@article{bb63a22b26b44dfab2ad726502f1f6cc,
title = " Blockade of MCU-Mediated Ca 2+ Uptake Perturbs Lipid Metabolism via PP4-Dependent AMPK Dephosphorylation ",
abstract = " Mitochondrial Ca 2+ uniporter (MCU)-mediated Ca 2+ uptake promotes the buildup of reducing equivalents that fuel oxidative phosphorylation for cellular metabolism. Although MCU modulates mitochondrial bioenergetics, its function in energy homeostasis in vivo remains elusive. Here we demonstrate that deletion of the Mcu gene in mouse liver (MCU Δhep ) and in Danio rerio by CRISPR/Cas9 inhibits mitochondrial Ca 2+ ( m Ca 2+ ) uptake, delays cytosolic Ca 2+ ( c Ca 2+ ) clearance, reduces oxidative phosphorylation, and leads to increased lipid accumulation. Elevated hepatic lipids in MCU Δhep were a direct result of extramitochondrial Ca 2+ -dependent protein phosphatase-4 (PP4) activity, which dephosphorylates AMPK. Loss of AMPK recapitulates hepatic lipid accumulation without changes in MCU-mediated Ca 2+ uptake. Furthermore, reconstitution of active AMPK, or PP4 knockdown, enhances lipid clearance in MCU Δhep hepatocytes. Conversely, gain-of-function MCU promotes rapid m Ca 2+ uptake, decreases PP4 levels, and reduces hepatic lipid accumulation. Thus, our work uncovers an MCU/PP4/AMPK molecular cascade that links Ca 2+ dynamics to hepatic lipid metabolism. Hepatic mitochondrial Ca 2+ shapes bioenergetics and lipid homeostasis. Tomar et al. demonstrate that MCU-mediated c Ca 2+ buffering serves as a crucial step in controlling hepatic fuel metabolism through an MCU/PP4/AMPK molecular cascade. Identification of these molecular signaling events aids in understanding how perturbation of mitochondrial ion homeostasis may contribute to the etiology of metabolic disorders.",
keywords = "AMPK, MCU, bioenergetics, calcium, diabetes, hepatocyte, lipid metabolism, metabolic diseases, mitochondrial Ca uniporter, phosphatase",
author = "Dhanendra Tomar and Fabi{\'a}n Ja{\~n}a and Zhiwei Dong and Quinn, {William J.} and Pooja Jadiya and Breves, {Sarah L.} and Daw, {Cassidy C.} and Subramanya Srikantan and Santhanam Shanmughapriya and Neeharika Nemani and Edmund Carvalho and Aparna Tripathi and Worth, {Alison M.} and Xueqian Zhang and Roshanak Razmpour and Ajay Seelam and Stephen Rhode and Mehta, {Anuj V.} and Michael Murray and Daniel Slade and Ramirez, {Servio H.} and Prashant Mishra and Gerhard, {Glenn S.} and Jeffrey Caplan and Luke Norton and Kumar Sharma and Sudarsan Rajan and Darius Balciunas and Wijesinghe, {Dayanjan S.} and Ahima, {Rexford S.} and Baur, {Joseph A.} and Muniswamy Madesh",
note = "Funding Information: We thank Anne Brunet for sharing the AMPK-HA construct. The AMPKα1 T172D-expressing plasmid was provided by Dr. Kenneth R. Hallows (University of Southern California Keck School of Medicine, Los Angeles, CA, USA). We thank Dr. John W. Elrod for sharing the MCU fl/fl mice. We thank Dr. Mitchell A. Lazar and Dr. Kevin J. Foskett (University of Pennsylvania, Philadelphia, PA, USA) for critically reading the manuscript, helpful thoughts, and suggestions. We also thank Jean L. Ross and Shannon Modla for EM sample preparation and image acquisition. This research was funded by the NIH ( R01GM109882 , R01HL086699 , R01HL142673 , and 1S10RR027327 to M. Madesh and U01HD087198 to D.S.W.). D.T. is supported by American Heart Association postdoctoral fellowship grant 17POST33660251 . This work also received support via a Young Investigator Award from SCIEX for clinical lipidomic research (to D.S.W.). Services and products in support of the research project were generated by the VCU Massey Cancer Center Lipidomics Shared Resource (Developing Core) supported in part NIH-NCI Cancer Center support grant P30 CA016059 . Metabolic profiling was performed by the University of Pennsylvania Diabetes Research Center Mouse Phenotyping, Physiology and Metabolism Core ( NIH grant P30-DK19525 ). Funding Information: We thank Anne Brunet for sharing the AMPK-HA construct. The AMPKα1 T172D-expressing plasmid was provided by Dr. Kenneth R. Hallows (University of Southern California Keck School of Medicine, Los Angeles, CA, USA). We thank Dr. John W. Elrod for sharing the MCUfl/fl mice. We thank Dr. Mitchell A. Lazar and Dr. Kevin J. Foskett (University of Pennsylvania, Philadelphia, PA, USA) for critically reading the manuscript, helpful thoughts, and suggestions. We also thank Jean L. Ross and Shannon Modla for EM sample preparation and image acquisition. This research was funded by the NIH (R01GM109882, R01HL086699, R01HL142673, and 1S10RR027327 to M. Madesh and U01HD087198 to D.S.W.). D.T. is supported by American Heart Association postdoctoral fellowship grant 17POST33660251. This work also received support via a Young Investigator Award from SCIEX for clinical lipidomic research (to D.S.W.). Services and products in support of the research project were generated by the VCU Massey Cancer Center Lipidomics Shared Resource (Developing Core) supported in part NIH-NCI Cancer Center support grant P30 CA016059. Metabolic profiling was performed by the University of Pennsylvania Diabetes Research Center Mouse Phenotyping, Physiology and Metabolism Core (NIH grant P30-DK19525). Funding Information: We thank Anne Brunet for sharing the AMPK-HA construct. The AMPK?1 T172D-expressing plasmid was provided by Dr. Kenneth R. Hallows (University of Southern California Keck School of Medicine, Los Angeles, CA, USA). We thank Dr. John W. Elrod for sharing the MCUfl/fl mice. We thank Dr. Mitchell A. Lazar and Dr. Kevin J. Foskett (University of Pennsylvania, Philadelphia, PA, USA) for critically reading the manuscript, helpful thoughts, and suggestions. We also thank Jean L. Ross and Shannon Modla for EM sample preparation and image acquisition. This research was funded by the NIH (R01GM109882, R01HL086699, R01HL142673, and 1S10RR027327 to M. Madesh and U01HD087198 to D.S.W.). D.T. is supported by American Heart Association postdoctoral fellowship grant 17POST33660251. This work also received support via a Young Investigator Award from SCIEX for clinical lipidomic research (to D.S.W.). Services and products in support of the research project were generated by the VCU Massey Cancer Center Lipidomics Shared Resource (Developing Core) supported in part NIH-NCI Cancer Center support grant P30 CA016059. Metabolic profiling was performed by the University of Pennsylvania Diabetes Research Center Mouse Phenotyping, Physiology and Metabolism Core (NIH grant P30-DK19525). D.T. P.J. F.J. C.C.D. and S. Srikantan performed hepatocyte isolation and culture, mCa2+ and cCa2+ measurements, confocal imaging, biochemical assays, and mitochondrial respiration experiments. Z.D. isolated mitoplasts, and X.Z. performed patch-clamping. S.L.B. A.T. N.N. S. Shanmughapriya, A.M.W. M. Murray, and D.S. contributed reagents and experimental tools and performed mouse genotyping. D.T. R.R. A.S. S. Rhode, and S.H.R. performed liver histology and imaging. D.T. S. Rajan, and M. Madesh designed and established the mouse model. S. Rajan, A.V.M. D.B. and M. Madesh designed and developed the zebrafish model. D.T. and N.N. performed zebrafish confocal imaging. J.C. performed electron microscopy. D.S.W. performed liver lipidomics profiling. W.J.Q. and J.A.B. generated AMPK?1/?2?hep mice. W.J.Q. J.A.B. L.N. K.S. and R.S.A. performed liver and plasma biochemistry and the in vivo CLAM study. S. Rajan, D.T. and S.L.B. performed molecular experiments, and guidance was provided by G.S.G. P.M. J.A.B. R.S.A. and M. Madesh. D.T. and M. Madesh conceived and designed the experiments and interpreted the experimental data. D.T. S.L.B. and M. Madesh wrote the manuscript with contributions from all the authors. The authors declare no competing interests. Publisher Copyright: {\textcopyright} 2019 The Authors",
year = "2019",
month = mar,
day = "26",
doi = "10.1016/j.celrep.2019.02.107",
language = "English (US)",
volume = "26",
pages = "3709--3725.e7",
journal = "Cell Reports",
issn = "2211-1247",
publisher = "Cell Press",
number = "13",
}