The persistent pool of HIV-1. Antiretroviral therapy can prevent the creation of new latently infected cells, but it does
not affect cells in which latency was initially established. Intermittent bursts of viremia originate in part from this latent
reservoir. Forcing these cells to exit the latent state without enhancing new infection could make the virus vulnerable
to clearance by an HIV-targeted immune response. Blocking the proliferation of these latently infected T cells could
deplete the pool if its stability is driven by such multiplication.
Expression of HIV from
formerly latently infected cells
Other T cells
( T stem cells or
memory CD4+ T cells
HIV New infection
blocked by ART
have compared neutron and x-ray diffraction data from both species. The authors
first replaced exchangeable protons with
deuterons in large (1 mm) crystals of CcP,
and then treated a CcP crystal with hydrogen peroxide to produce CcP-I. The results
show that the catalytic imidazole of His52 is
not protonated in resting CcP, as expected
(see the figure). In CcP-I, both nitrogen atoms in His52 are protonated (deuterated),
which was unexpected according to the
generally accepted mechanism. This means
that the protons required for O-O bond
cleavage in CcP-0 (see the figure) must have
come from another source, such as an adjacent water molecule. In this snapshot of the
catalytic cycle, the ferryl oxygen, Fe(IV)=O,
of CcP-I is not protonated, and the short
iron-oxygen distance expected for the ferryl
Visualizing the mechanistically pertinent
protons has important implications for the
mechanism of O-O bond scission mediated
by CcP and other heme proteins. A “wet” version of the peroxidase mechanism has been
proposed, in which a water molecule adjacent to His52 mediates O-O bond cleavage (1,
8, 9). Retention of the His52 proton adjacent
to the ferryl heme after O-O bond heterolysis, as revealed by Casadei et al., suggests
that another proton, likely traveling through
an aqueduct of water molecules leading to
the active-site cavity, is also necessary (see
CcP-catalyzed peroxide bond heterolysis thus seems to occur via a proton relay
mechanism similar to that of cytochrome
P450 (10), with electrons arriving through
Trp191 (see the figure). In this scenario, depro-tonation of His52 would occur during subsequent reduction of CcP-I by another enzyme,
ferrocytochrome c. This realization points to
a water-mediated, acid-catalyzed process for
O-O bond heterolysis, which is mechanistically satisfying because of its analogies to
other proton relay mechanisms, such as that
of cytochrome P450. Further, the need for a
water channel and an external proton in peroxidase catalysis are highly informative for
the design and construction of new heme-iron biocatalysts. ■
1. T. L. Poulos, Chem. Rev. 114, 3919 (2014).
2. C. M. Casadei et al ., Science 345, 193 (2014).
3. M. G. Cuypers etal ., Angew.Chem.Int.Ed. 52, 1022 (2013).
4. S.J. Tomanicek et al., J. Biol. Chem. 288,4715(2013).
5. J. T.Groves, J. Inorg. Biochem. 100,434(2006).
6. J. E. Penner-Hahn et al ., J. Am. Chem. Soc. 108, 7819
7. T.H.Yosca et al., Science 342,825(2013).
8. P. Vidossich et al ., J. Phys. Chem. B 114, 5161 (2010).
9. K.Sen, W. Thiel, J. Phys. Chem. B 118,2810(2014).
10. X. Wang, S.Peter, M.Kinne,M.Hofrichter,J. T.Groves,J.
Am. Chem. Soc. 134, 12897 (2012).
10.1126/science.1256754 I L L U
The persistence of HIV-1 infected cells in individuals on antiretroviral therapy (ART) presents an obstacle for cure of infection. ART is the best available remedy for millions of in- fected people, but treatment must
be life-long because HIV establishes latent
infection that is unaffected by antiretrovirals and is invisible to immune surveillance.
Because decades of treatment may be unsustainable, there is intense interest in reversing latency. If quiescent HIV in CD4+ T
cells can be identified and activated without enhancing new infection, HIV-targeted
immune response might be able to control
or even clear infection. On page 179 in this
issue and in this week’s Science Express,
Maldarelli et al. (1) and Wagner et al. (2),
respectively, raise a new challenge for these
efforts suggesting that proliferation of latently infected cells may be a key factor in
sustaining this durable viral reservoir.
Latent HIV proviruses (viral genome in-
tegrated into the host cell DNA) are found
most often in resting CD4+ T cells within the
central memory arm of the immune system
(3–5), although other T cell subpopulations
have been implicated (6, 7). The latent pool
shows minimal or absent decay (8), which is
not fully understood. One possible explana-
tion is that ongoing low-level HIV replication
during ART replenishes the pool. However,
viral genetic diversity does not increase over
time in individuals during ART (9), at odds
with this view.
Other evidence suggests that virus
emerges from the pool of latently infected
cells periodically. Even patients whose viral
load is well suppressed show intermittent
bursts of viremia (“blips”), and in many patients viremia is detectable in specialized assays (10, 11). Given that the pool of latently
infected cells must be primarily established
before ART, it is difficult to understand why
such periodic induction of the pool does not
lead to it running dry.
Homeostatic proliferation of infected
transitional memory T cells (6) has been
proposed as a source that could maintain the
pool, but this does not explain persistence in
the dominant central memory reservoir. Latently infected stem cell–like memory T cells
could proliferate (7), and it will be of great
interest to compare integration patterns
seen in these cells and in more differentiated
Maldarelli et al. and Wagner et al. harvested DNA from the blood cells of HIV-infected individuals after a decade of successful
ART, and analyzed the distribution of sites of
proviral integration in the human genome.
Typically, HIV favors integration in regions of
the genome that are transcriptionally active
(12), but a unique pattern was seen in rare
proviruses from well-suppressed patients.
Both groups found expanded proviral clones
that were enriched for proviruses in or near
a limited set of cellular genes, some of which
Persistence by proliferation?
By David Margolis1 and Frederic Bushman2
Latently HIV-infected cells driven to proliferate may raise a
further challenge for eradication strategies