© The Rockefeller University Press,
0021-9525/1997//995 $5.00
The Journal of Cell Biology, Volume 136, Number 5,
, 1997 995-1005
Inner but Not Outer Membrane Leaflets Control the Transition from Glycosylphosphatidylinositol-anchored Influenza Hemagglutinin-induced Hemifusion to Full Fusion
Grigory B. Melikyan,
Sofya A. Brener,
Dong C. Ok, and
Fredric S. Cohen
Department of Molecular Biophysics and Physiology, Rush Medical College, Chicago, Illinois 60612
Cells that express wild-type influenza hemagglutinin (HA) fully fuse to RBCs, while cells that express the HA-ectodomain anchored to membranes by glycosylphosphatidylinositol, rather than by a transmembrane domain, only hemifuse to RBCs. Amphipaths were inserted into inner and outer membrane leaflets to determine the contribution of each leaflet in the transition from hemifusion to fusion. When inserted into outer leaflets, amphipaths did not promote the transition, independent of whether the agent induces monolayers to bend outward (conferring positive spontaneous monolayer curvature) or inward (negative curvature). In contrast, when incorporated into inner leaflets, positive curvature agents led to full fusion. This suggests that fusion is completed when a lipidic fusion pore with net positive curvature is formed by the inner leaflets that compose a hemifusion diaphragm. Suboptimal fusion conditions were established for RBCs bound to cells expressing wild-type HA so that lipid but not aqueous dye spread was observed. While this is the same pattern of dye spread as in stable hemifusion, for this "stunted" fusion, lower concentrations of amphipaths in inner leaflets were required to promote transfer of aqueous dyes. Also, these amphipaths induced larger pores for stunted fusion than they generated within a stable hemifusion diaphragm. Therefore, spontaneous curvature of inner leaflets can affect formation and enlargement of fusion pores induced by HA. We propose that after the HA-ectodomain induces hemifusion, the transmembrane domain causes pore formation by conferring positive spontaneous curvature to leaflets of the hemifusion diaphragm.
Abbreviations used in this paper: CF, 6-carboxyfluorescein; CPZ, chlorpromazine; M-CPZ, metho-chlorpromazine; DB, dibucaine; DOPC, dioleoylphosphatidylcholine; DOPE, dioleoylphosphatidylethanolamine; GPI, glycosylphosphatidylinositol; HA, hemagglutinin; HD, hemifusion diaphragm; LPC, lysophosphatidylcholine; L-LPC, lyso-lauroylphosphatidylcholine; S-LPC, lyso-stearoylphosphatidylcholine; MPCA, membranepermeable, cationic amphipaths; NBD-t, NBD-taurine; PS, phosphatidylserine; RD, tetramethylrhodamine-dextran; R18, octadecylrhodamine B chloride; TFP, trifluoperazine; WT, wild type.
Address all correspondence to Fredric S. Cohen, Department of Molecular Biophysics and Physiology, Rush Medical College, 1653 W. Congress Parkway, Chicago, IL 60612. Tel.: (312) 942-6753. Fax: (312) 942-8711. e-mail: fcohen{at}rpslmc.edu
2. CPZ was more efficient in promoting aqueous dye redistribution between RBCs and GPI cells in these experiments than in the experiment of Fig. 3 A. For Fig. 3 A, NBD-t–loaded RBCs were treated with 4,4'-diisothiocyanato-stilbene-2,2'-disulfonic acid (DIDS; Sigma Chemical Co.) to prevent leakage of NBD-t (Sarkar et al., 1989). However, DIDS treatment inhibited R18 redistribution (not shown) and was omitted in this series of experiments to avoid possible interference with LPC or CPZ.

CiteULike
Complore
Connotea
Del.icio.us
Digg
Facebook
Reddit
Technorati
Twitter What's this?
This article has been cited by other articles:
-
Mima, J., Wickner, W.
(2009). Complex Lipid Requirements for SNARE- and SNARE Chaperone-dependent Membrane Fusion. J. Biol. Chem.
284: 27114-27122
[Abstract]
[Full Text]
-
Markosyan, R. M., Leung, M. Y., Cohen, F. S.
(2009). The Six-Helix Bundle of Human Immunodeficiency Virus Env Controls Pore Formation and Enlargement and Is Initiated at Residues Proximal to the Hairpin Turn. J. Virol.
83: 10048-10057
[Abstract]
[Full Text]
-
Li, Z., Blissard, G. W.
(2009). The Autographa californica Multicapsid Nucleopolyhedrovirus GP64 Protein: Analysis of Transmembrane Domain Length and Sequence Requirements. J. Virol.
83: 4447-4461
[Abstract]
[Full Text]
-
Clancy, E. K., Duncan, R.
(2009). Reovirus FAST Protein Transmembrane Domains Function in a Modular, Primary Sequence-Independent Manner To Mediate Cell-Cell Membrane Fusion. J. Virol.
83: 2941-2950
[Abstract]
[Full Text]
-
Niu, X., Gupta, K., Yang, J. T., Shamblott, M. J., Levchenko, A.
(2009). Physical transfer of membrane and cytoplasmic components as a general mechanism of cell-cell communication. J. Cell Sci.
122: 600-610
[Abstract]
[Full Text]
-
Schowalter, R. M., Chang, A., Robach, J. G., Buchholz, U. J., Dutch, R. E.
(2009). Low-pH Triggering of Human Metapneumovirus Fusion: Essential Residues and Importance in Entry. J. Virol.
83: 1511-1522
[Abstract]
[Full Text]
-
Raviv, Y., Blumenthal, R., Tompkins, S. M., Humberd, J., Hogan, R. J., Viard, M.
(2008). Hydrophobic Inactivation of Influenza Viruses Confers Preservation of Viral Structure with Enhanced Immunogenicity. J. Virol.
82: 4612-4619
[Abstract]
[Full Text]
-
Ryu, B. Y., Zavorotinskaya, T., Trentin, B., Albritton, L. M.
(2008). The block to membrane fusion differs with the site of ligand insertion in modified retroviral envelope proteins. J. Gen. Virol.
89: 1049-1058
[Abstract]
[Full Text]
-
Markosyan, R. M., Kielian, M., Cohen, F. S.
(2007). Fusion Induced by a Class II Viral Fusion Protein, Semliki Forest Virus E1, Is Dependent on the Voltage of the Target Cell. J. Virol.
81: 11218-11225
[Abstract]
[Full Text]
-
Jun, Y., Wickner, W.
(2007). Assays of vacuole fusion resolve the stages of docking, lipid mixing, and content mixing. Proc. Natl. Acad. Sci. USA
104: 13010-13015
[Abstract]
[Full Text]
-
Aguilar, H. C., Matreyek, K. A., Choi, D. Y., Filone, C. M., Young, S., Lee, B.
(2007). Polybasic KKR Motif in the Cytoplasmic Tail of Nipah Virus Fusion Protein Modulates Membrane Fusion by Inside-Out Signaling. J. Virol.
81: 4520-4532
[Abstract]
[Full Text]
-
Chernomordik, L. V., Zimmerberg, J., Kozlov, M. M.
(2006). Membranes of the world unite!. JCB
175: 201-207
[Abstract]
[Full Text]
-
Melikyan, G. B., Egelhofer, M., von Laer, D.
(2006). Membrane-Anchored Inhibitory Peptides Capture Human Immunodeficiency Virus Type 1 gp41 Conformations That Engage the Target Membrane prior to Fusion.. J. Virol.
80: 3249-3258
[Abstract]
[Full Text]
-
Garg, H., Blumenthal, R.
(2006). HIV gp41-induced apoptosis is mediated by caspase-3-dependent mitochondrial depolarization, which is inhibited by HIV protease inhibitor nelfinavir. J. Leukoc. Biol.
79: 351-362
[Abstract]
[Full Text]
-
Wallin, M., Loving, R., Ekstrom, M., Li, K., Garoff, H.
(2005). Kinetic Analyses of the Surface-Transmembrane Disulfide Bond Isomerization-Controlled Fusion Activation Pathway in Moloney Murine Leukemia Virus. J. Virol.
79: 13856-13864
[Abstract]
[Full Text]
-
Li, Y., Han, X., Lai, A. L., Bushweller, J. H., Cafiso, D. S., Tamm, L. K.
(2005). Membrane Structures of the Hemifusion-Inducing Fusion Peptide Mutant G1S and the Fusion-Blocking Mutant G1V of Influenza Virus Hemagglutinin Suggest a Mechanism for Pore Opening in Membrane Fusion. J. Virol.
79: 12065-12076
[Abstract]
[Full Text]
-
Giraudo, C. G., Hu, C., You, D., Slovic, A. M., Mosharov, E. V., Sulzer, D., Melia, T. J., Rothman, J. E.
(2005). SNAREs can promote complete fusion and hemifusion as alternative outcomes. JCB
170: 249-260
[Abstract]
[Full Text]
-
Melikyan, G. B., Barnard, R. J. O., Abrahamyan, L. G., Mothes, W., Young, J. A. T.
(2005). Imaging individual retroviral fusion events: From hemifusion to pore formation and growth. Proc. Natl. Acad. Sci. USA
102: 8728-8733
[Abstract]
[Full Text]
-
Zaitseva, E., Mittal, A., Griffin, D. E., Chernomordik, L. V.
(2005). Class II fusion protein of alphaviruses drives membrane fusion through the same pathway as class I proteins. JCB
169: 167-177
[Abstract]
[Full Text]
-
Leikina, E., Mittal, A., Cho, M.-S., Melikov, K., Kozlov, M. M., Chernomordik, L. V.
(2004). Influenza Hemagglutinins Outside of the Contact Zone Are Necessary for Fusion Pore Expansion. J. Biol. Chem.
279: 26526-26532
[Abstract]
[Full Text]
-
Melikyan, G. B., Barnard, R. J. O., Markosyan, R. M., Young, J. A. T., Cohen, F. S.
(2004). Low pH Is Required for Avian Sarcoma and Leukosis Virus Env-Induced Hemifusion and Fusion Pore Formation but Not for Pore Growth. J. Virol.
78: 3753-3762
[Abstract]
[Full Text]
-
Zavorotinskaya, T., Qian, Z., Franks, J., Albritton, L. M.
(2004). A Point Mutation in the Binding Subunit of a Retroviral Envelope Protein Arrests Virus Entry at Hemifusion. J. Virol.
78: 473-481
[Abstract]
[Full Text]
-
Qian, Z., Albritton, L. M.
(2004). An Aromatic Side Chain Is Required at Residue 8 of SU for Fusion of Ecotropic Murine Leukemia Virus. J. Virol.
78: 508-512
[Abstract]
[Full Text]
-
Lin, X., Derdeyn, C. A., Blumenthal, R., West, J., Hunter, E.
(2003). Progressive Truncations C Terminal to the Membrane-Spanning Domain of Simian Immunodeficiency Virus Env Reduce Fusogenicity and Increase Concentration Dependence of Env for Fusion. J. Virol.
77: 7067-7077
[Abstract]
[Full Text]
-
Abrahamyan, L. G., Markosyan, R. M., Moore, J. P., Cohen, F. S., Melikyan, G. B.
(2003). Human Immunodeficiency Virus Type 1 Env with an Intersubunit Disulfide Bond Engages Coreceptors but Requires Bond Reduction after Engagement To Induce Fusion. J. Virol.
77: 5829-5836
[Abstract]
[Full Text]
-
Liu, T., Li, R., Pan, T., Liu, D., Petersen, R. B., Wong, B.-S., Gambetti, P., Sy, M. S.
(2002). Intercellular Transfer of the Cellular Prion Protein. J. Biol. Chem.
277: 47671-47678
[Abstract]
[Full Text]
-
Jeetendra, E., Robison, C. S., Albritton, L. M., Whitt, M. A.
(2002). The Membrane-Proximal Domain of Vesicular Stomatitis Virus G Protein Functions as a Membrane Fusion Potentiator and Can Induce Hemifusion. J. Virol.
76: 12300-12311
[Abstract]
[Full Text]
-
Harman, A., Browne, H., Minson, T.
(2002). The Transmembrane Domain and Cytoplasmic Tail of Herpes Simplex Virus Type 1 Glycoprotein H Play a Role in Membrane Fusion. J. Virol.
76: 10708-10716
[Abstract]
[Full Text]
-
Melikyan, G. B., Markosyan, R. M., Roth, M. G., Cohen, F. S.
(2000). A Point Mutation in the Transmembrane Domain of the Hemagglutinin of Influenza Virus Stabilizes a Hemifusion Intermediate That Can Transit to Fusion. Mol. Biol. Cell
11: 3765-3775
[Abstract]
[Full Text]
-
Melikyan, G. B., Markosyan, R. M., Hemmati, H., Delmedico, M. K., Lambert, D. M., Cohen, F. S.
(2000). Evidence That the Transition of HIV-1 Gp41 into a Six-Helix Bundle, Not the Bundle Configuration, Induces Membrane Fusion. JCB
151: 413-424
[Abstract]
[Full Text]
-
Grote, E., Baba, M., Ohsumi, Y., Novick, P. J.
(2000). Geranylgeranylated Snares Are Dominant Inhibitors of Membrane Fusion. JCB
151: 453-466
[Abstract]
[Full Text]
-
Armstrong, R. T., Kushnir, A. S., White, J. M.
(2000). The Transmembrane Domain of Influenza Hemagglutinin Exhibits a Stringent Length Requirement to Support the Hemifusion to Fusion Transition. JCB
151: 425-438
[Abstract]
[Full Text]
-
Gaudin, Y.
(2000). Rabies Virus-Induced Membrane Fusion Pathway. JCB
150: 601-612
[Abstract]
[Full Text]
-
Leikina, E., Chernomordik, L. V.
(2000). Reversible Merger of Membranes at the Early Stage of Influenza Hemagglutinin-mediated Fusion. Mol. Biol. Cell
11: 2359-2371
[Abstract]
[Full Text]
-
Markosyan, R. M., Cohen, F. S., Melikyan, G. B.
(2000). The Lipid-anchored Ectodomain of Influenza Virus Hemagglutinin (GPI-HA) Is Capable of Inducing Nonenlarging Fusion Pores. Mol. Biol. Cell
11: 1143-1152
[Abstract]
[Full Text]
-
Melikyan, G. B., Markosyan, R. M., Brener, S. A., Rozenberg, Y., Cohen, F. S.
(2000). Role of the Cytoplasmic Tail of Ecotropic Moloney Murine Leukemia Virus Env Protein in Fusion Pore Formation. J. Virol.
74: 447-455
[Abstract]
[Full Text]
-
Qiao, H., Armstrong, R. T., Melikyan, G. B., Cohen, F. S., White, J. M.
(1999). A Specific Point Mutant at Position 1 of the Influenza Hemagglutinin Fusion Peptide Displays a Hemifusion Phenotype. Mol. Biol. Cell
10: 2759-2769
[Abstract]
[Full Text]
-
Melikyan, G. B., Lin, S., Roth, M. G., Cohen, F. S.
(1999). Amino Acid Sequence Requirements of the Transmembrane and Cytoplasmic Domains of Influenza Virus Hemagglutinin for Viable Membrane Fusion. Mol. Biol. Cell
10: 1821-1836
[Abstract]
[Full Text]
-
Kobe, B., Center, R. J., Kemp, B. E., Poumbourios, P.
(1999). Crystal structure of human T cell leukemia virus type 1 gp21 ectodomain crystallized as a maltose-binding protein chimera reveals structural evolution of retroviral transmembrane proteins. Proc. Natl. Acad. Sci. USA
96: 4319-4324
[Abstract]
[Full Text]
-
Wild, P., Schraner, E. M., Peter, J., Loepfe, E., Engels, M.
(1998). Novel Entry Pathway of Bovine Herpesvirus 1 and 5. J. Virol.
72: 9561-9566
[Abstract]
[Full Text]
-
Markovic, I., Pulyaeva, H., Sokoloff, A., Chernomordik, L. V.
(1998). Membrane Fusion Mediated by Baculovirus gp64 Involves Assembly of Stable gp64 Trimers into Multiprotein Aggregates. JCB
143: 1155-1166
[Abstract]
[Full Text]
-
Razinkov, V. I., Melikyan, G. B., Epand, R. M., Epand, R. F., Cohen, F. S.
(1998). Effects of Spontaneous Bilayer Curvature on Influenza Virus-mediated Fusion Pores. JGP
112: 409-422
[Abstract]
[Full Text]
-
Cleverley, D. Z., Lenard, J.
(1998). The transmembrane domain in viral fusion: Essential role for a conserved glycine residue in vesicular stomatitis virus G protein. Proc. Natl. Acad. Sci. USA
95: 3425-3430
[Abstract]
[Full Text]
-
Chernomordik, L. V., Frolov, V. A., Leikina, E., Bronk, P., Zimmerberg, J.
(1998). The Pathway of Membrane Fusion Catalyzed by Influenza Hemagglutinin: Restriction of Lipids, Hemifusion, and Lipidic Fusion Pore Formation. JCB
140: 1369-1382
[Abstract]
[Full Text]
-
Munoz-Barroso, I., Durell, S., Sakaguchi, K., Appella, E., Blumenthal, R.
(1998). Dilation of the Human Immunodeficiency Virus-1 Envelope Glycoprotein Fusion Pore Revealed by the Inhibitory Action of a Synthetic Peptide from gp41. JCB
140: 315-323
[Abstract]
[Full Text]
-
Schroth-Diez, B., Ponimaskin, E., Reverey, H., Schmidt, M. F. G., Herrmann, A.
(1998). Fusion Activity of Transmembrane and Cytoplasmic Domain Chimeras of the Influenza Virus Glycoprotein Hemagglutinin. J. Virol.
72: 133-141
[Abstract]
[Full Text]
-
Beh, C. T., Brizzio, V., Rose, M. D.
(1997). KAR5 Encodes a Novel Pheromone-inducible Protein Required for Homotypic Nuclear Fusion. JCB
139: 1063-1076
[Abstract]
[Full Text]
-
Chang, D.-K., Cheng, S.-F., Deo Trivedi, V., Yang, S.-H.
(2000). The Amino-terminal Region of the Fusion Peptide of Influenza Virus Hemagglutinin HA2 Inserts into Sodium Dodecyl Sulfate Micelle with Residues 16-18 at the Aqueous Boundary at Acidic pH. OLIGOMERIZATION AND THE CONFORMATIONAL FLEXIBILITY. J. Biol. Chem.
275: 19150-19158
[Abstract]
[Full Text]
-
Markovic, I., Leikina, E., Zhukovsky, M., Zimmerberg, J., Chernomordik, L. V.
(2001). Synchronized activation and refolding of influenza hemagglutinin in multimeric fusion machines. JCB
155: 833-844
[Abstract]
[Full Text]