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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
HIV cell-to-cell transmission
 Coculture of HIV presenting cells (chronically infected MOLT cell lines)
and uninfected cells (primary lymphocytes).
Cellular contacts
Effector cell
Transfer of HIV antigens
Target cell
Untreated
Effector Cells
(MOLT cells)
Trypsin treated
Trypsin treated
NL4-3
42 %
81 %
BaL
Target cells
(T CD4+ lymphocytes)
HIV-1 CA p24
34 %
74 %
HIV-1 cell-to-cell transmission depends on the
interaction between gp120 and CD4
50
Ca p24 + cells
(% of target cells)
45
40
35
No Drug
mAb IgGb12
C-34
30
25
20
15
10
5
0
-
Tryp
Uninfected
MOLT
-
Tryp
HIV-1NL4-3
MOLT
-
Tryp
HIV-1 BaL
MOLT
Internalized HIV resides in vesicular
compartments even in the presence of coreceptor
Ca p24 + cells (% of target cells)
and fusion inhibitors
100
80
No Drug
AMD3100
60
AZT
40
TAK-779
C-34
20
mAb IgGb12
0
Uninfected
MOLT
HIV-1NL4-3
MOLT
HIV-1 BaL
MOLT
Endocytosis
Objectives
 To characterize the subcellular compartments witch contain the
internalized HIV particles.
 To elucidate the fate of the internalized HIV particles.
HIV-1 particles transferred to CD4+ T lymphocytes
contained CD63
CD63 + cells (% of target cells)
80
60
Surface
40
20
0
80
Intracellular
60
40
20
0
No Drug
mAb IgGb12
C-34
Endocyted HIV was later released in the
supernatant as infectious viral particles
Purification of
CD4 T cells
MOLT/BaL +
CD4 T cells (6 h.)
Trypsin
treatment
12 h
Incubation

Infection of
U87.CD4.CCR5.
80000
60000
Infectivity
(Infectious units/ml)
Released CA p24
(pg/ml)
100000
40000
20000
Cell-associated
CA p24 staining (MFI)
0
0
10
20
30
40
50
24
20
16
600
450
300
150
0
Uninfected
12
8
No Drug
4
mAb IgGb12
TAK-779
0
0
10
20
30
Time (hours)
40
50
BaL
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.
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