Presentacin de PowerPoint

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Cell-to-cell HIV-1 transmission through a
clathrin-dependent endocytic pathway
Berta Bosch, Boyan Grigorov, Jordi Senserrich,
Bonaventura Clotet, Jean-Luc Darlix, Delphine
Muriaux and José A. Este
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HIV cell-to-cell transmission

• Coculture of HIV presenting cells (chronically
  infected MOLT cell lines) and uninfected cells
  (primary lymphocytes).

Untreated
Trypsin treated
NL4-3
BaL
HIV-1 CA p24
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HIV-1 cell-to-cell transmission depends on the
interaction between gp120 and CD4
No Drug
Ca p24 cells ( of target cells)
mAb IgGb12
C-34
-
-
-
Tryp
Tryp
Tryp
HIV-1 BaL MOLT
Uninfected MOLT
HIV-1NL4-3 MOLT
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Internalized HIV resides in vesicular
compartments even in the presence of coreceptor
and fusion inhibitors
No Drug
AMD3100
Ca p24 cells ( of target cells)
AZT
TAK-779
C-34
mAb IgGb12
Uninfected MOLT
HIV-1 BaL MOLT
HIV-1NL4-3 MOLT
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Endocytosis
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Objectives

• To characterize the subcellular compartments
  witch contain the internalized HIV particles.
• To elucidate the fate of the internalized HIV
  particles.

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HIV-1 particles transferred to CD4 T lymphocytes
contained CD63
Surface
CD63 cells ( of target cells)
Intracellular
CD63
CD63
CD63
CD63
Isotype
Isotype
Isotype
Isotype
HIV-1BaL MOLT
HIV-1CI-1-SI MOLT
HIV-1NL4-3 MOLT
Uninfected MOLT
No Drug
mAb IgGb12
C-34
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Endocyted HIV was later released in the
supernatant as infectious viral particles
Purification of CD4 T cells
Trypsin treatment
12 h Incubation
MOLT/BaL CD4 T cells (6 h.)
Infection of U87.CD4.CCR5.
?
Released CA p24 (pg/ml)
Infectivity (Infectious units/ml)
Cell-associated CA p24 staining (MFI)
Uninfected
No Drug
TAK-779
Uninfected
No Drug
mAb IgGb12
mAb IgGb12
TAK-779
BaL
Time (hours)
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Conclusions

• After cell-to-cell contacts, complete HIV
  particles are transferred into target cells
  (stimulated or unstimulated CD4 T lymphocytes).
  These viral particles are localized in
  compartments
• CLATHRIN
• DYNAMIN
• EEA1
• CD63
• CAVEOLIN -
• LAMP1 -
• independently of the viral coreceptor usage.
• These data suggest that T cell to T cell HIV
  transmission may occur through the
  clathrin-mediated endocytic pathway leading to
  the subsequent release of infectious viral
  particles. Only a small percentage of the
  internalized virus is targeted to the
  lysosomal-degradative pathway.

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Acknowledgements
Jordi Senserrich Bonaventura Clotet José A. Esté
Boyan Grigorov Delphine Muriaux Jean-Luc Darlix


Work funded by European Consortium Targeting
Replication and Integration of HIV