Probing Endoplasmic Reticulum Dynamics using Fluorescence Imaging and Photobleaching Techniques

Lindsey Costantini1, Erik Snapp1

1 Department of Anatomy and Structural Biology, Albert Einstein College of Medicine, Bronx, New York
Publication Name:  Current Protocols in Cell Biology
Unit Number:  Unit 21.7
DOI:  10.1002/0471143030.cb2107s60
Online Posting Date:  September, 2013
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This unit describes approaches and tools for studying the dynamics and organization of endoplasmic reticulum (ER) membranes and proteins in living cells using fluorescence microscopy. The ER plays a key role in secretory protein biogenesis, calcium regulation, and lipid synthesis. However, study of these processes has often been restricted to biochemical assays that average millions of lysed cells or imaging of static fixed cells. With new fluorescent protein (FP) reporter tools, sensitive commercial microscopes, and photobleaching techniques, investigators can interrogate the behaviors of ER proteins, membranes, and stress pathways in single live cells. Solutions are described for imaging challenges relevant to the ER, including the mobility of ER membranes, a range of ER structures, and the influence of post‐translational modifications on FP reporters. Considerations for performing photobleaching assays for ER proteins are discussed. Finally, reporters and drugs for studying misfolded secretory protein stress and the unfolded protein response are described. Curr. Protoc. Cell Biol. 60:21.7.1‐21.7.29. © 2013 by John Wiley & Sons, Inc.

Keywords: FRAP; FLIP; confocal microscopy; live cell imaging; superfolder green fluorescent protein; diffusion; membrane; tubule; microtubule

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Table of Contents

  • Introduction
  • Strategic Planning
  • Basic Protocol 1: Imaging Endoplasmic Reticulum Structures in Fixed Cells
  • Alternate Protocol 1: Imaging Endoplasmic Reticulum Structures in Live Cells
  • Reagents and Solutions
  • Commentary
  • Literature Cited
  • Figures
  • Tables
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Basic Protocol 1: Imaging Endoplasmic Reticulum Structures in Fixed Cells

  • Cultures of adherent cells or cells in suspension expressing fluorescent protein reporter or ER‐specific antigen (see Strategic Planning)
  • Imaging medium (see recipe)
  • Drug treatment stock solutions
    • 10 mg/ml A23187 (Sigma‐Aldrich, cat. no. C7522)/dimethyl sulfoxide (DMSO) or ethanol: store up to 6 months at −20°C
    • 5 mg/ml brefeldin A (Sigma‐Aldrich, cat. no. B7651)/ethanol: store up to 1 year at −20°C
    • 1 M dithiothreitol (DTT; Sigma‐Aldrich, cat. no. D0632)/deionized water: store up to 6 months at −20°C; avoid repeated freeze‐thawing
    • 5 mg/ml nocodazole (Sigma‐Aldrich, cat. no. M1404)/DMSO: store up to 1 year at −20°C
    • 1 M thapsigargin (Sigma‐Aldrich, cat. no. T9033)/DMSO: store up to 6 months at −20°C
    • 5 mg/ml tunicamycin (Tm; Sigma‐Aldrich, cat. no. T7765)/DMSO: store up to 1 year at −20°C; avoid repeated freeze‐thawing
  • Phosphate‐buffered saline (PBS, appendix 2A), warmed to the appropriate cell growth temperature
  • Fixative solution (see recipe)
  • Permeabilization solution (see recipe)
  • Blocking solution: 10% (v/v) fetal bovine serum (FBS)/PBS ( appendix 2A)
  • Primary antibody against ER antigen, e.g., rabbit anti‐PDI (ADI‐SPA‐890‐D; Assay Designs or Thermo Fisher Scientific) and secondary antibody labeled with fluorescent dye
  • Mattek glass‐bottom dishes or Labtek coverglass chambers (Nunc), or similar (treated with concentrated poly‐L‐lysine if using cells in suspension; see receipe)
  • 63× NA 1.4 oil plan apochromat objective and immersion oil supplied by the lens manufacturer
  • Confocal laser scanning (e.g., Carl Zeiss 710, Leica SP8, Olympus Fluoview 1000, or Nikon A1+), high‐speed confocal (e.g., Nikon C2+, Carl Zeiss Live, or Live Duoscan), spinning disk (e.g., Carl Zeiss Cell Observer SD or Olympus DSU), or wide‐field fluorescence microscope adapted for deconvolution (e.g., Deltavision, Applied Precision), with appropriate filter sets and illumination sources.
  • Microscope software, e.g., ImageJ ( or Volocity (Perkin Elmer) to create a three‐dimensional reconstructions
  • Additional reagents and equipment for carrying out confocal microscopy (unit 4.5), wide‐field fluorescence microscopy (unit 4.2), and fluorescence recovery after photobleaching (FRAP; unit 21.1)
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Literature Cited

Literature Cited
  Ai, H.W., Shaner, N.C., Cheng, Z., Tsien, R.Y., and Campbell, R.E. 2007. Exploration of new chromophore structures leads to the identification of improved blue fluorescent proteins. Biochemistry 46:5904‐5910.
  Aronson, D.E., Costantini, L.M., and Snapp, E.L. 2011. Superfolder GFP is fluorescent in oxidizing environments when targeted via the Sec translocon. Traffic 12:543‐548.
  Boyce, M., Bryant, K.F., Jousse, C., Long, K., Harding, H.P., Scheuner, D., Kaufman, R.J., Ma, D., Coen, D.M., Ron, D., and Yuan, J. 2005. A selective inhibitor of eIF2α dephosphorylation protects cells from ER stress. Science 307: 935‐939.
  Brodsky, J.L. and Skach, W.R. 2011. Protein folding and quality control in the endoplasmic reticulum: Recent lessons from yeast and mammalian cell systems. Curr. Opin. Cell Biol. 23:464‐475.
  Brunsing, R., Omori, S.A., Weber, F., Bicknell, A., Friend, L., Rickert, R., and Niwa, M. 2008. B‐ and T‐cell development both involve activity of the unfolded protein response pathway. J. Biol. Chem. 283:17954‐17961.
  Calfon, M., Zeng, H., Urano, F., Till, J.H., Hubbard, S.R., Harding, H.P., Clark, S.G., and Ron, D. 2002. IRE1 couples endoplasmic reticulum load to secretory capacity by processing the XBP‐1 mRNA. Nature 415:92‐96.
  Campbell, R.E., Tour, O., Palmer, A.E., Steinbach, P.A., Baird, G.S., Zacharias, D.A., and Tsien, R.Y. 2002. A monomeric red fluorescent protein. Proc. Natl. Acad. Sci. U.S.A. 99:7877‐7882.
  Cawley, K., Deegan, S., Samali, A., and Gupta, S. 2011. Assays for detecting the unfolded protein response. Methods Enzymol. 490:31‐51.
  Chen, C., Bonifacino, J.S., Yuan, L.C., and Klausner, R.D. 1988. Selective degradation of T cell antigen receptor chains retained in a pre‐Golgi compartment. J. Cell Biol. 107:2149‐2161.
  Chen, X., Shen, J., and Prywes, R. 2002. The luminal domain of ATF6 senses endoplasmic reticulum (ER) stress and causes translocation of ATF6 from the ER to the Golgi. J. Biol. Chem. 277:13045‐13052.
  Costantini, L.M., Fossati, M., Francolini, M., and Snapp, E.L. 2012. Assessing the tendency of fluorescent proteins to oligomerize under physiologic conditions. Traffic 13:643‐649.
  Costantini, L.M., Subach, O.M., Jaureguiberry‐Bravo, M., Verkhusha, V.V., and Snapp, E.L. 2013. Cysteineless nonglycosylated monomeric blue fluorescent protein, secBFP2, for studies in the eukaryotic secretory pathway. Biochem. Biophys. Res. Commun. 430:1114‐1119.
  Crisp, M., Liu, Q., Roux, K., Rattner, J.B., Shanahan, C., Burke, B., Stahl, P.D., and Hodzic, D. 2006. Coupling of the nucleus and cytoplasm: Role of the LINC complex. J. Cell Biol. 172:41‐53.
  Cross, B.C., Bond, P.J., Sadowski, P.G., Jha, B.K., Zak, J., Goodman, J.M., Silverman, R.H., Neubert, T.A., Baxendale, I.R., Ron, D., and Harding, H.P. 2012. The molecular basis for selective inhibition of unconventional mRNA splicing by an IRE1‐binding small molecule. Proc. Natl. Acad. Sci. U.S.A. 109: E869‐E878.
  Cubitt, A.B., Woollenweber, L.A., and Heim, R. 1999. Understanding structure‐function relationships in the Aequorea victoria green fluorescent protein. Methods Cell Biol. 58:19‐30.
  Dayel, M.J., Hom, E.F., and Verkman, A.S. 1999. Diffusion of green fluorescent protein in the aqueous‐phase lumen of endoplasmic reticulum. Biophys. J. 76:2843‐2851.
  DuRose, J.B., Tam, A.B., and Niwa, M. 2006. Intrinsic capacities of molecular sensors of the unfolded protein response to sense alternate forms of endoplasmic reticulum stress. Mol. Biol. Cell 17:3095‐3107.
  Ellenberg, J., Siggia, E.D., Moreira, J.E., Smith, C.L., Presley, J.F., Worman, H.J., and Lippincott‐Schwartz, J. 1997. Nuclear membrane dynamics and reassembly in living cells: Targeting of an inner nuclear membrane protein in interphase and mitosis. J. Cell Biol. 138:1193‐1206.
  English, A.R. and Voeltz, G.K. 2012. Rab10 GTPase regulates ER dynamics and morphology. Nat. Cell Biol. 15:169‐178.
  English, A.R., Zurek, N., and Voeltz, G.K. 2009. Peripheral ER structure and function. Curr. Opin. Cell Biol. 21:596‐602.
  Fribley, A., Zhang, K., and Kaufman, R.J. 2009. Regulation of apoptosis by the unfolded protein response. Methods Mol. Biol. 559:191‐204.
  Friedman, J.R. and Voeltz, G.K. 2011. The ER in 3D: A multifunctional dynamic membrane network. Trends Cell Biol. 21:709‐717.
  Friedman, J.R., Lackner, L.L., West, M., DiBenedetto, J.R., Nunnari, J., and Voeltz, G.K. 2011. ER tubules mark sites of mitochondrial division. Science 334:358‐362.
  Gaietta, G., Deerinck, T.J., Adams, S.R., Bouwer, J., Tour, O., Laird, D.W., Sosinsky, G.E., Tsien, R.Y., and Ellisman, M.H. 2002. Multicolor and electron microscopic imaging of connexin trafficking. Science 296:503‐507.
  Gambin, Y., Lopez‐Esparza, R., Reffay, M., Sierecki, E., Gov, N.S., Genest, M., Hodges, R.S., and Urbach, W. 2006. Lateral mobility of proteins in liquid membranes revisited. Proc. Natl. Acad. Sci. U.S.A. 103:2098‐2102.
  Gass, J.N., Gifford, N.M., and Brewer, J.W. 2002. Activation of an unfolded protein response during differentiation of antibody‐secreting B cells. J. Biol. Chem. 277:49047‐49054.
  Griffiths, G., Warren, G., Quinn, P., Mathieu‐Costello, O., and Hoppeler, H. 1984. Density of newly synthesized plasma membrane proteins in intracellular membranes. I. Stereological studies. J. Cell Biol. 98:2133‐2141.
  Gruber, C.W., Cemazar, M., Heras, B., Martin, J.L., and Craik, D.J. 2006. Protein disulfide isomerase: The structure of oxidative folding. Trends Biochem. Sci. 31:455‐464.
  Guo, F. and Snapp, E.L. 2013. ERdj3 regulates BiP occupancy in living cells. J Cell Sci. 126:1429‐1439.
  Harding, H.P., Novoa, I., Zhang, Y., Zeng, H., Wek, R., Schapira, M., and Ron, D. 2000a. Regulated translation initiation controls stress‐induced gene expression in mammalian cells. Mol. Cell 6:1099‐1108.
  Harding, H.P., Zhang, Y., Bertolotti, A., Zeng, H., and Ron, D. 2000b. Perk is essential for translational regulation and cell survival during the unfolded protein response. Mol. Cell 5: 897‐904.
  Harding, H.P., Zyryanova, A.F., and Ron, D. 2012. Uncoupling proteostasis and development in vitro with a small molecule inhibitor of the pancreatic endoplasmic reticulum kinase, PERK. J. Biol. Chem. 287:44338‐44344.
  Hegde, R.S. and Bernstein, H.D. 2006. The surprising complexity of signal sequences. Trends Biochem. Sci. 31:563‐571.
  Iizuka, R., Yamagishi‐Shirasaki, M., and Funatsu, T. 2011. Kinetic study of de novo chromophore maturation of fluorescent proteins. Anal. Biochem. 414:173‐178.
  Ingolia, N.T., Lareau, L.F., and Weissman, J.S. 2011. Ribosome profiling of mouse embryonic stem cells reveals the complexity and dynamics of mammalian proteomes. Cell 147:789‐802.
  Iwakoshi, N.N., Lee, A.H., Vallabhajosyula, P., Otipoby, K.L., Rajewsky, K., and Glimcher, L.H. 2003. Plasma cell differentiation and the unfolded protein response intersect at the transcription factor XBP‐1. Nat. Immunol. 4:321‐329.
  Iwawaki, T., Akai, R., Kohno, K., and Miura, M. 2004. A transgenic mouse model for monitoring endoplasmic reticulum stress. Nat. Med. 10:98‐102.
  Jackson, M.R., Nilsson, T., and Peterson, P.A. 1993. Retrieval of transmembrane proteins to the endoplasmic reticulum. J. Cell Biol. 121: 317‐333.
  Jain, R.K., Joyce, P.B., Molinete, M., Halban, P.A., and Gorr, S.U. 2001. Oligomerization of green fluorescent protein in the secretory pathway of endocrine cells. Biochem. J. 360:645‐649.
  Kaji, E.H. and Lodish, H.F. 1993. In vitro unfolding of retinol‐binding protein by dithiothreitol. Endoplasmic reticulum‐associated factors. J. Biol. Chem. 268:22195‐22202.
  Kelkar, D.A., Khushoo, A., Yang, Z., and Skach, W.R. 2012. Kinetic analysis of ribosome‐bound fluorescent proteins reveals an early, stable, cotranslational folding intermediate. J. Biol. Chem. 287:2568‐2578.
  Koch, G.L., Booth, C., and Wooding, F.B. 1988. Dissociation and re‐assembly of the endoplasmic reticulum in live cells. J. Cell Sci. 91:511‐522.
  Kwon, S.E. and Chapman, E.R. 2012. Glycosylation is dispensable for sorting of synaptotagmin 1 but is critical for targeting of SV2 and synaptophysin to recycling synaptic vesicles. J. Biol. Chem. 287:35658‐35668.
  Lai, C.W., Aronson, D.E., and Snapp, E.L. 2010. BiP availability distinguishes states of homeostasis and stress in the endoplasmic reticulum of living cells. Mol. Biol. Cell 21:1909‐1921.
  Lai, C.W., Otero, J.H., Hendershot, L.M., and Snapp, E. 2012. ERdj4 protein is a soluble endoplasmic reticulum (ER) DnaJ family protein that interacts with ER‐associated degradation machinery. J. Biol. Chem. 287:7969‐7978.
  Lajoie, P., Moir, R.D., Willis, I.M., and Snapp, E.L. 2012. Kar2p Availability defines distinct forms of endoplasmic reticulum stress in living cells. Mol. Biol. Cell 23:955‐964.
  Lee, A.H., Iwakoshi, N.N., Anderson, K.C., and Glimcher, L.H. 2003. Proteasome inhibitors disrupt the unfolded protein response in myeloma cells. Proc. Natl. Acad. Sci. U.S.A. 100:9946‐9951.
  Lee, C. and Chen, L.B. 1988. Dynamic behavior of endoplasmic reticulum in living cells. Cell 54:37‐46.
  Lippincott‐Schwartz, J., Roberts, T.H., and Hirschberg, K. 2000. Secretory protein trafficking and organelle dynamics in living cells. Annu. Rev. Cell Dev. Biol. 16:557‐589.
  Lodish, H.F. and Kong, N. 1993. The secretory pathway is normal in dithiothreitol‐treated cells, but disulfide‐bonded proteins are reduced and reversibly retained in the endoplasmic reticulum. J. Biol. Chem. 268:20598‐20605.
  Lu, L., Ladinsky, M.S., and Kirchhausen, T. 2009. Cisternal organization of the endoplasmic reticulum during mitosis. Mol. Biol. Cell 20:3471‐3480.
  Luo, S. and Lee, A.S. 2002. Requirement of the p38 mitogen‐activated protein kinase signalling pathway for the induction of the 78 kDa glucose‐regulated protein/immunoglobulin heavy‐chain binding protein by azetidine stress: Activating transcription factor 6 as a target for stress‐induced phosphorylation. Biochem. J. 366: 787‐795.
  Ma, Y. and Hendershot, L.M. 2004. ER chaperone functions during normal and stress conditions. J. Chem. Neuroanat. 28:51‐65.
  Maiuolo, J., Bulotta, S., Verderio, C., Benfante, R., and Borgese, N. 2011. Selective activation of the transcription factor ATF6 mediates endoplasmic reticulum proliferation triggered by a membrane protein. Proc. Natl. Acad. Sci. U.S.A. 108:7832‐7837.
  Marguet, D., Spiliotis, E.T., Pentcheva, T., Lebowitz, M., Schneck, J., and Edidin, M. 1999. Lateral diffusion of GFP‐tagged H2Ld molecules and of GFP‐TAP1 reports on the assembly and retention of these molecules in the endoplasmic reticulum. Immunity 11:231‐240.
  Merksamer, P.I., Trusina, A., and Papa, F.R. 2008. Real‐time redox measurements during endoplasmic reticulum stress reveal interlinked protein folding functions. Cell 135:933‐947.
  Michelangeli, F. and East, J.M. 2011. A diversity of SERCA Ca2+ pump inhibitors. Biochem. Soc. Trans. 39:789‐797.
  Munro, S. and Pelham, H.R.B. 1987. A C‐terminal signal prevents secretion of luminal ER proteins. Cell 48:899‐907.
  Nagai, N., Hosokawa, M., Itohara, S., Adachi, E., Matsushita, T., Hosokawa, N., and Nagata, K. 2000. Embryonic lethality of molecular chaperone hsp47 knockout mice is associated with defects in collagen biosynthesis. J. Cell Biol. 150:1499‐1506.
  Nagaya, H., Tamura, T., Higa‐Nishiyama, A., Ohashi, K., Takeuchi, M., Hashimoto, H., Hatsuzawa, K., Kinjo, M., Okada, T., and Wada, I. 2008. Regulated motion of glycoproteins revealed by direct visualization of a single cargo in the endoplasmic reticulum. J. Cell Biol. 180:129‐143.
  Nehls, S., Snapp, E.L., Cole, N.B., Zaal, K.J., Kenworthy, A.K., Roberts, T.H., Ellenberg, J., Presley, J.F., Siggia, E., and Lippincott‐Schwartz, J. 2000. Dynamics and retention of misfolded proteins in native ER membranes. Nat. Cell Biol. 2:288‐295.
  Olveczky, B.P. and Verkman, A.S. 1998. Monte Carlo analysis of obstructed diffusion in three dimensions: Application to molecular diffusion in organelles. Biophys. J. 74:2722‐2730.
  Ordóñez, A., Snapp, E.L., Tan, L., Miranda, E., Marciniak, S.J., and Lomas, D.A. 2012. Endoplasmic reticulum polymers impair luminal protein mobility and sensitise to cellular stress in α (1) ‐antitrypsin deficiency. Hepatology 57:2049‐2060.
  Oslowski, C.M. and Urano, F. 2011. Measuring ER stress and the unfolded protein response using mammalian tissue culture system. Methods Enzymol. 490:71‐92.
  Ostrovsky, O., Makarewich, C.A., Snapp, E.L., and Argon, Y. 2009. An essential role for ATP binding and hydrolysis in the chaperone activity of GRP94 in cells. Proc. Natl. Acad. Sci. U.S.A. 106:11600‐11605.
  Pedelacq, J.D., Cabantous, S., Tran, T., Terwilliger, T.C., and Waldo, G.S. 2006. Engineering and characterization of a superfolder green fluorescent protein. Nat. Biotechnol. 24:79‐88.
  Pincus, D., Chevalier, M.W., Aragon, T., van Anken, E., Vidal, S.E., El‐Samad, H., and Walter, P. 2010. BiP binding to the ER‐stress sensor Ire1 tunes the homeostatic behavior of the unfolded protein response. PLoS Biol. 8:e100415.
  Rutkowski, D.T., Arnold, S.M., Miller, C.N., Wu, J., Li, J., Gunnison, K.M., Mori, K., Sadighi Akha, A.A., Raden, D., and Kaufman, R.J. 2006. Adaptation to ER stress is mediated by differential stabilities of pro‐survival and pro‐apoptotic mRNAs and proteins. PLoS Biol. 4:e374.
  Saffman, P.G. and Delbruck, M. 1975. Brownian motion in biological membranes. Proc. Natl. Acad. Sci. U.S.A. 72:3111‐3113.
  Sbalzarini, I.F., Mezzacasa, A., Helenius, A., and Koumoutsakos, P. 2005. Effects of organelle shape on fluorescence recovery after photobleaching. Biophys. J. 89:1482‐1492.
  Shaner, N.C., Campbell, R.E., Steinbach, P.A., Giepmans, B.N., Palmer, A.E., and Tsien, R.Y. 2004. Improved monomeric red, orange and yellow fluorescent proteins derived from Discosoma sp. red fluorescent protein. Nat. Biotechnol. 22:1567‐1572.
  Shaner, N.C., Lin, M.Z., McKeown, M.R., Steinbach, P.A., Hazelwood, K.L., Davidson, M.W., and Tsien, R.Y. 2008. Improving the photostability of bright monomeric orange and red fluorescent proteins. Nat. Methods 5: 545‐551.
  Shibata, Y., Voeltz, G.K., and Rapoport, T.A. 2006. Rough sheets and smooth tubules. Cell 126:435‐439.
  Shibata, Y., Shemesh, T., Prinz, W.A., Palazzo, A.F., Kozlov, M.M., and Rapoport, T.A. 2010. Mechanisms determining the morphology of the peripheral ER. Cell 143:774‐788.
  Siggia, E.D., Lippincott‐Schwartz, J., and Bekiranov, S. 2000. Diffusion in an inhomogeneous media: Theory and simulations applied to a whole cell photobleach recovery. Biophys. J. 79:1761‐1770.
  Snapp, E.L. 2003. ER biogenesis: Proliferation and differentiation. In The Biogenesis of Cellular Organelles (C. Mullins, ed.) pp. 63‐95. Landes Bioscience and Kluwer Academic/Plenum Publishers, New York.
  Snapp, E.L. 2009. Fluorescent proteins: A cell biologist's user guide. Trends Cell Biol. 19: 649‐655.
  Snapp, E.L., Hegde, R.S., Francolini, M., Lombardo, F., Colombo, S., Pedrazzini, E., Borgese, N., and Lippincott‐Schwartz, J. 2003. Formation of stacked ER cisternae by low affinity protein interactions. J. Cell Biol. 163:257‐269.
  Snapp, E.L., Sharma, A., Lippincott‐Schwartz, J., and Hegde, R.S. 2006. Monitoring chaperone engagement of substrates in the endoplasmic reticulum of live cells. Proc. Natl. Acad. Sci. U.S.A. 103:6536‐6541.
  Spencer, S.L., Gaudet, S., Albeck, J.G., Burke, J.M., and Sorger, P.K. 2009. Non‐genetic origins of cell‐to‐cell variability in TRAIL‐induced apoptosis. Nature 459:428‐432.
  Subramanian, K. and Meyer, T. 1997. Calcium‐induced restructuring of nuclear envelope and endoplasmic reticulum calcium stores. Cell 89:963‐971.
  Suzuki, T., Arai, S., Takeuchi, M., Sakurai, C., Ebana, H., Higashi, T., Hashimoto, H., Hatsuzawa, K., and Wada, I. 2012. Development of cysteine‐free fluorescent proteins for the oxidative environment. PLoS One 7:e37551.
  Swedlow, J.R. 2007. Quantitative fluorescence microscopy and image deconvolution. Methods Cell Biol. 81:447‐465.
  Tabas, I. and Ron, D. 2011. Integrating the mechanisms of apoptosis induced by endoplasmic reticulum stress. Nat. Cell Biol. 13:184‐190.
  Taylor, C.W. and Broad, L.M. 1998. Pharmacological analysis of intracellular Ca2+ signalling: Problems and pitfalls. Trends Pharmacol. Sci. 19:370‐375.
  Terasaki, M. and Jaffe, L.A. 1991. Organization of the sea urchin egg endoplasmic reticulum and its reorganization at fertilization. J. Cell Biol. 114:929‐940.
  Terasaki, M., Chen, L.B., and Fujiwara, K. 1986. Microtubules and the endoplasmic reticulum are highly interdependent structures. J. Cell Biol. 103:1557‐1568.
  Tsaytler, P., Harding, H.P., Ron, D., and Bertolotti, A. 2011. Selective inhibition of a regulatory subunit of protein phosphatase 1 restores proteostasis. Science 332:91‐94.
  Upton, J.P., Wang, L., Han, D., Wang, E.S., Huskey, N.E., Lim, L., Truitt, M., McManus, M.T., Ruggero, D., Goga, A., Papa, F.R., and Oakes, S.A. 2012. IRE1α cleaves select microRNAs during ER stress to derepress translation of proapoptotic Caspase‐2. Science 338:818‐822.
  Voeltz, G.K., Rolls, M.M., and Rapoport, T.A. 2002. Structural organization of the endoplasmic reticulum. EMBO Rep. 3:944‐950.
  Volkmann, K., Lucas, J.L., Vuga, D., Wang, X., Brumm, D., Stiles, C., Kriebel, D., Der‐Sarkissian, A., Krishnan, K., Schweitzer, C., Liu, Z., Malyankar, U.M., Chiovitti, D., Canny, M., Durocher, D., Sicheri, F., and Patterson, J.B. 2011. Potent and selective inhibitors of the inositol‐requiring enzyme 1 endoribonuclease. J. Biol. Chem. 286:12743‐12755.
  Wallace, W., Schaefer, L.H., and Swedlow, J.R. 2001. A workingperson's guide to deconvolution in light microscopy. Biotechniques 31:1076‐1078, 1080, 1082 passim.
  Walter, P. and Ron, D. 2011. The unfolded protein response: From stress pathway to homeostatic regulation. Science 334:1081‐1086.
  Wang, S. and Kaufman, R.J. 2012. The impact of the unfolded protein response on human disease. J. Cell Biol. 197:857‐867.
  West, M., Zurek, N., Hoenger, A., and Voeltz, G.K. 2011. A 3D analysis of yeast ER structure reveals how ER domains are organized by membrane curvature. J. Cell Biol. 193:333‐346.
  Wu, J., Rutkowski, D.T., Dubois, M., Swathirajan, J., Saunders, T., Wang, J., Song, B., Yau, G.D., and Kaufman, R.J. 2007. ATF6α optimizes long‐term endoplasmic reticulum function to protect cells from chronic stress. Dev. Cell 13:351‐364.
  Zeligs, J.D. and Wollman, S.H. 1979. Mitosis in rat thyroid epithelial cells in vivo. J. Ultrastruct. Res. 66:53‐77.
  Zaal, K.J.M., Smith, C.L., Polishchuk, R.S., Altan, N., Cole, N.B., Ellenberg, J., Hirschberg, K., Presley, J.F., Roberts, T.H., Siggia, E., Phair, R.D., and Lippincott‐Schwartz, J. 1999. Golgi membranes are absorbed into and reemerge from the ER during mitosis. Cell 99:589‐601.
  Zacharias, D.A., Violin, J.D., Newton, A.C., and Tsien, R.Y. 2002. Partitioning of lipid‐modified monomeric GFPs into membrane microdomains of live cells. Science 296:913‐916.
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