Sec61β facilitates the maintenance of endoplasmic reticulum homeostasis by associating microtubules

Abstract

Sec61β, a subunit of the Sec61 translocon complex, is not essential in yeast and commonly used as a marker of endoplasmic reticulum (ER). In higher eukaryotes, such as Drosophila, deletion of Sec61β causes lethality, but its physiological role is unclear. Here, we show that Sec61β interacts directly with microtubules. Overexpression of Sec61β containing small epitope tags, but not a RFP tag, induces dramatic bundling of the ER and microtubule. A basic region in the cytosolic domain of Sec61β is critical for microtubule association. Depletion of Sec61β induces ER stress in both mammalian cells and Caenorhabditis elegans, and subsequent restoration of ER homeostasis correlates with the microtubule binding ability of Sec61β. Loss of Sec61β causes increased mobility of translocon complexes and reduced level of membrane-bound ribosomes. These results suggest that Sec61β may stabilize protein translocation by linking translocon complex to microtubule and provide insight into the physiological function of ER-microtubule interaction.

Keywords

ER stress / Microtubule / Sec61β / Translocon / Ribosome

References

[1] Abell BM, Pool MR, Schlenker O, Sinning I, High S (2004) Signal recognition particle mediates post-translational targeting in eukaryotes. EMBO J 23:2755–2764

[2] Abell BM, Rabu C, Leznicki P, Young JC, High S (2007) Posttranslational integration of tail-anchored proteins is facilitated by defined molecular chaperones. J Cell Sci 120:1743–1751

[3] Barlowe C (2010) ER sheets get roughed up. Cell 143:665–666

[4] Bernales S, Papa FR, Walter P (2006) Intracellular signaling by the unfolded protein response. Annu Rev Cell Dev Biol 22:487–508

[5] Brenner S (1974) The genetics of Caenorhabditis elegans. Genetics 77:71–94

[6] Calfon M, Zeng H, Urano F, Till JH, Hubbard SR, Harding HP, Clark SG, Ron D (2002) IRE1 couples endoplasmic reticulum load to secretory capacity by processing the XBP-1 mRNA. Nature 415:92–96

[7] Carrasco S, Meyer T (2011) STIM proteins and the endoplasmic reticulum-plasma membrane junctions. Annu Rev Biochem 80:973–1000

[8] Favaloro V, Spasic M, Schwappach B, Dobberstein B (2008) Distinct targeting pathways for the membrane insertion of tail-anchored (TA) proteins. J Cell Sci 121:1832–1840

[9] Feng D, Zhao X, Soromani C, Toikkanen J, Romisch K, Vembar SS, Brodsky JL, Keranen S, Jantti J (2007) The transmembrane domain is sufficient for Sbh1p function, its association with the Sec61 complex, and interaction with Rtn1p. J Biol Chem 282:30618–30628

[10] Finke K, Plath K, Panzner S, Prehn S, Rapoport TA, Hartmann E, Sommer T (1996) A second trimeric complex containing homologs of the Sec61p complex functions in protein transport across the ER membrane of S. cerevisiae. EMBO J 15:1482–1494

[11] Friedman JR, Voeltz GK (2011) The ER in 3D: a multifunctional dynamic membrane network. Trends Cell Biol 21:709–717

[12] Gamerdinger M, Hanebuth MA, Frickey T, Deuerling E (2015) The principle of antagonism ensures protein targeting specificity at the endoplasmic reticulum. Science 348:201–207

[13] Goyal U, Blackstone C (2013) Untangling the web: mechanisms underlying ER network formation. Biochim Biophys Acta 1833:2492–2498

[14] Grigoriev I, Gouveia SM, van der Vaart B, Demmers J, Smyth JT, Honnappa S, Splinter D, Steinmetz MO, Putney JW Jr, Hoogenraad CC, Akhmanova A (2008) STIM1 is a MT-plus-end-tracking protein involved in remodeling of the ER. Curr Biol 18:177–182

[15] Harding HP, Zhang Y, Ron D (1999) Protein translation and folding are coupled by an endoplasmic-reticulum-resident kinase. Nature 397:271–274

[16] Harding HP, Zhang Y, Bertolotti A, Zeng H, Ron D (2000) Perk is essential for translational regulation and cell survival during the unfolded protein response. Mol Cell 5:897–904

[17] Haze K, Yoshida H, Yanagi H, Yura T, Mori K (1999) Mammalian transcription factor ATF6 is synthesized as a transmembrane protein and activated by proteolysis in response to endoplasmic reticulum stress. Mol Biol Cell 10:3787–3799

[18] Hu J, Shibata Y, Voss C, Shemesh T, Li Z, Coughlin M, Kozlov MM, Rapoport TA, Prinz WA (2008) Membrane proteins of the endoplasmic reticulum induce high-curvature tubules. Science 319:1247–1250

[19] Hu J, Prinz WA, Rapoport TA (2011) Weaving the web of ER tubules. Cell 147:1226–1231

[20] Kalies KU, Rapoport TA, Hartmann E (1998) The beta subunit of the Sec61 complex facilitates cotranslational protein transport and interacts with the signal peptidase during translocation. J Cell Biol 141:887–894

[21] Kelkar A, Dobberstein B (2009) Sec61beta, a subunit of the Sec61 protein translocation channel at the endoplasmic reticulum, is involved in the transport of Gurken to the plasma membrane. BMC Cell Biol 10:11

[22] Klopfenstein DR, Kappeler F, Hauri HP (1998) A novel direct interaction of endoplasmic reticulum with microtubules. EMBO J 17:6168–6177

[23] Lee K, Tirasophon W, Shen X, Michalak M, Prywes R, Okada T, Yoshida H, Mori K, Kaufman RJ (2002) IRE1-mediated unconventional mRNA splicing and S2P-mediated ATF6 cleavage merge to regulate XBP1 in signaling the unfolded protein response. Genes Dev 16:452–466

[24] Leroux A, Rokeach LA (2008) Inter-species complementation of the translocon beta subunit requires only its transmembrane domain. PLoS ONE 3:e3880

[25] Levy R, Wiedmann M, Kreibich G (2001) In vitro binding of ribosomes to the beta subunit of the Sec61p protein translocation complex. J Biol Chem 276:2340–2346

[26] Li H, Korennykh AV, Behrman SL, Walter P (2010) Mammalian endoplasmic reticulum stress sensor IRE1 signals by dynamic clustering. Proc Natl Acad Sci USA 107:16113–16118

[27] Liao HJ, Carpenter G (2007) Role of the Sec61 translocon in EGF receptor trafficking to the nucleus and gene expression. Mol Biol Cell 18:1064–1072

[28] Liao HJ, Carpenter G (2009) Cetuximab/C225-induced intracellular trafficking of epidermal growth factor receptor. Cancer Res 69:6179–6183

[29] Lipschutz JH, Lingappa VR, Mostov KE (2003) The exocyst affects protein synthesis by acting on the translocation machinery of the endoplasmic reticulum. J Biol Chem 278:20954–20960

[30] Meyer HA, Grau H, Kraft R, Kostka S, Prehn S, Kalies KU, Hartmann E (2000) Mammalian Sec61 is associated with Sec62 and Sec63. J Biol Chem 275:14550–14557

[31] Miyazaki K, Wakana Y, Noda C, Arasaki K, Furuno A, Tagaya M (2012) Contribution of the long form of syntaxin 5 to the organization of the endoplasmic reticulum. J Cell Sci 125:5658–5666

[32] Nikonov AV, Snapp E, Lippincott-Schwartz J, Kreibich G (2002) Active translocon complexes labeled with GFP-Dad1 diffuse slowly as large polysome arrays in the endoplasmic reticulum. J Cell Biol 158:497–506

[33] Nikonov AV, Hauri HP, Lauring B, Kreibich G (2007) Climp-63-mediated binding of microtubules to the ER affects the lateral mobility of translocon complexes. J Cell Sci 120:2248–2258

[34] Park CY, Hoover PJ, Mullins FM, Bachhawat P, Covington ED, Raunser S, Walz T, Garcia KC, Dolmetsch RE, Lewis RS (2009) STIM1 clusters and activates CRAC channels via direct binding of a cytosolic domain to Orai1. Cell 136:876–890

[35] Park SH, Zhu PP, Parker RL, Blackstone C (2010) Hereditary spastic paraplegia proteins REEP1, spastin, and atlastin-1 coordinate microtubule interactions with the tubular ER network. J Clin Invest 120:1097–1110

[36] Plumb R, Zhang ZR, Appathurai S, Mariappan M (2015) A functional link between the co-translational protein translocation pathway and the UPR. Elife 4:e07426

[37] Rapoport TA (2007) Protein translocation across the eukaryotic endoplasmic reticulum and bacterial plasma membranes. Nature 450:663–669

[38] Ron D, Walter P (2007) Signal integration in the endoplasmic reticulum unfolded protein response. Nat Rev Mol Cell Biol 8:519–529

[39] Schroder M, Kaufman RJ (2005) The mammalian unfolded protein response. Annu Rev Biochem 74:739–789

[40] Shibata Y, Voeltz GK, Rapoport TA (2006) Rough sheets and smooth tubules. Cell. 126:435–439

[41] Shibata Y, Shemesh T, Prinz WA, Palazzo AF, Kozlov MM, Rapoport TA (2010) Mechanisms determining the morphology of the peripheral ER. Cell 143:774–788

[42] Staehelin LA (1997) The plant ER: a dynamic organelle composed of a large number of discrete functional domains. Plant J 11:1151–1165

[43] Stefanovic S, Hegde RS (2007) Identification of a targeting factor for posttranslational membrane protein insertion into the ER. Cell 128:1147–1159

[44] Sundaram A, Plumb R, Appathurai S, Mariappan M (2017) The Sec61 translocon limits IRE1alpha signaling during the unfolded protein response. Elife 6:12

[45] Szczesna-Skorupa E, Chen CD, Liu H, Kemper B (2004) Gene expression changes associated with the endoplasmic reticulum stress response induced by microsomal cytochrome p450 overproduction. J Biol Chem 279:13953–13961

[46] Tabas I, Ron D (2011) Integrating the mechanisms of apoptosis induced by endoplasmic reticulum stress. Nat Cell Biol 13:184–190

[47] Terasaki M, Chen LB, Fujiwara K (1986) Microtubules and the endoplasmic reticulum are highly interdependent structures. J Cell Biol 103:1557–1568

[48] Toikkanen JH, Miller KJ, Soderlund H, Jantti J, Keranen S (2003) The beta subunit of the Sec61p endoplasmic reticulum translocon interacts with the exocyst complex in Saccharomyces cerevisiae. J Biol Chem 278:20946–20953

[49] Valcarcel R, Weber U, Jackson DB, Benes V, Ansorge W, Bohmann D, Mlodzik M (1999) Sec61beta, a subunit of the protein translocation channel, is required during Drosophila development. J Cell Sci 112(Pt 23):4389–4396

[50] Vedrenne C, Klopfenstein DR, Hauri HP (2005) Phosphorylation controls CLIMP-63-mediated anchoring of the endoplasmic reticulum to microtubules. Mol Biol Cell 16:1928–1937

[51] Voeltz GK, Rolls MM, Rapoport TA (2002) Structural organization of the endoplasmic reticulum. EMBO Rep 3:944–950

[52] Voeltz GK, Prinz WA, Shibata Y, Rist JM, Rapoport TA (2006) A class of membrane proteins shaping the tubular endoplasmic reticulum. Cell 124:573–586

[53] Walter P, Ron D (2011) The unfolded protein response: from stress pathway to homeostatic regulation. Science 334:1081–1086

[54] Wang YN, Yamaguchi H, Huo L, Du Y, Lee HJ, Lee HH, Wang H, Hsu JM, Hung MC (2010) The translocon Sec61beta localized in the inner nuclear membrane transports membrane-embedded EGF receptor to the nucleus. J Biol Chem 285:38720–38729

[55] Wang S, Romano FB, Field CM, Mitchison TJ, Rapoport TA (2013) Multiple mechanisms determine ER network morphology during the cell cycle in Xenopus egg extracts. J Cell Biol 203:801–814

[56] Wang X, Li S, Wang H, Shui W, Hu J (2017) Quantitative proteomics reveal proteins enriched in tubular endoplasmic reticulum of Saccharomyces cerevisiae. Elife 6:e23816

[57] Yoshida H, Matsui T, Yamamoto A, Okada T, Mori K (2001) XBP1 mRNA is induced by ATF6 and spliced by IRE1 in response to ER stress to produce a highly active transcription factor. Cell 107:881–891

[58] Zhang H, Hu J (2016) Shaping the endoplasmic reticulum into a social network. Trends Cell Biol 26:934–943

[59] Zhang H, Zhou M (2012) Polysome preparation, RNA isolation and analysis. Bio Protoc 2:e286

[60] Zurek N, Sparks L, Voeltz G (2011) Reticulon short hairpin transmembrane domains are used to shape ER tubules. Traffic 12:28–41

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