Cryo Electron Tomography of Native HIV-1 Budding Sites


The structure of immature and mature HIV-1 particles has been analyzed in detail by cryo electron microscopy, while no such studies have been reported for cellular HIV-1 budding sites. Here, we established a system for studying HIV-1 virus-like particle assembly and release by cryo electron tomography of intact human cells. The lattice of the structural Gag protein in budding sites was indistinguishable from that of the released immature virion, suggesting that its organization is determined at the assembly site without major subsequent rearrangements. Besides the immature lattice, a previously not described Gag lattice was detected in some budding sites and released particles; this lattice was found at high frequencies in a subset of infected T-cells. It displays the same hexagonal symmetry and spacing in the MA-CA layer as the immature lattice, but lacks density corresponding to NC-RNA-p6. Buds and released particles carrying this lattice consistently lacked the viral ribonucleoprotein complex, suggesting that they correspond to aberrant products due to premature proteolytic activation. We hypothesize that cellular and/or viral factors normally control the onset of proteolytic maturation during assembly and release, and that this control has been lost in a subset of infected T-cells leading to formation of aberrant particles.


Vyšlo v časopise: Cryo Electron Tomography of Native HIV-1 Budding Sites. PLoS Pathog 6(11): e32767. doi:10.1371/journal.ppat.1001173
Kategorie: Research Article
prolekare.web.journal.doi_sk: https://doi.org/10.1371/journal.ppat.1001173

Souhrn

The structure of immature and mature HIV-1 particles has been analyzed in detail by cryo electron microscopy, while no such studies have been reported for cellular HIV-1 budding sites. Here, we established a system for studying HIV-1 virus-like particle assembly and release by cryo electron tomography of intact human cells. The lattice of the structural Gag protein in budding sites was indistinguishable from that of the released immature virion, suggesting that its organization is determined at the assembly site without major subsequent rearrangements. Besides the immature lattice, a previously not described Gag lattice was detected in some budding sites and released particles; this lattice was found at high frequencies in a subset of infected T-cells. It displays the same hexagonal symmetry and spacing in the MA-CA layer as the immature lattice, but lacks density corresponding to NC-RNA-p6. Buds and released particles carrying this lattice consistently lacked the viral ribonucleoprotein complex, suggesting that they correspond to aberrant products due to premature proteolytic activation. We hypothesize that cellular and/or viral factors normally control the onset of proteolytic maturation during assembly and release, and that this control has been lost in a subset of infected T-cells leading to formation of aberrant particles.


Zdroje

1. FreedEO

1998 HIV-1 gag proteins: diverse functions in the virus life cycle. Virology 251 1 15

2. HurleyJH

2008 ESCRT complexes and the biogenesis of multivesicular bodies. Curr Opin Cell Biol 20 4 11

3. MoritaE

SundquistWI

2004 Retrovirus budding. Ann Rev Cell Dev Biol 20 395 425

4. WilliamsRL

UrbeS

2007 The emerging shape of the ESCRT machinery. Nat Rev Mol Cell Biol 8 355 368

5. BriggsJAG

SimonMN

GrossI

KräusslichHG

FullerSD

2004 The stoichiometry of Gag protein in HIV-1. Nat Struct Mol Biol 11 672 675

6. GarrusJE

von SchwedlerUK

PornillosOW

MorhamSG

ZavitzKH

2001 Tsg101 and the vacuolar protein sorting pathway are essential for HIV-1 budding. Cell 107 55 65

7. VerPlankL

BouamrF

LaGrassaTJ

AgrestaB

KikonyogoA

2001 Tsg101, a homologue of ubiquitin-conjugating (E2) enzymes, binds the L domain in HIV type 1 Pr55(Gag). Proc Natl Acad Sci U S A 98 7724 7729

8. CoffinJM

HughesSH

VarmusHE

1997 Retroviruses: Cold Spring Harbor Laboratory Press.

9. PettitSC

MoodyMD

WehbieRS

KaplanAH

NantermetPV

1994 The p2 domain of human immunodeficiency virus type 1 Gag regulates sequential proteolytic processing and is required to produce fully infectious virions. J Virol 68 8017 8027

10. WiegersK

RutterG

KottlerH

TessmerU

HohenbergH

1998 Sequential steps in human immunodeficiency virus particle maturation revealed by alterations of individual Gag polyprotein cleavage sites. J Virol 72 2846 2854

11. CorenLV

ThomasJA

ChertovaE

SowderRC

GagliardiTD

2007 Mutational analysis of the C-terminal Gag cleavage sites in human immunodeficiency virus type 1. J Virol 81 10047 10054

12. CheckleyMA

LuttgeBG

SoheilianF

NagashimaK

FreedEO

2010 The capsid-spacer peptide 1 Gag processing intermediate is a dominant-negative inhibitor of HIV-1 maturation. Virology 400 137 144

13. LeeSK

HarrisJ

SwanstromR

2009 A strongly transdominant mutation in the human immunodeficiency virus type 1 gag gene defines an Achilles heel in the virus life cycle. J Virol 83 8536 8543

14. MüllerB

AndersM

AkiyamaH

WelschS

GlassB

2009 HIV-1 Gag processing intermediates trans-dominantly interfere with HIV-1 infectivity. J Biol Chem 284 29692 29703

15. KräusslichHG

1991 Human immunodeficiency virus proteinase dimer as component of the viral polyprotein prevents particle assembly and viral infectivity. Proc Natl Acad Sci U S A 88 3213 3217

16. KaplanAH

ZackJA

KniggeM

PaulDA

KempfDJ

1993 Partial inhibition of the human immunodeficiency virus type 1 protease results in aberrant virus assembly and the formation of noninfectious particles. J Virol 67 4050 4055

17. MooreMD

FuW

SoheilianF

NagashimaK

PtakRG

2008 Suboptimal inhibition of protease activity in human immunodeficiency virus type 1: effects on virion morphogenesis and RNA maturation. Virology 379 152 160

18. BenjaminJ

Ganser-PornillosBK

TivolWF

SundquistWI

JensenGJ

2005 Three-dimensional structure of HIV-1 virus-like particles by electron cryotomography. J Mol Biol 346 577 588

19. BriggsJA

RichesJD

GlassB

BartonovaV

ZanettiG

2009 Structure and assembly of immature HIV. Proc Natl Acad Sci U S A 106 11090 11095

20. BriggsJAG

GrünewaldK

GlassB

FörsterF

KräusslichHG

2006 The mechanism of HIV-1 core assembly: Insights from three-dimensional reconstructions of authentic virions. Structure 14 15 20

21. LiuJ

BartesaghiA

BorgniaMJ

SapiroG

SubramaniamS

2008 Molecular architecture of native HIV-1 gp120 trimers. Nature 455 109 113

22. WrightER

SchoolerJB

DingHJ

KiefferC

FillmoreC

2007 Electron cryotomography of immature HIV-1 virions reveals the structure of the CA and SP1 Gag shells. EMBO J 26 2218 2226

23. ZhuP

LiuJ

BessJJr

ChertovaE

LifsonJD

2006 Distribution and three-dimensional structure of AIDS virus envelope spikes. Nature 441 847 852

24. ZhuP

WinklerH

ChertovaE

TaylorKA

RouxKH

2008 Cryoelectron tomography of HIV-1 envelope spikes: further evidence for tripod-like legs. PLoS Pathog 4 e1000203

25. BriggsJAG

WilkT

WelkerR

KräusslichHG

FullerSD

2003 Structural organization of authentic, mature HIV-1 virions and cores. EMBO J 22 1707 1715

26. CarlsonLA

BriggsJA

GlassB

RichesJD

SimonMN

2008 Three-dimensional analysis of budding sites and released virus suggests a revised model for HIV-1 morphogenesis. Cell Host Microbe 4 592 599

27. LiS

HillCP

SundquistWI

FinchJT

2000 Image reconstructions of helical assemblies of the HIV-1CA protein. Nature 407 409 413

28. GanserBK

LiS

KlishkoVY

FinchJT

SundquistWI

1999 Assembly and analysis of conical models for the HIV-1 core. Science 283 80 83

29. WagnerR

GrafM

BielerK

WolfH

GrunwaldT

2000 Rev-independent expression of synthetic gag-pol genes of human immunodeficiency virus type 1 and simian immunodeficiency virus: implications for the safety of lentiviral vectors. Hum Gene Ther 11 2403 2413

30. MedaliaO

WeberI

FrangakisAS

NicastroD

GerischG

2002 Macromolecular architecture in eukaryotic cells visualized by cryoelectron tomography. Science 298 1209 1213

31. OttDE

CorenLV

KaneBP

BuschLK

JohnsonDG

1996 Cytoskeletal proteins inside human immunodeficiency virus type 1 virions. J Virol 70 7734 7743

32. GladnikoffM

ShimoniE

GovNS

RoussoI

2009 Retroviral assembly and budding occur through an actin-driven mechanism. Biophys J 97 2419 2428

33. CristRM

DattaSA

StephenAG

SoheilianF

MirroJ

2009 Assembly properties of human immunodeficiency virus type 1 Gag-leucine zipper chimeras: implications for retrovirus assembly. J Virol 83 2216 2225

34. HockleyDJ

NermutMV

GriefC

JowettJB

JonesIM

1994 Comparative morphology of Gag protein structures produced by mutants of the gag gene of human immunodeficiency virus type 1. J Gen Virol 75 Pt 11 2985 2997

35. OttDE

CorenLV

ChertovaEN

GagliardiTD

NagashimaK

2003 Elimination of protease activity restores efficient virion production to a human immunodeficiency virus type 1 nucleocapsid deletion mutant. J Virol 77 5547 5556

36. PopovaE

PopovS

GottlingerHG

2010 Human Immunodeficiency Virus Type 1 Nucleocapsid p1 Confers ESCRT Pathway Dependence. J Virol 84 6590 6597

37. MastronardeDN

2005 Automated electron microscope tomography using robust prediction of specimen movements. J Struct Biol 152 36 51

38. FrankJ

2006 Electron Tomography: Methods for Three-Dimensional Visualization of Structures in the Cell Berlin: Springer

39. NickellS

FörsterF

LinaroudisA

Del NetW

BeekF

2005 TOM software toolbox: acquisition and analysis for electron tomography. Journal of Structural Biology 149 227 234

40. KremerJR

MastronardeDN

McIntoshJR

1996 Computer visualization of three-dimensional image data using IMOD. J Struct Biol 116 71 76

41. WestfallPH

YoungSS

1993 Resampling-based multiple testing: examples and methods for P-value adjustment: Wiley-Interscience 360

Štítky
Hygiena a epidemiológia Infekčné lekárstvo Laboratórium

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