Dissection of the Influenza A Virus Endocytic Routes Reveals Macropinocytosis as an Alternative Entry Pathway


Influenza A virus (IAV) enters host cells upon binding of its hemagglutinin glycoprotein to sialylated host cell receptors. Whereas dynamin-dependent, clathrin-mediated endocytosis (CME) is generally considered as the IAV infection pathway, some observations suggest the occurrence of an as yet uncharacterized alternative entry route. By manipulating entry parameters we established experimental conditions that allow the separate analysis of dynamin-dependent and -independent entry of IAV. Whereas entry of IAV in phosphate-buffered saline could be completely inhibited by dynasore, a specific inhibitor of dynamin, a dynasore-insensitive entry pathway became functional in the presence of fetal calf serum. This finding was confirmed with the use of small interfering RNAs targeting dynamin-2. In the presence of serum, both IAV entry pathways were operational. Under these conditions entry could be fully blocked by combined treatment with dynasore and the amiloride derivative EIPA, the hallmark inhibitor of macropinocytosis, whereas either drug alone had no effect. The sensitivity of the dynamin-independent entry pathway to inhibitors or dominant-negative mutants affecting actomyosin dynamics as well as to a number of specific inhibitors of growth factor receptor tyrosine kinases and downstream effectors thereof all point to the involvement of macropinocytosis in IAV entry. Consistently, IAV particles and soluble FITC-dextran were shown to co-localize in cells in the same vesicles. Thus, in addition to the classical dynamin-dependent, clathrin-mediated endocytosis pathway, IAV enters host cells by a dynamin-independent route that has all the characteristics of macropinocytosis.


Vyšlo v časopise: Dissection of the Influenza A Virus Endocytic Routes Reveals Macropinocytosis as an Alternative Entry Pathway. PLoS Pathog 7(3): e32767. doi:10.1371/journal.ppat.1001329
Kategorie: Research Article
prolekare.web.journal.doi_sk: https://doi.org/10.1371/journal.ppat.1001329

Souhrn

Influenza A virus (IAV) enters host cells upon binding of its hemagglutinin glycoprotein to sialylated host cell receptors. Whereas dynamin-dependent, clathrin-mediated endocytosis (CME) is generally considered as the IAV infection pathway, some observations suggest the occurrence of an as yet uncharacterized alternative entry route. By manipulating entry parameters we established experimental conditions that allow the separate analysis of dynamin-dependent and -independent entry of IAV. Whereas entry of IAV in phosphate-buffered saline could be completely inhibited by dynasore, a specific inhibitor of dynamin, a dynasore-insensitive entry pathway became functional in the presence of fetal calf serum. This finding was confirmed with the use of small interfering RNAs targeting dynamin-2. In the presence of serum, both IAV entry pathways were operational. Under these conditions entry could be fully blocked by combined treatment with dynasore and the amiloride derivative EIPA, the hallmark inhibitor of macropinocytosis, whereas either drug alone had no effect. The sensitivity of the dynamin-independent entry pathway to inhibitors or dominant-negative mutants affecting actomyosin dynamics as well as to a number of specific inhibitors of growth factor receptor tyrosine kinases and downstream effectors thereof all point to the involvement of macropinocytosis in IAV entry. Consistently, IAV particles and soluble FITC-dextran were shown to co-localize in cells in the same vesicles. Thus, in addition to the classical dynamin-dependent, clathrin-mediated endocytosis pathway, IAV enters host cells by a dynamin-independent route that has all the characteristics of macropinocytosis.


Zdroje

1. PattersonS

OxfordJS

DourmashkinRR

1979 Studies on the mechanism of influenza-virus entry into cells. J Gen Virol 43 223 229

2. MatlinKS

ReggioH

HeleniusA

SimonsK

1981 Infectious entry pathway of Influenza Virus in a Canine Kidney Cell line. J Cell Biol 91 601 613

3. RustMJ

LakadamyaliM

ZhangF

ZhuangX

2004 Assembly of endocytotic machinery around individual influenza viruses during viral entry. Nat Struct Mol Biol 11 567 573

4. ChenC

ZhuangX

2008 Epsin 1 is a cargo-specific adaptor for the clathrin-mediated endocytosis of influenza virus. Proc Natl Acad Sci U S A 105 11790 11795

5. ChuVC

WhittakerGR

2004 Influenza virus entry and infection require host cell N-linked glycoprotein. Proc Natl Acad Sci U S A 101 18153 18158

6. SieczkarskiSB

WhittakerGR

2002 Influenza virus can enter and infect cells in the absence of clathrin-mediated endocytosis. J Virol 76 10455 10464

7. LakadamyaliM

RustMJ

ZhuangX

2004 Endocytosis of influenza viruses. Microb Inf 6 929 936

8. DohertyGJ

McMahonHT

2009 Mechanisms of Endocytosis. Annu Rev Biochem 78 857 902

9. HansenCG

NicholsBJ

2009 Molecular mechanisms of clathrin-independent endocytosis. J Cell Sci 122 1713 1721

10. KumariS

MgS

MayorS

2010 Endocytosis unplugged: multiple ways to enter the cell. Cell Res 20 256 275

11. MarshM

HeleniusA

2006 Virus entry: open sesame. Cell 124 729 740

12. MercerJ

SchelhaasM

HeleniusA

2010 Virus entry by endocytosis. Annu Rev Biochem 79 803 833

13. KerrMC

TeasdaleRD

2009 Defining Macropinocytosis. Traffic 10 364 371

14. MercerJ

HeleniusA

2009 Virus entry by macropinocytosis. Nat Cell Biol 11 510 520

15. FrancisCL

RyanTA

JonesBD

SmithSJ

FalkowS

1993 Ruffles induced by Salmonella and other stimuli direct macropinocytosis of bacteria. Nature 364 639 642

16. HoffmannPR

deCathelineauAM

OgdenCA

LeverrierY

BrattonDL

2001 Phosphatidylserine (PS) induces PS receptor-mediated macropinocytosis and promotes clearance of apoptotic cells. J Cell Biol 155 649 659

17. AmyereM

MettlenM

Van Der SmissenP

PlatekA

PayrastreB

2002 Origin, originality, functions, subversions and molecular signaling of macropinocytosis. Int J Med Microbiol 291 487 494

18. DonaldsonJG

Porat-ShliomN

CohenLA

2009 Clathrin-independent endocytosis: a unique platform for cell signaling and PM remodeling. Cell Signal 21 1 6

19. MayorS

PaganoRE

2007 Pathways of clathrin-independent endocytosis. Nat Rev Mol Cell Biol 8 603 612

20. McClureSJ

RobinsonPJ

1996 Dynamin, endocytosis and intracellular signaling. Mol Membr Biol 13 189 215

21. BonazziM

SpanòS

TuracchioG

CericolaC

ValenteC

2005 CtBP3/BARS drives membrane fission in dynamin-independent transport pathways. Nat Cell Biol 7 570 580

22. LutzA

DyallJ

OlivoPD

PekoszA

2005 Virus-inducible reporter genes as a tool for detecting and quantifying IAV replication. J Virol Meth 126 13 20

23. PerezL

CarrascoL

1994 Involvement of the vacuolar H(+)-ATPase in animal virus entry. J Gen Virol 75 2595 2606

24. SunX

YauVK

BriggsBJ

WhittakerGR

2005 Role of clathrin-mediated endocytosis during vesicular stomatitis virus into different host cells. Virology 338 53 60

25. CuretonDK

MassolRH

SaffarianS

KirchhausenTL

WhelanSP

2009 Vesicular stomatitis virus enters cells through vesicles incompletely coated with clathrin that depend upon actin for internalization. PLOS Pathog 5 e1000394

26. TscherneDM

ManicassamyB

García-SastreA

2010 An enzymatic virus-like particle assay for sensitive detection of virus entry. J Virol Methods 163 336 343

27. KönigR

StertzS

ZhouY

InoueA

HoffmannHH

2010 Human host factors required for influenza virus replication. Nature 463 813 817

28. TangJ

ColacinoJM

LarsenSH

SpitzerW

1990 Virucidal activity of hypericin against enveloped and non-enveloped DNA and RNA viruses. Antiviral Res 13 313 325

29. HoffmannHH

PaleseP

ShawML

2008 Modulation of influenza virus replication by alteration of sodium ion transport and protein kinase C activity. Antivir Res 80 124 134

30. BainJ

McLauchlanH

ElliottM

CohenP

2003 The specificities of protein kinase inhibitors: an update. Biochem J 371 199 204

31. ZerialM

McBrideH

2001 Rab proteins as membrane organizers. Nat Rev Mol Cell Biol 2 107 117

32. SieczkarskiSB

WhittakerGR

2003 Differential requirements of Rab5 and Rab7 for endocytosis of influenza and other enveloped viruses. Traffic 4 333 343

33. SwanssonJA

WattsC

1995 Macropinocytosis. Trends Cell Biol 5 424 428

34. MercerJ

KnebelS

SchmidtFI

CrouseJ

BurkardC

2010 Vaccinia virus strains use distinct forms of macropinocytosis for host-cell entry. Proc Natl Acad Sci U S A 107 9346 9351

35. CoyneCB

BergelsonJM

2006 Virus-induced Abl and Fyn kinase signals permit coxsackievirus entry through the epithelial tight junctions. Cell 124 119 131

36. ArakiN

EgamiY

WatanabeY

HataeT

2007 Phosphoinositide metabolism during membrane ruffling and macropinosome formation in EGF-stimulated A431 cells. Exp Cell Res 313 1496 1507

37. RidleyAJ

1994 Membrane ruffling and signal transduction. Bioessays 16 321 327

38. DharmawardhaneS

SchurmannA

SellsMA

ChernoffJ

SchmidSL

2000 Regulation of macropinocytosis by p21-activated kinase-1. Mol Biol Cell 11 3341 3352

39. DeaconSW

BeeserA

FukuiJA

RennefahrtUE

MyersC

2008 An isoform-selective, small-molecule inhibitor targets the autoregulatory mechanism of p21-activated kinase. Chem Biol 15 305 306

40. RidleyAJ

PatersonHF

JohnstonCL

DiekmannD

HallA

1992 The small GTP-binding protein rac regulates growth factor-induced membrane ruffling. Cell 70 401 410

41. FiorentiniC

FalzanoL

FabbriA

StringaroA

LogozziM

2001 Activation of rho GTPases by cytotoxic necrotizing factor 1 induces macropinocytosis and scavenging activity in epithelial cells. Cell 12 2061 2073

42. HallA

1992 Ras-related GTPases and the cytoskeleton. Mol Biol Cell 3 475 479

43. MercerJ

HeleniusA

2008 Vaccinia virus uses macropinocytosis and apoptotic mimicry to enter host cells. Science 320 531 535

44. RidleyAJ

2006 Rho GTPases and actin dynamics in membrane protrusions and vesicle trafficking. Trends Cell Biol 16 522 529

45. MikiH

TakenawaT

2003 Regulation of actin dynamics by WASP family proteins. J Biochem 134 309 313

46. PelishHE

CieslaW

TanakaN

ReddyK

ShairMD

2006 The Cdc42 inhibitor secramine B prevents cAMP-induced K+ conductance in intestinal epithelial cells. Biochem Pharmacol 71 1720 1726

47. GaoY

DickersonJB

GuoF

ZhengJ

ZhengY

2004 Rational design and characterization of a Rac GTPase-specific small molecule inhibitor. Proc Natl Acad Sci U S A 101 7618 23

48. PetersonJR

BickfordLC

MorganD

KimAS

OuerfelliO

2004 Chemical inhibition of N-WASP by stabilization of a native autoinhibited conformation. Nat Struct Mol Biol 11 747 755

49. KasaharaK

NakayamaY

SatoI

IkedaK

HoshinoM

2007 Role of Src-family kinases in formation and trafficking of macropinosomes. J Cell Physiol 21 220 232

50. MettlenM

PlatekA

Van Der SmissenP

CarpentierS

AmyereM

2006 Src triggers circular ruffling and macropinocytosis at the apical surface of polarized MDCK cells. Traffic 7 589 603

51. DonepudiM

ReshMD

2008 c-Src trafficking and co-localization with the EGF receptor promotes EGF ligand-independent EGF receptor activation and signaling. Cell Signal 20 1359 1367

52. LuskováP

DráberP

2004 Modulation of the Fcepsilon receptor I signaling by tyrosine kinase inhibitors: search for therapeutic targets of inflammatory and allergy diseases. Curr Pharm Des 10 1727 1737

53. PorterJR

GeJ

LeeJ

NormantE

WestK

2009 Ansamycin inhibitors of Hsp90: nature's prototype for anti-chaperone therapy. Curr Top Med Chem 9 1386 1418

54. CrèvecoeurJ

MervilleMP

PietteJ

GloireG

2008 Geldanamycin inhibits tyrosine phosphorylation-dependent NF-kappaB activation. Biochem Pharmacol 75 2183 2191

55. WestMA

BretscherMS

WattsC

1989 Distinct endocytotic pathways in epidermal growth factor-stimulated human carcinoma A431 cells. J Cell Biol 109 2731 2739

56. MeierO

BouckeK

HammerSV

KellerS

StidwillRP

2002 Adenovirus triggers macropinocytosis and endosomal leakage together with its clathrin-mediated uptake. J Cell Biol 1119 1131

57. GoldS

MonaghanP

MertensP

JacksonT

2010 A Clathrin Independent Macropinocytosis-Like Entry Mechanism Used by Bluetongue Virus-1 during Infection of BHK Cells. PLoS ONE 5 e11360

58. RaghuH

Sharma-WaliaN

VeettilMV

SadagopanS

ChandranB

2009 Kaposi's sarcoma-associated herpesvirus utilizes an actin polymerization-dependent macropinocytic pathway to enter human dermal microvascular endothelial and human umbilical vein endothelial cells. J Virol 83 4895 4911

59. RoyAM

ParkerJS

ParrishCR

WhittakerGR

2000 Early stages of influenza virus entry in Mv-1 Lung Cells: involvement of dynamin. Virology 267 17 28

60. MaciaE

EhrlichM

MassolR

BoucrotE

BrunnerC

2006 Dynasore, a cell-permeable inhibitor of dynamin. Dev Cell 10 839 850

61. GlaserL

StevensJ

ZamarinD

WilsonIA

García-SastreA

2005 A single amino acid substitution in 1918 influenza virus hemagglutinin changes receptor binding specificity. J Virol 79 11533 11536

62. KoivusaloM

WelchC

HayashiH

ScottCC

KimM

2010 Amiloride inhibits macropinocytosis by lowering submembranous pH and preventing Rac1 and Cdc42 signaling. J Cell Biol 547 563

63. SnijderB

SacherR

RämöP

DammEM

LiberaliP

2009 Population context determines cell-to-cell variability in endocytosis and virus infection. Nature 461 520 5233

64. SunX

WhittakerGR

2007 Role of the actin cytoskeleton during influenza virus internalization into polarized epithelial cells. Cell Microbiol 9 1672 1682

65. Van den BroekeC

RaduM

ChernoffJ

FavoreelHW

2010 An emerging role for p21-activated kinases (Paks) in viral infections. Trends Cell Biol 20 160 169

66. GaoY

HubbertCC

LuJ

LeeYS

LeeJY

2007 Histone deacetylase 6 regulates growth factor-induced actin remodeling and endocytosis. Mol Cell Biol 27 8637 8647

67. RubinBK

2010 Mucus and mucins. Otolaryngol Clin North Am 43 27 34

68. GharibSA

VaisarT

AitkenML

ParkDR

HeineckeJW

2009 Mapping the lung proteome in cystic fibrosis. J Proteome Res 8 3020 3028

69. GovenderP

DunnMJ

DonnellySC

2009 Proteomics and the lung: Analysis of bronchoalveolar lavage fluid. Proteom Clin Appl 3 1044 1051

70. MagiB

BargagliE

BiniL

RottoliP

2006 Proteome analysis of bronchoalveolar lavage in lung diseases. Proteomics 6 6354 6369

71. KettererMR

ShaoJQ

HornickDB

BuscherB

BandlVK

1999 Infection of Primary Human Bronchial Epithelial Cells by Haemophilus influenza: Macropinocytosis as a Mechanism of Airway Epithelial Cell Entry. Infect Immun 67 4161 4170

72. LuX

TumpeyTM

MorkenT

ZakiSR

CoxNJ

1999 A mouse model for the evaluation of pathogenesis and immunity to influenza A (H5N1) viruses isolated from humans. J Virol 73 5903 5911

73. GaoP

WatanabeS

ItoT

GotoH

WellsK

1999 Biological heterogeneity, including systemic replication in mice, of H5N1 influenza A virus isolates from humans in Hong Kong. J Virol 73 3184 3189

74. FislovaT

GocnikM

SladkovaT

DurmanovaV

RajcaniJ

2009 Multiorgan distribution of human influenza A virus strains observed in a mouse model. Arch Virol 154 409 419

75. RodriguezJF

RodriguezD

RodriguezJR

McGowanEB

EstebanM

1988 Expression of the firefly luciferase gene in vaccinia virus: a highly sensitive gene marker to follow virus dissemination in tissues of infected animals. Proc Natl Acad Sci U S A 85 1667 1671

76. PowerAT

WangTJ

FallsJM

PattersonKA

ParatoBD

2007 Carrier cell-based delivery of an oncolytic virus circumvents antiviral immunity. Mol Ther 15 123 130

77. RaabenM

Groot KoerkampMJ

RottierPJ

de HaanCA

2007 Mouse hepatitis coronavirus replication induces host translational shutoff and mRNA decay, with concomitant formation of stress granules and processing bodies. Cell Microbiol 9 2218 2229

78. JinY

AtkinsonSJ

MarrsJA

GallagherPJ

2001 Myosin ii light chain phosphorylation regulates membrane localization and apoptotic signaling of tumor necrosis factor receptor-1. J Biol Chem 276 30342 30349

79. IwataK

ItoK

FukuzakiA

InakiK

HagaT

1999 Dynamin and rab5 regulate GRK2-dependent internalization of dopamine D2 receptors. Eur J Biochem 263 596 602

80. ReyM

Valenzuela-FernandezA

UrzainquiA

Yanez-MoM

Perez-MartinezM

2007 Myosin IIA is involved in the endocytosis of CXCR4 induced by SDF-1α. J Cell Sci 120 1126 1133

81. DalyRJ

2004 Cortactin signalling and dynamic actin networks. Biochem J 382 13 15

Štítky
Hygiena a epidemiológia Infekčné lekárstvo Laboratórium

Článok vyšiel v časopise

PLOS Pathogens


2011 Číslo 3
Najčítanejšie tento týždeň
Najčítanejšie v tomto čísle
Kurzy

Zvýšte si kvalifikáciu online z pohodlia domova

Získaná hemofilie - Povědomí o nemoci a její diagnostika
nový kurz

Eozinofilní granulomatóza s polyangiitidou
Autori: doc. MUDr. Martina Doubková, Ph.D.

Všetky kurzy
Prihlásenie
Zabudnuté heslo

Zadajte e-mailovú adresu, s ktorou ste vytvárali účet. Budú Vám na ňu zasielané informácie k nastaveniu nového hesla.

Prihlásenie

Nemáte účet?  Registrujte sa