title: The Lhs1/GRP170 chaperones facilitate the endoplasmic reticulum-associated degradation of the epithelial sodium channel.
abstract: The epithelial sodium channel, ENaC, plays a critical role in maintaining salt and water homeostasis, and not surprisingly defects in ENaC function are associated with disease. Like many other membrane-spanning proteins, this trimeric protein complex folds and assembles inefficiently in the endoplasmic reticulum (ER), which results in a substantial percentage of the channel being targeted for ER-associated degradation (ERAD). Because the spectrum of factors that facilitates the degradation of ENaC is incomplete, we developed yeast expression systems for each ENaC subunit. We discovered that a conserved Hsp70-like chaperone, Lhs1, is required for maximal turnover of the ENaC α subunit. By expressing Lhs1 ATP binding mutants, we also found that the nucleotide exchange properties of this chaperone are dispensable for ENaC degradation. Consistent with the precipitation of an Lhs1-αENaC complex, Lhs1 holdase activity was instead most likely required to support the ERAD of αENaC. Moreover, a complex containing the mammalian Lhs1 homolog GRP170 and αENaC co-precipitated, and GRP170 also facilitated ENaC degradation in human, HEK293 cells, and in a Xenopus oocyte expression system. In both yeast and higher cell types, the effect of Lhs1 on the ERAD of αENaC was selective for the unglycosylated form of the protein. These data establish the first evidence that Lhs1/Grp170 chaperones can act as mediators of ERAD substrate selection.
authors: Department of Biological Sciences, University of Pittsburgh, Pittsburgh, Pennsylvania 15260, USA.
authors: Teresa M
authors: TM
authors: Buck
authors: Lindsay
authors: L
authors: Plavchak
authors: Ankita
authors: A
authors: Roy
authors: Bridget F
authors: BF
authors: Donnelly
authors: Ossama B
authors: OB
authors: Kashlan
authors: Thomas R
authors: TR
authors: Kleyman
authors: Arohan R
authors: AR
authors: Subramanya
authors: Jeffrey L
authors: JL
authors: Brodsky
mesh_terms: D000255:Adenosine Triphosphate / Q000378:metabolism; D000584:Amiloride / Q000494:pharmacology; D000818:Animals; D004721:Endoplasmic Reticulum / Q000378:metabolism; D060746:Endoplasmic Reticulum-Associated Degradation; D053503:Epithelial Sodium Channels / Q000235:genetics / Q000378:metabolism*; D005260:Female; D006023:Glycoproteins / Q000235:genetics / Q000378:metabolism*; D057809:HEK293 Cells; D018840:HSP70 Heat-Shock Proteins / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D015151:Immunoblotting; D017136:Ion Transport / Q000187:drug effects; D008564:Membrane Potentials / Q000187:drug effects; D009154:Mutation; D009865:Oocytes / Q000378:metabolism / Q000502:physiology; D011485:Protein Binding; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics / Q000378:metabolism*; D012964:Sodium / Q000378:metabolism; D014981:Xenopus
chemical_list: D053503:Epithelial Sodium Channels; D006023:Glycoproteins; D018840:HSP70 Heat-Shock Proteins; C102115:LHS1 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins; C098410:glucose-regulated protein 170; D000584:Amiloride; D000255:Adenosine Triphosphate; D012964:Sodium
keywords: ENaC; ER-associated Degradation; Endoplasmic Reticulum (ER); Molecular Chaperone; Proteasome; Sodium Channels
references: J Biol Chem. 2010 Oct 1;285(40):30363-9
references: 20624922
references: Semin Cell Dev Biol. 2007 Dec;18(6):751-61
references: 17964199
references: Proc Natl Acad Sci U S A. 1995 Jul 3;92(14):6224-8
references: 7603976
references: Mol Biol Cell. 2001 Oct;12(10):3226-41
references: 11598205
references: Biochemistry. 1996 Jun 11;35(23):7299-307
references: 8652506
references: EMBO J. 2000 Dec 1;19(23):6440-52
references: 11101517
references: Nature. 2001 Dec 6;414(6864):652-6
references: 11740563
references: Cell. 2008 Jun 13;133(6):1068-79
references: 18555782
references: J Biol Chem. 2007 Sep 7;282(36):26140-9
references: 17631501
references: Yeast. 1994 Apr;10 Suppl A:S69-74
references: 8091863
references: J Biol Chem. 2001 Apr 20;276(16):12903-10
references: 11278712
references: Nat Cell Biol. 2005 Aug;7(8):766-72
references: 16056268
references: Mol Cell Endocrinol. 2012 Mar 24;350(2):242-7
references: 22101317
references: Cell. 2000 Apr 28;101(3):249-58
references: 10847680
references: J Biol Chem. 2010 Jul 9;285(28):21600-6
references: 20430899
references: J Biol Chem. 1996 May 31;271(22):13215-20
references: 8662752
references: Am J Physiol. 1994 Dec;267(6 Pt 1):C1682-90
references: 7810611
references: J Biol Chem. 1996 Sep 13;271(37):22791-5
references: 8798455
references: J Biol Chem. 2010 Apr 16;285(16):12445-53
references: 20177057
references: EMBO J. 1996 Jun 3;15(11):2640-50
references: 8654361
references: Mol Cell. 2005 Sep 16;19(6):741-51
references: 16168370
references: J Cell Biol. 2012 Dec 10;199(6):907-18
references: 23229898
references: Mol Biol Cell. 2010 Mar 15;21(6):1047-58
references: 20110346
references: Mol Biol Cell. 2004 Nov;15(11):4787-97
references: 15342786
references: Physiol Rev. 2012 Apr;92(2):537-76
references: 22535891
references: Biochem J. 2011 Sep 15;438(3):447-55
references: 21675960
references: Cell Mol Life Sci. 2006 Nov;63(22):2560-70
references: 16952052
references: Nature. 1997 Aug 28;388(6645):891-5
references: 9278052
references: Nat Rev Mol Cell Biol. 2010 Aug;11(8):579-92
references: 20651708
references: J Biol Chem. 2012 Jun 1;287(23):19255-65
references: 22496374
references: Biochim Biophys Acta. 1999 Jul 13;1432(2):214-21
references: 10407143
references: Mol Biol Cell. 2012 Feb;23(4):520-32
references: 22190736
references: Mol Cell. 2005 Sep 16;19(6):765-75
references: 16168372
references: Mol Biol Cell. 2005 Jan;16(1):40-50
references: 15525676
references: Cell. 2006 Jul 28;126(2):237-9
references: 16873052
references: Cell Stress Chaperones. 2000 Oct;5(4):276-90
references: 11048651
references: Am J Physiol. 1996 Aug;271(2 Pt 1):C605-11
references: 8770001
references: EMBO J. 2000 Feb 15;19(4):550-61
references: 10675324
references: Mol Biol Cell. 1992 Feb;3(2):129-42
references: 1550957
references: Am J Physiol Gastrointest Liver Physiol. 2005 Sep;289(3):G444-55
references: 15845869
references: Science. 2011 Nov 25;334(6059):1086-90
references: 22116878
references: J Biol Chem. 1997 Oct 24;272(43):27295-300
references: 9341177
references: Curr Opin Cell Biol. 2011 Apr;23(2):157-66
references: 21094034
references: J Cell Biol. 1997 May 19;137(4):813-24
references: 9151684
references: Am J Physiol. 1999 Jun;276(6):C1346-51
references: 10362597
references: Gene. 1995 Apr 14;156(1):119-22
references: 7737504
references: Mol Biol Cell. 2007 Mar;18(3):806-14
references: 17182856
references: Trends Mol Med. 2012 Feb;18(2):81-91
references: 22138491
references: Cell. 2009 Jan 23;136(2):272-83
references: 19167329
references: FEBS Lett. 2002 Nov 6;531(2):339-42
references: 12417338
references: Cell. 2006 Jul 28;126(2):349-59
references: 16873065
references: EMBO J. 1997 Mar 3;16(5):899-907
references: 9118951
references: J Cell Biol. 1995 Jul;130(1):41-9
references: 7790376
references: Gene. 1989 Apr 15;77(1):51-9
references: 2744487
references: Mol Biol Cell. 2001 May;12(5):1303-14
references: 11359923
references: Cell. 2007 Oct 5;131(1):106-20
references: 17923091
references: Protein Sci. 2005 Jul;14(7):1697-709
references: 15987899
references: J Cell Biol. 2009 Jan 12;184(1):159-72
references: 19124653
references: J Biol Chem. 1998 Jan 9;273(2):681-4
references: 9422716
references: J Biol Chem. 2000 Dec 22;275(51):39886-93
references: 10978318
references: Methods Mol Med. 2002;70:257-65
references: 11917528
references: Proc Natl Acad Sci U S A. 1996 Nov 26;93(24):13797-801
references: 8943015
references: J Biol Chem. 2003 Sep 26;278(39):37073-82
references: 12871941
references: Semin Cell Dev Biol. 2007 Dec;18(6):780-91
references: 17942349
references: Mol Biol Cell. 2001 Dec;12(12):4114-28
references: 11739805
references: J Biol Chem. 2001 Jul 6;276(27):24891-900
references: 11333259
references: Nat Rev Mol Cell Biol. 2008 Dec;9(12):944-57
references: 19002207
references: J Biol Chem. 2007 Sep 21;282(38):28149-56
references: 17664274
references: J Biol Chem. 2007 Nov 16;282(46):33475-33483
references: 17855354
references: J Clin Invest. 1999 Oct;104(7):R19-23
references: 10510339
references: Mol Cell. 2008 Dec 26;32(6):870-7
references: 19111666
references: Cell. 2012 Dec 7;151(6):1163-7
references: 23217703
references: EMBO J. 2008 Jan 9;27(1):201-11
references: 18046456
references: Trends Biochem Sci. 2010 Feb;35(2):74-82
references: 19853458
references: J Biol Chem. 1999 Feb 5;274(6):3453-60
references: 9920890
references: Endocrinology. 2005 Dec;146(12):5079-85
references: 16150899
references: J Biol Chem. 2010 Oct 1;285(40):30453-62
references: 20663869
references: J Cell Biol. 2003 May 26;161(4):715-25
references: 12756234
references: Cell Stress Chaperones. 2007 Spring;12(1):1-8
references: 17441502
references: Cancer Res. 2006 Jan 15;66(2):1161-8
references: 16424054
references: Proc Natl Acad Sci U S A. 2006 Apr 11;103(15):5817-22
references: 16585520
references: Mol Cell Biol. 2002 Jan;22(2):626-34
references: 11756557
references: J Biol Chem. 2009 Nov 13;284(46):31564-71
references: 19759005
references: Mol Cell. 2005 Sep 16;19(6):753-64
references: 16168371
references: Genetics. 2012 Nov;192(3):889-903
references: 22923381
references: Exp Cell Res. 2012 May 15;318(9):1011-9
references: 22405998
references: EMBO J. 1998 Dec 1;17(23):6879-87
references: 9843494
references: J Biol Chem. 1994 Sep 30;269(39):24379-83
references: 7929098
references: Mol Biol Cell. 2007 Oct;18(10):3776-87
references: 17634286
references: J Biol Chem. 1995 Mar 3;270(9):4563-9
references: 7876226
references: Trends Cell Biol. 2012 Jan;22(1):22-32
references: 22055166
references: Curr Top Microbiol Immunol. 2005;300:41-56
references: 16573236
references: Annu Rev Biochem. 2004;73:1019-49
references: 15189166
references: J Biol Chem. 2002 May 3;277(18):15947-56
references: 11836248
references: Mol Biol Cell. 1993 Nov;4(11):1109-19
references: 8305733
references: J Am Soc Nephrol. 2008 Oct;19(10):1845-54
references: 18753254
references: FEBS Lett. 2006 Oct 2;580(22):5237-40
references: 16962589
references: Nature. 1994 Feb 3;367(6462):463-7
references: 8107805
references: EMBO J. 1997 Nov 3;16(21):6325-36
references: 9351815
references: Mol Biol Cell. 2002 Dec;13(12):4456-69
references: 12475965
references: J Biol Chem. 2010 Oct 8;285(41):31261-7
references: 20667832
references: Science. 2004 Jan 2;303(5654):98-101
references: 14704430
references: J Biol Chem. 2012 Jun 22;287(26):21914-25
references: 22474283
references: Mol Biol Cell. 2008 Jun;19(6):2620-30
references: 18400946
references: Cell. 2008 Jan 11;132(1):101-12
references: 18191224
references: J Biol Chem. 2011 Dec 23;286(51):43611-43621
references: 22027832
references: Trends Biochem Sci. 2012 Oct;37(10):404-10
references: 22921611
references: Mol Biol Cell. 2004 Sep;15(9):4125-35
references: 15215312
references: J Cell Biol. 2001 May 28;153(5):1061-70
references: 11381090
references: Biochemistry. 2003 Dec 23;42(50):14893-902
references: 14674765
references: Mol Cell Biol. 1996 Nov;16(11):6444-56
references: 8887673
references: J Biol Chem. 1998 Nov 13;273(46):30344-51
references: 9804797
references: EMBO J. 1996 May 15;15(10):2371-80
references: 8665844
references: J Biol Chem. 2007 Nov 9;282(45):32665-75
references: 17823116
references: EMBO J. 1999 Mar 15;18(6):1492-505
references: 10075921