its central role in energy homeostasis.

AMPK regulation by small molecules

The molecule adenosine triphosphate (ATP) is regarded as the molecular unit of currency of intracellular energy transfer. It provides the energy used to drive virtually every cellular process, from muscle contraction to DNA synthesis. Human adults make ~50kg ATP daily due to rapid turnover to the low energy monophosphate form AMP. AMPK is able to sense elevations in AMP/ATP ratio (indicative of energy shortfall) via 3 nucleotide binding sites within its γ regulatory subunit, triggering phosphorylation of AMPK by upstream kinase LKB1 and CaMKK2 and subsequent AMPK signalling. We are interested in examining the molecular mechanisms by which this occurs.

An estimated 380 million people worldwide have type 2 diabetes. The metabolic dimensions of this disease, along with cardiovascular disease, obesity and cancer have encouraged efforts to develop small compound, direct-acting AMPK regulators as novel therapeutics. Crystal structures of AMPK/drug complexes have shown two distinct drug sites exist in the AMPK complex; one at an interface formed between the β-subunit carbohydrate binding module (CBM) and α-subunit kinase domain (occupied by drugs such as A-769662), the other within the γ-subunit (compound C2). Our research aims to understand how drug-binding at these sites leads to regulation of AMPK signalling, thereby driving development of treatments for metabolic diseases.

Student Projects

Staff

Publication Highlights

  1. Morrison KR, Smiles WJ, Ling NXY, Hoque A, Shea G, Ngoei KRW, Yu D, Murray-Segal L, Scott JW, Galic S, Kemp BE, Petersen J & Oakhill JS. An AMPKα2-specific phospho-switch controls lysosomal targeting for activation. Cell Reports, 38:110365 (2022).
  2. Pinkosky SL, Scott JW, Desjardins EM, Smith BK, Day EA, Ford RJ, Langendorf CG, Ling NXY, Nero TL, Loh K, Galic S, Hoque A, Smiles WJ, Ngoei KRW, Parker MW, Yan Y, Melcher K, Kemp BE, Oakhill JS* & Steinberg GR*. Long-chain fatty acyl-CoA esters regulate metabolism via allosteric control of AMPK beta1 isoforms. *co-senior authors. Nature Metabolism, 2:873–881 (2020).
  3. Ling NXY, Kaczmarek A, Hoque A, Davie E, Ngoei KRW, Morrison KR, Smiles WJ, Forte GM, Wang T, Lie S, Dite TA, Langendorf CG, Scott JW, Oakhill JS* & Petersen J*. mTORC1 directly inhibits AMPK to promote cell proliferation under nutrient stress. *co-senior authors. Nature Metabolism, 2:41-49 (2020).
  4. Dite TA, Langendorf CG, Hoque A, Galic S, Rebello RJ, Ovens AJ, Lindqvist LM, Ngoei KRW, Ling NXY, Furic L, Kemp BE, Scott JW & Oakhill JS. AMP-activated protein kinase selectively inhibited by the type II inhibitor SBI-0206965. Journal of Biological Chemistry, 293:8874-8885 (2018).
  5. Ngoei KRW, Langendorf CG, Ling NXY, Hoque A, Varghese S, Camerino MA, Walker SR, Bozikis YE, Dite TA, Ovens AJ, Smiles WJ, Jacobs R, Huang H, Parker MW, Scott JW, Rider MH, Foitzik RC, Kemp BE, Baell JB & Oakhill JS. Structural Determinants for Small-Molecule Activation of Skeletal Muscle AMPK α2β2γ1 by the Glucose Importagog SC4. Cell Chemical Biology, 25:728-737 (2018).
  6. Dite TA, Ling NXY, Scott JW, Hoque A, Galic S, Parker BL, Ngoei KRW, Langendorf CG, O’Brien MT, Kundu M, Viollet B, Steinberg GR, Sakamoto K, Kemp BE, Oakhill JS. The autophagy initiator ULK1 sensitizes AMPK to allosteric drugs. Nature Communications, 8:571 (2017).
  7. Langendorf CG, Ngoei KR, Scott JW, Ling NX, Issa SM, Gorman MA, Parker MW, Sakamoto K, Oakhill JS & Kemp BE. Structural basis of allosteric and synergistic activation of AMPK by furan-2-phosphonic derivative C2 binding. Nature Communications, 7:10912 (2016).
  8. Scott JW, Ling NXY, Issa SMA, Dite TA, O’Brien MT, Chen ZP, Galic S, Langendorf CG, Steinberg GR, Kemp BE & Oakhill JS. Drugs and AMP unite to switch on naive AMPK. Chemistry & Biology, 21:619-627 (2014).
  9. Oakhill JS, Steel R, Chen ZP, Scott JW, Ling N, Tam S & Kemp BE. AMPK is a direct adenylate charge-regulated protein kinase. Science, 332:1433-1435 (2011).
  10. Oakhill JS, Chen ZP, Scott JW, Steel R, Castelli LA, Ling N, Macaulay SL & Kemp BE. β-Subunit myristoylation is the gatekeeper for initiating metabolic stress sensing by AMP-activated protein kinase (AMPK). PNAS, 107:19237-19241 (2010).