AUP1 (Ancient Ubiquitous Protein 1) Is a Key Determinant of Hepatic Very-Low-Density Lipoprotein Assembly and Secretion.
AUP1 (ancient ubiquitous protein 1) is an endoplasmic reticulum-associated protein that also localizes to the surface of lipid droplets (LDs), with dual role in protein quality control and LD regulation. Here, we investigated the role of AUP1 in hepatic lipid mobilization and demonstrate critical roles in intracellular biogenesis of apoB100 (apolipoprotein B-100), LD mobilization, and very-low-density lipoprotein (VLDL) assembly and secretion. APPROACH AND RESULTS: siRNA (short/small interfering RNA) knockdown of AUP1 significantly increased secretion of VLDL-sized apoB100-containing particles from HepG2 cells, correcting a key metabolic defect in these cells that normally do not secrete much VLDL. Secreted particles contained higher levels of metabolically labeled triglyceride, and AUP1-deficient cells displayed a larger average size of LDs, suggesting a role for AUP1 in lipid mobilization. Importantly, AUP1 was also found to directly interact with apoB100, and this interaction was enhanced with proteasomal inhibition. Knockdown of AUP1 reduced apoB100 ubiquitination, decreased intracellular degradation of newly synthesized apoB100, and enhanced extracellular apoB100 secretion. Interestingly, the stimulatory effect of AUP1 knockdown on VLDL assembly was reminiscent of the effect previously observed after MEK-ERK (mitogen-activated protein kinase kinase-extracellular signal-regulated kinase) inhibition; however, further studies indicated that the AUP1 effect was independent of MEK-ERK signaling.



In summary, our findings reveal an important role for AUP1 as a regulator of apoB100 stability, hepatic LD metabolism, and intracellular lipidation of VLDL particles. AUP1 may be a crucial factor in apoB100 quality control, determining the rate at which apoB100 is degraded or lipidated to enable VLDL particle assembly and secretion.
From the Molecular Structure and Function Program, Research Institute, The Hospital for Sick Children, Toronto, Ontario, Canada (J.Z., M.Z., J.T., K.A.); Department of Biochemistry (M.Z., K.A.) and Department of Laboratory Medicine and Pathobiology (J.T., K.A.), University of Toronto, Ontario, Canada; Biochemistry and Cell Biology of Lipids Unit, LIMES Institute, University of Bonn, Germany (C.T.); and Department of Biochemistry, Microbiology, and Immunology, University of Ottawa, Ontario, Canada (M.A.A., Z.Y.).
Jing
J
Zhang
From the Molecular Structure and Function Program, Research Institute, The Hospital for Sick Children, Toronto, Ontario, Canada (J.Z., M.Z., J.T., K.A.); Department of Biochemistry (M.Z., K.A.) and Department of Laboratory Medicine and Pathobiology (J.T., K.A.), University of Toronto, Ontario, Canada; Biochemistry and Cell Biology of Lipids Unit, LIMES Institute, University of Bonn, Germany (C.T.); and Department of Biochemistry, Microbiology, and Immunology, University of Ottawa, Ontario, Canada (M.A.A., Z.Y.).
Mostafa
M
Zamani
From the Molecular Structure and Function Program, Research Institute, The Hospital for Sick Children, Toronto, Ontario, Canada (J.Z., M.Z., J.T., K.A.); Department of Biochemistry (M.Z., K.A.) and Department of Laboratory Medicine and Pathobiology (J.T., K.A.), University of Toronto, Ontario, Canada; Biochemistry and Cell Biology of Lipids Unit, LIMES Institute, University of Bonn, Germany (C.T.); and Department of Biochemistry, Microbiology, and Immunology, University of Ottawa, Ontario, Canada (M.A.A., Z.Y.).
Christoph
C
Thiele
From the Molecular Structure and Function Program, Research Institute, The Hospital for Sick Children, Toronto, Ontario, Canada (J.Z., M.Z., J.T., K.A.); Department of Biochemistry (M.Z., K.A.) and Department of Laboratory Medicine and Pathobiology (J.T., K.A.), University of Toronto, Ontario, Canada; Biochemistry and Cell Biology of Lipids Unit, LIMES Institute, University of Bonn, Germany (C.T.); and Department of Biochemistry, Microbiology, and Immunology, University of Ottawa, Ontario, Canada (M.A.A., Z.Y.).
Jennifer
J
Taher
From the Molecular Structure and Function Program, Research Institute, The Hospital for Sick Children, Toronto, Ontario, Canada (J.Z., M.Z., J.T., K.A.); Department of Biochemistry (M.Z., K.A.) and Department of Laboratory Medicine and Pathobiology (J.T., K.A.), University of Toronto, Ontario, Canada; Biochemistry and Cell Biology of Lipids Unit, LIMES Institute, University of Bonn, Germany (C.T.); and Department of Biochemistry, Microbiology, and Immunology, University of Ottawa, Ontario, Canada (M.A.A., Z.Y.).
Mohsen
M
Amir Alipour
From the Molecular Structure and Function Program, Research Institute, The Hospital for Sick Children, Toronto, Ontario, Canada (J.Z., M.Z., J.T., K.A.); Department of Biochemistry (M.Z., K.A.) and Department of Laboratory Medicine and Pathobiology (J.T., K.A.), University of Toronto, Ontario, Canada; Biochemistry and Cell Biology of Lipids Unit, LIMES Institute, University of Bonn, Germany (C.T.); and Department of Biochemistry, Microbiology, and Immunology, University of Ottawa, Ontario, Canada (M.A.A., Z.Y.).
Zemin
Z
Yao
From the Molecular Structure and Function Program, Research Institute, The Hospital for Sick Children, Toronto, Ontario, Canada (J.Z., M.Z., J.T., K.A.); Department of Biochemistry (M.Z., K.A.) and Department of Laboratory Medicine and Pathobiology (J.T., K.A.), University of Toronto, Ontario, Canada; Biochemistry and Cell Biology of Lipids Unit, LIMES Institute, University of Bonn, Germany (C.T.); and Department of Biochemistry, Microbiology, and Immunology, University of Ottawa, Ontario, Canada (M.A.A., Z.Y.). khosrow.adeli@sickkids.ca.
Khosrow
K
Adeli
D053299:Apolipoprotein B-100 / Q000378:metabolism*; D051017:Apoptosis Regulatory Proteins / Q000378:metabolism; D002352:Carrier Proteins / Q000235:genetics / Q000378:metabolism*; D056945:Hep G2 Cells; D022781:Hepatocytes / Q000378:metabolism*; D006801:Humans; D066292:Lipid Droplets / Q000378:metabolism; D008079:Lipoproteins, VLDL / Q000378:metabolism*; D008099:Liver / Q000378:metabolism*; D008565:Membrane Proteins; D010316:Particle Size; D046988:Proteasome Endopeptidase Complex / Q000378:metabolism; D011485:Protein Binding; D055550:Protein Stability; D059748:Proteolysis; D034622:RNA Interference; D014162:Transfection; D014280:Triglycerides / Q000378:metabolism*; D054875:Ubiquitination
C464942:AUP1 protein, human; D053299:Apolipoprotein B-100; D051017:Apoptosis Regulatory Proteins; C112511:CIDEB protein, human; D002352:Carrier Proteins; D008079:Lipoproteins, VLDL; D008565:Membrane Proteins; D014280:Triglycerides; C030106:very low density lipoprotein triglyceride; D046988:Proteasome Endopeptidase Complex
AUP1; VLDL; apobB100; hepatocytes; lipid droplet