title: Regulation of amino acid transporter trafficking by mTORC1 in primary human trophoblast cells is mediated by the ubiquitin ligase Nedd4-2.
abstract: Changes in placental amino acid transfer directly contribute to altered fetal growth, which increases the risk for perinatal complications and predisposes for the development of obesity, diabetes and cardiovascular disease later in life. Placental amino acid transfer is critically dependent on the expression of specific transporters in the plasma membrane of the trophoblast, the transporting epithelium of the human placenta. However, the molecular mechanisms regulating this process are largely unknown. Nedd4-2 is an ubiquitin ligase that catalyses the ubiquitination of proteins, resulting in proteasomal degradation. We hypothesized that inhibition of mechanistic target of rapamycin complex 1 (mTORC1) decreases amino acid uptake in primary human trophoblast (PHT) cells by activation of Nedd4-2, which increases transporter ubiquitination resulting in decreased transporter expression in the plasma membrane. mTORC 1 inhibition increased the expression of Nedd4-2, promoted ubiquitination and decreased the plasma membrane expression of SNAT2 (an isoform of the System A amino acid transporter) and LAT1 (a System L amino acid transporter isoform), resulting in decreased cellular amino acid uptake. Nedd4-2 silencing markedly increased the trafficking of SNAT2 and LAT1 to the plasma membrane, which stimulated cellular amino acid uptake. mTORC1 inhibition by silencing of raptor failed to decrease amino acid transport following Nedd4-2 silencing. In conclusion, we have identified a novel link between mTORC1 signalling and ubiquitination, a common posttranslational modification. Because placental mTORC1 is inhibited in fetal growth restriction and activated in fetal overgrowth, we propose that regulation of placental amino acid transporter ubiquitination by mTORC1 and Nedd4-2 constitutes a molecular mechanisms underlying abnormal fetal growth.
authors.affiliation: Division of Reproductive Sciences, Department of OB/GYN, University of Colorado Anschutz Medical Campus, Aurora, CO 80045, U.S.A. fredrick.joseph@ucdenver.edu.
authors.forename: Fredrick J
authors.initials: FJ
authors.lastname: Rosario
authors.affiliation: Department of Medicine, Peter Doherty Institute, The University of Melbourne, Carlton, Victoria 3010, Australia.
authors.forename: Kris Genelyn
authors.initials: KG
authors.lastname: Dimasuay
authors.affiliation: Division of Bio-System Pharmacology, Department of Pharmacology, Graduate School of Medicine, Osaka University, Osaka, 565-0871, Japan.
authors.forename: Yoshikatsu
authors.initials: Y
authors.lastname: Kanai
authors.affiliation: Section of Neonatology, Department of Pediatrics, University of Colorado Anschutz Medical Campus, Aurora, CO 80045, U.S.A.
authors.forename: Theresa L
authors.initials: TL
authors.lastname: Powell
authors.affiliation: Division of Reproductive Sciences, Department of OB/GYN, University of Colorado Anschutz Medical Campus, Aurora, CO 80045, U.S.A.
authors.forename: Thomas
authors.initials: T
authors.lastname: Jansson
mesh_terms: D048868:Adaptor Proteins, Signal Transducing / Q000235:genetics / Q000378:metabolism*; D026924:Amino Acid Transport System A / Q000378:metabolism*; D027241:Amino Acid Transport System y+L; D002478:Cells, Cultured; D056827:Endosomal Sorting Complexes Required for Transport / Q000235:genetics / Q000378:metabolism*; D005260:Female; D027301:Fusion Regulatory Protein 1, Light Chains / Q000378:metabolism*; D006801:Humans; D047908:Intracellular Signaling Peptides and Proteins / Q000235:genetics / Q000378:metabolism*; D000076222:Mechanistic Target of Rapamycin Complex 1; D000076225:Mechanistic Target of Rapamycin Complex 2; D046912:Multiprotein Complexes / Q000235:genetics / Q000378:metabolism*; D000075702:Nedd4 Ubiquitin Protein Ligases; D011247:Pregnancy; D061251:Primary Cell Culture; D021381:Protein Transport; D034622:RNA Interference; D000076223:Regulatory-Associated Protein of mTOR; D015398:Signal Transduction; D058570:TOR Serine-Threonine Kinases / Q000235:genetics / Q000378:metabolism*; D014162:Transfection; D014327:Trophoblasts / Q000201:enzymology*; D044767:Ubiquitin-Protein Ligases / Q000235:genetics / Q000378:metabolism*; D054875:Ubiquitination
chemical_list: D048868:Adaptor Proteins, Signal Transducing; D026924:Amino Acid Transport System A; D027241:Amino Acid Transport System y+L; D056827:Endosomal Sorting Complexes Required for Transport; D027301:Fusion Regulatory Protein 1, Light Chains; D047908:Intracellular Signaling Peptides and Proteins; D046912:Multiprotein Complexes; C463659:RPTOR protein, human; D000076223:Regulatory-Associated Protein of mTOR; C445761:SLC38A1 protein, human; C421261:SLC7A7 protein, human; D000075702:Nedd4 Ubiquitin Protein Ligases; C000618483:Nedd4 protein, human; C000618485:Nedd4L protein, human; D044767:Ubiquitin-Protein Ligases; C556139:DEPTOR protein, human; D000076222:Mechanistic Target of Rapamycin Complex 1; D000076225:Mechanistic Target of Rapamycin Complex 2; D058570:TOR Serine-Threonine Kinases
keywords: LAT1; SNAT2; placenta; ubiquitination