title: Contribution of topology determinants of a viral movement protein to its membrane association, intracellular traffic, and viral cell-to-cell movement.
abstract: The p7B movement protein (MP) of Melon necrotic spot virus (MNSV) is a single-pass membrane protein associated with the endoplasmic reticulum (ER), the Golgi apparatus (GA), and plasmodesmata (Pd). Experimental data presented here revealed that the p7B transmembrane domain (TMD) was sufficient to target the green fluorescent protein (GFP) to ER membranes. In addition, the short extramembrane regions of p7B were essential for subsequent ER export and transport to the GA and Pd. Microsomal partitioning and bimolecular fluorescence assays supported a type II topology of p7B in planta. Mutations affecting conventional determinants of p7B membrane topology, such as the TMD secondary structure, the overall hydrophobicity profile, the so-called "aromatic belt," and the net charge distribution on either side of the TMD, were engineered into infectious RNAs to investigate the relationship between the MP structure and MNSV cell-to-cell movement. The results revealed that (i) the overall hydrophobic profile and the α-helix integrity of the TMD were relevant for virus movement, (ii) modification of the net charge balance of the regions flanking both TMD sides drastically reduced cell-to-cell movement, (iii) localization of p7B to the GA was necessary but not sufficient for virus movement, and (iv) membrane insertion was essential for p7B function in virus movement. Our results therefore indicate that MNSV cell-to-cell movement requires sequential transport of p7B from the ER via the GA to Pd, which is modulated by a combination of several signals with different strengths in the extramembrane regions and TMD of the MP.
authors: Instituto de Biología Molecular y Celular de Plantas (IBMCP), UPV-CSIC, Avenida Ingeniero Fausto Elio, s/n, 46022, Valencia, Spain.
authors: A
authors: A
authors: Genovés
authors: V
authors: V
authors: Pallás
authors: J A
authors: JA
authors: Navarro
references: Biochemistry. 1997 Aug 19;36(33):10213-20
references: 9254619
references: J Gen Physiol. 2007 May;129(5):353-6
references: 17438115
references: Eur J Biochem. 1992 May 1;205(3):1207-15
references: 1577002
references: Proc Natl Acad Sci U S A. 1998 Sep 15;95(19):11169-74
references: 9736708
references: Biophys J. 2001 Oct;81(4):2251-6
references: 11566795
references: EMBO J. 2005 Jul 20;24(14):2533-42
references: 15973434
references: Annu Rev Biochem. 2009;78:515-40
references: 19489728
references: J Gen Virol. 2006 Jun;87(Pt 6):1745-1750
references: 16690941
references: J Gen Virol. 2010 Jul;91(Pt 7):1865-70
references: 20219894
references: J Virol. 2009 Jun;83(11):5535-43
references: 19321624
references: J Mol Biol. 2001 Feb 16;306(2):349-62
references: 11237604
references: Methods Mol Biol. 2008;451:33-54
references: 18370246
references: Bioorg Med Chem. 1999 Jan;7(1):1-7
references: 10199651
references: J Gen Virol. 2006 Sep;87(Pt 9):2699-2707
references: 16894211
references: Plant J. 2006 Apr;46(1):145-54
references: 16553902
references: Nat Rev Mol Cell Biol. 2006 Dec;7(12):909-18
references: 17139331
references: Plant Cell. 2002 May;14(5):1077-92
references: 12034898
references: Mol Plant Microbe Interact. 2001 Sep;14(9):1063-74
references: 11551071
references: Trends Cell Biol. 2003 Jun;13(6):295-300
references: 12791295
references: Proc Natl Acad Sci U S A. 2009 Oct 13;106(41):17594-9
references: 19805075
references: J Biol Chem. 2000 Jun 23;275(25):19409-15
references: 10781599
references: Proc Natl Acad Sci U S A. 2004 Apr 20;101(16):6291-6
references: 15079061
references: Proc Natl Acad Sci U S A. 1998 Feb 17;95(4):1933-7
references: 9465120
references: Mol Plant Microbe Interact. 2010 Mar;23(3):263-72
references: 20121448
references: EMBO J. 2003 Jul 15;22(14):3645-53
references: 12853479
references: Plant Cell. 2005 Jun;17(6):1788-800
references: 15879561
references: Science. 2005 Mar 4;307(5714):1427
references: 15681341
references: Virology. 2007 Oct 25;367(2):348-57
references: 17610929
references: Annu Rev Phytopathol. 2010;48:69-91
references: 20337516
references: Proc Natl Acad Sci U S A. 2008 Oct 14;105(41):15702-7
references: 18840693
references: Plant J. 2011 Mar;65(6):882-96
references: 21251105
references: Curr Opin Struct Biol. 2005 Aug;15(4):378-86
references: 16043344
references: PLoS Biol. 2008 Jan;6(1):e7
references: 18215111
references: Proc Natl Acad Sci U S A. 2008 Mar 18;105(11):4127-32
references: 18326626
references: Proc Natl Acad Sci U S A. 2000 Jun 20;97(13):7112-7
references: 10840061
references: Virology. 2006 Dec 5-20;356(1-2):57-67
references: 16950492
references: J Gen Virol. 2010 Feb;91(Pt 2):404-14
references: 19846669
references: J Mol Biol. 2005 Feb 11;346(1):377-85
references: 15663952
references: J Mol Biol. 1991 Apr 5;218(3):499-503
references: 2016741
references: Biochemistry. 2004 Oct 12;43(40):12716-22
references: 15461443
references: Nature. 2007 Dec 13;450(7172):1026-30
references: 18075582
references: Virology. 2009 Dec 5;395(1):133-42
references: 19796783
references: Mol Plant Microbe Interact. 2008 Mar;21(3):335-45
references: 18257683
references: Nature. 2005 Jan 27;433(7024):377-81
references: 15674282
references: EMBO J. 1995 Aug 1;14(15):3609-16
references: 7641680
references: J Gen Virol. 2006 Aug;87(Pt 8):2371-2380
references: 16847133
references: Trends Plant Sci. 2005 Aug;10(8):376-82
references: 16023398
references: Virology. 2006 Jan 5;344(1):169-84
references: 16364748
references: Nat Struct Biol. 2002 May;9(5):318-9
references: 11976722
references: J Mol Biol. 2007 Feb 16;366(2):366-74
references: 17169373
references: J Mol Biol. 2009 Mar 20;387(1):113-28
references: 19302797
references: J Mol Biol. 2004 May 28;339(2):471-9
references: 15136048
references: Proteins. 2005 Sep 1;60(4):606-16
references: 16028222
references: Trends Biochem Sci. 2005 Jul;30(7):355-7
references: 15935679
references: BMC Cell Biol. 2009 Jun 19;10:48
references: 19538755
references: Plant Signal Behav. 2009 Jan;4(1):35-8
references: 19704702
references: Mol Plant Microbe Interact. 2010 Oct;23(10):1231-47
references: 20831404
references: Proc Natl Acad Sci U S A. 2004 Aug 3;101(31):11263-8
references: 15280537
references: J Virol. 2004 Apr;78(7):3704-9
references: 15016890