1. PUBLISHABLE SUMMARY
a) Summary description of project objectives
The HIV innate immunity project aims at investigating the antiviral effect of the SAMHD1
protein in dendritic cells (DCs) during HIV infection by combining several virological
and immunological tools.
b) Description of the work performed since the beginning of the project
Dendritic cells are professional antigen-presenting cells linking innate and adaptive
immune responses, however they remain poorly sensitive to HIV-1 infection. SAMHD1 is
a restriction factor that blocks HIV-1 infection by decreasing the levels of deoxy
nucleotide triphosphates (dNTPs) in the cell, which in turns stops the reverse
transcription step required to complete the virus life cycle {Goldstone, #1} {Powell,
#3}. On the other hand, the viral protein Vpx, present in HIV-2 and SIV from macaques,
triggers the destruction of SAMHD1 facilitating the infection of DCs by HIV-1 when Vpx
is supplied inside the cells. HIV-1 is more efficiently transmitted between cells, through
virologycal synapses, than as free viral particles, a mechanism that allows a faster
spread of the infection.
In the first period of the project (2012-2013) we studied whether HIV-1 can be
transmitted from infected lymphocytes to cocultivated myeloid derived dendritic cells
(MDDCs) and examined the effect of SAMHD1 on viral transmission and triggering of the
type I IFN response. Previous reports showed that MDDC were poorly sensitive to HIV-1,
which means that they do not induce a canonical antiviral type I IFN response{Manel,
#2}. We wanted to explore if HIV-1-infected cells are more potent inducers of type I IFN
than free virions, a feature already observed in plasmacytoid DC {Lepelley, #4}.
Generally restriction factors are effective at low multiplicity of infection, so we wished to
determine whether the antiviral effect of SAMHD1 is saturable. We performed Vpx
treatment or silencing of SAMHD1 in MDDCs, to unravel the role of SAMHD1 during
infection and immune detection of HIV.
During the second period (2013-2014) we aimed at studying HIV-2 infection of MDDC,
as its genome encodes for Vpx, the protein that removes SAMHD1 restriction. Previous
reports have failed to observe HIV-2 infection in DCs {Duvall, 2007 #5}, but these results
are in controversy with a recent article that observed HIV-2 infection in DC {Lahaye,
#6}. In order to reconcile this two reports, we have performed coculture experiments
between HIV-2 infected cells and MDDC. The aim is to explore the differences between
HIV-1 and HIV-2 in terms of infection and detection of the viruses by DC, a feature that
could explain the lower pathogenesis of HIV-2 as compared to HIV-1.
c) A description of the main results achieved so far
I have characterized the effect of SAMHD1 during cell-to-cell HIV-1 transmission. To do
so, I have performed coculture experiments between HIV-1 infected T cells and MDDC
(Figure). I have stained the intracellular HIV-1 capsid protein (Gag) and SAMHD1 and
measured the expression by flow cytometry. Data showed that addition of Vpx
significantly enhances HIV-1 transmission from T cells to MDDC. Moreover, silencing of
SAMHD1 in MDDC confirmed the same results suggesting that SAMHD1 plays a role on
the restriction of cell-to-cell HIV transmission to MDDCs . Moreover, I measured the
immune detection of HIV-1 in MDDC by analyzing the intracellular levels of the MxA
interferon-stimulated protein and the release of the antiviral cytokine type I IFN in the
supernatant of the cocultures. These data showed for the first time that SAMHD1 is
active during cell-to-cell HIV-1 transmission.
Afterwards, we measured the infection of MDDc by HIV-2 infected T cells. Data showed
that HIV-2, despite having Vpx, was not able to infect MDDC even under coculture
conditions. Only VSV pseudotyped HIV-2 viruses were able to infect MDDC suggesting
that HIV-2 has an entry block. The poor infection of HIV-2 in MDDC correlates with a low
or absent detection of this virus in terms of type I IFN response. These results
confirmed published data {Duvall, 2007 #5} and showed for the first time that cell-tocell HIV-2 transmission cannot overcome this restriction. However, further studies are
required to explore the HIV-2 blockade in MDDCs.
d) The expected final results and their potential impact and use (including the socioeconomic impact and the wider societal implications of the project so far)
The low sensitivity of MDDC to productive HIV-1 infection has important consequences
on the type of innate immune response awakened in HIV-1 infected patients.
Understanding how the pathways of innate interactions between HIV-1 and different
types of DC operate during pathogenic and nonpathogenic infections may be particularly
relevant for HIV-1 vaccination.
Cell-to-cell HIV transmission is a potent way of viral dissemination and sometimes it is
underestimated in many of the studies performed in vitro. Thus, we aimed at
characterizing this pathway and its importance when it comes to understanding the
intrinsic mechanism of HIV restriction.
The relevance of this work is to be able to design target strategies aimed at eradicating
HIV-1 and boosting the virus that remains hidden in treated patients. Finding a final
cure for HIV-1 patients and a prevention to HIV-1 infection could have a huge impact on
the economical sanitary cost that the globe is facing these days with HIV-1 infected
people specially in countries with a significant AIDS population.
Duval M.G, Lore K, et al. Dendritic cells are less susceptible to HIV-2 infection than to HIV-1 infection.
Journal of virology 2007, 81(24):13486-98
Goldstone D.C, Ennis-Adeniran V, et al. HIV-1 restriction factor SAMHD1 is a deoxynucleoside
triphosphate triphosphohydrolase. Nature, 480(7377):379-82
Powell R.D, Holland P.J, et al. Aircadi-Goutiere syndrome gene and HIV-1 restriction factor SAMHD1
is a dGTP-regulated deoxynucleotide triphosphohydrolase. Journal of biologycal chemistry,
286(51):43596-600
Lahaye X, Satoh T, et al. The capsids of HIV-1 and HIV-2 determine immune detection of the viral
cDNA by the innate sensor cGAS in dendritic cells. Immunity 2014 ,39(6):1132-42
Lepelley A, Louis S, et al. Innate sensing of HIV-infected cells. PLoSPathogens 2011, 7(2)
Manel N, Hogstad B et al. A cryptic sensor for HIV-1 activates antiviral innate immunity in dendritic
cells. Nature 2010, 467(7312): 214-7