Live-cell coimaging of the genomic RNAs and Gag proteins of two lentiviruses.
Human immunodeficiency virus type 1 (HIV-1) Gag and genomic RNA determinants required for encapsidation are well established, but where and when encapsidation occurs in the cell is unknown. We constructed MS2 phage coat protein labeling systems to track spatial dynamics of primate and nonprimate lentiviral genomic RNAs (HIV-1 and feline immunodeficiency virus [FIV]) vis-à-vis their Gag proteins in live cells. Genomic RNAs of both lentiviral genera were observed to traffic into the cytoplasm, and this was Rev dependent. In transit, FIV Gag and genomic RNA accumulated independently of each other at the nuclear envelope, and focal colocalizations of genomic RNA with an intact packaging signal (psi) and Gag were observed to extend outward from the cytoplasmic face. In contrast, although HIV-1 genomic RNA was detected at the nuclear envelope, HIV-1 Gag was not. For both lentiviruses, genomic RNAs were seen at the plasma membrane if and only if Gag was present and psi was intact. In addition, HIV-1 and FIV genomes accumulated with Gag in late endosomal foci, again, only psi dependently. Thus, lentiviral genomic RNAs require specific Gag binding to accumulate at the plasma membrane, packaged genomes cointernalize with Gag into the endosomal pathway, and plasma membrane RNA incorporation by Gag does not trigger committed lentiviral particle egress from the cell. Based on the FIV results, we hypothesize that the Gag-genome association may initiate at the nuclear envelope.
Kemler
Iris
I
Department of Molecular Medicine, Guggenheim 18, Mayo Clinic College of Medicine, 200 First Street SW, Rochester, MN 55905, USA.
Meehan
Anne
A
Poeschla
Eric M
EM
D000818:Animals; D002460:Cell Line; D002462:Cell Membrane / Q000737:chemistry; D002467:Cell Nucleus / Q000737:chemistry; D003593:Cytoplasm / Q000737:chemistry; D011992:Endosomes / Q000737:chemistry; D015683:Gene Products, gag / Q000378:metabolism*; D015497:HIV-1 / Q000502:physiology*; D006801:Humans; D007091:Image Processing, Computer-Assisted / Q000379:methods; D016089:Immunodeficiency Virus, Feline / Q000502:physiology*; D012367:RNA, Viral / Q000378:metabolism*; D019065:Virus Assembly
D015683:Gene Products, gag; D012367:RNA, Viral
PLoS Pathog. 2009 Oct;5(10):e1000627
19834549
J Virol. 2002 Dec;76(23):11889-903
12414931
Dev Cell. 2003 Jul;5(1):161-74
12852860
J Virol. 2001 Dec;75(24):12058-69
11711596
Traffic. 2003 Nov;4(11):785-801
14617360
Nat Biotechnol. 1997 Sep;15(9):871-5
9306402
Traffic. 2006 Sep;7(9):1177-93
17004321
Cell. 2004 Oct 29;119(3):381-92
15507209
Methods Mol Biol. 2003;229:251-71
12824636
Virology. 2004 Dec 20;330(2):471-80
15567440
Methods Enzymol. 2000;318:493-506
10890008
J Virol. 1992 Jul;66(7):4144-53
1602537
Proc Natl Acad Sci U S A. 2009 Nov 10;106(45):19114-9
19861549
Traffic. 2006 Jun;7(6):731-45
16683918
J Cell Sci. 2003 Feb 15;116(Pt 4):587-97
12538759
Curr Biol. 2003 Jan 21;13(2):161-167
12546792
Annu Rev Cell Dev Biol. 2004;20:395-425
15473846
Cell. 2005 Mar 11;120(5):663-74
15766529
Virology. 2006 Jan 5;344(1):55-63
16364736
Proc Natl Acad Sci U S A. 1994 Feb 15;91(4):1256-60
8108397
Nat Rev Microbiol. 2005 Aug;3(8):643-55
16064056
J Mol Biol. 2005 Nov 25;354(2):330-9
16253274
J Virol. 2007 Oct;81(19):10718-28
17634229
Virology. 2004 Sep 15;327(1):111-20
15327902
Nat Med. 1998 Mar;4(3):354-7
9500613
Science. 1998 Jan 16;279(5349):384-8
9430589
J Virol. 2007 Jul;81(14):7476-90
17507489
J Virol. 2002 Dec;76(23):12381-7
12414982
Invest Ophthalmol Vis Sci. 2002 Dec;43(12):3686-90
12454037
Traffic. 2005 Sep;6(9):741-55
16101678
PLoS Pathog. 2007 Mar;3(3):e36
17381240
Mol Cell. 1998 Apr;1(5):649-59
9660949
J Cell Biol. 2007 Apr 23;177(2):329-41
17438075
Traffic. 2003 Dec;4(12):902-10
14617353
Virus Res. 2003 Jun;93(2):199-209
12782368
Nature. 2008 Jul 10;454(7201):236-40
18500329
J Virol. 2005 Apr;79(7):4055-65
15767407
Traffic. 2003 Oct;4(10):660-70
12956869
J Virol. 1996 May;70(5):2963-73
8627772
J Virol. 2007 Jun;81(12):6216-30
17428849
Mol Cell. 1998 Oct;2(4):437-45
9809065
Prog Mol Subcell Biol. 2004;35:135-50
15113083
Nat Biotechnol. 2004 Dec;22(12):1567-72
15558047
Traffic. 2003 Oct;4(10):671-80
12956870
Mol Cell Biol. 2004 Apr;24(7):2637-48
15024055
J Virol. 2007 Apr;81(8):4002-11
17267488
J Cell Biol. 2002 Nov 11;159(3):441-52
12417576
PLoS Pathog. 2009 Nov;5(11):e1000652
19893629
J Virol. 1998 Aug;72(8):6858-66
9658135
PLoS Biol. 2006 Dec;4(12):e435
17147474
Vet Immunol Immunopathol. 2010 Mar 15;134(1-2):3-13
19910057
J Virol. 2008 Mar;82(5):2106-19
18094166
Proc Natl Acad Sci U S A. 1989 Aug;86(15):5781-5
2788277
J Virol. 1989 Sep;63(9):4085-7
2760989
Proc Natl Acad Sci U S A. 2002 Mar 19;99(6):3944-9
11891341