b Macrophages were treated with different A2ti (25 or 50?M), an anti-A2 antibody (25?g/mL), or maraviroc (25?g/mL) prior to exposure to HIV-1JR-CSF (MOI?=?1). A2 and S100A10 were tested for OICR-0547 their ability to impair productive HIV-1 contamination of macrophages. Our data suggest that interactions between HIV-1 gp120 and A2 exist, though this conversation may be indirect. Furthermore, an anti-A2 antibody impaired Rabbit Polyclonal to Akt HIV-1 particle production in macrophages in vitro, whereas A2ti did not indicating that annexin A2 OICR-0547 may promote HIV-1 contamination of macrophages in its monomeric rather than tetrameric form. strong class=”kwd-title” Keywords: Annexin A2, Annexin A2 heterotetramer, HIV-1, Inhibitor, Macrophage, Receptor Introduction During sexual transmission of human immunodeficiency computer virus (HIV), macrophages of the cervical, anal, and foreskin epithelium are among the first immune cells to encounter the virus, which makes them initial targets for HIV contamination [1, 2]. It is well OICR-0547 established that secretory leukocyte protease inhibitor (SLPI), a protein found in high concentrations in mucosal fluids, protects against HIV-1 contamination of macrophages impartial of its anti-protease activity [3, 4]. Moreover, when the host-cell membrane constituent phospholipid phosphatidylserine (PS) is usually incorporated into the viral envelope during the budding process, it acts as a cofactor for HIV-1 contamination of macrophages . The ability of host-derived PS to influence HIV-1 contamination led to the prediction that an unknown factor on target-cell membranes facilitated viral binding and/or fusion through PS. It was later revealed that SLPI directly interacted with annexin A2 (A2), a PS-binding moiety, and that SLPI could disrupt the conversation between A2 and PS around the HIV-1 envelope to prevent contamination in vitro  (also observe Fig.?1d). Additionally, antibodies against A2 or RNA silencing of A2 significantly inhibited HIV-1 contamination comparable to that of SLPI. It was also shown that A2 is usually involved in HIV-1 replication in monocyte-derived macrophages (MDMs) , and that HIV-1 produced from MDMs that had been treated with A2 siRNA exhibited decreased infectivity . Open in a separate windows Fig. 1 A2 from macrophage lysates is usually captured on HIV-1 gp120-coated SiMPull slides. Lysis buffer (a) or macrophage cell lysates (b) were flowed onto SiMPull slides coated with increasing amounts of biotinylated gp120, and the number of captured complexes (c) were detected following staining with a rabbit anti-A2 antibody and an anti-rabbit 568-conjugated secondary antibody using TIRF microscopy, where each white dot OICR-0547 represents one protein-protein complex (scale bar?=?5?m). Controls included no gp120 and no lysate. Data are offered as the means??SD of five fields of view of a representative example of an experiment performed three times. * em p /em ? ?0.05 ** em p /em ? ?0.01 as determined by a one-way ANOVA followed by a Kruskal-Wallis multiple comparisons test against the no gp120 control group. d In a separate experiment, lysates were flowed onto SiMPull slides coated with an anti-A2 antibody, and captured complexes were detected with mouse anti-S100A10 or anti-SLPI main antibodies and an anti-mouse OICR-0547 568-conjugated secondary antibody. *** em p /em ? ?0.001 as determined by an unpaired two-tailed Students em T /em -test against the no capture control group Generally, HIV-1 infects macrophages through the canonical CD4 receptor CCR5 coreceptor pathway [2, 9], though numerous cofactors can affect the efficiency of this process and the rate of contamination [5, 6]. Access inhibitors, such as the CCR5 antagonist maraviroc , often lead to the emergence of resistant HIV-1 strains that can use alternate pathways . Moreover, option pathways of HIV-1 contamination are likely to differ in macrophages and CD4+ T cells as they express different membrane components such as PS and A2, which are found around the macrophage cell membrane but not on viable T cells [4, 7]. A2 can be found around the cell surface as a heterotetramer (A2t) consisting of two A2 monomers and an S100A10 dimer , which are co-expressed by macrophages . Additionally, data from your HIV-1 Human Conversation Database from your National Center for Biotechnology Information (NCBI) suggests that there may be interactions between HIV-1 gp120 and host A2 , though direct evidence is lacking. Recently, our collaborators developed triazole-based small molecule inhibitors of A2t (A2ti) that specifically disrupt the conversation between A2 and S100A10 , and we showed that these small molecules block contamination of the A2t-utilizing human papillomavirus type 16 (HPV16) , but have yet to be explored in the context of HIV. While A2 has already been implicated in HIV-1.
b Macrophages were treated with different A2ti (25 or 50?M), an anti-A2 antibody (25?g/mL), or maraviroc (25?g/mL) prior to exposure to HIV-1JR-CSF (MOI?=?1)
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