The CD8+ T Cell Noncytotoxic Antiviral Responses

Abstract

The CD8⁺ T cell noncytotoxic antiviral response (CNAR) was discovered during studies of asymptomatic HIV-infected subjects more than 30 years ago. In contrast to CD8⁺ T cell cytotoxic lymphocyte (CTL) activity, CNAR suppresses HIV replication without target cell killing. This activity has characteristics of innate immunity: it acts on all retroviruses and thus is neither epitope specific nor HLA restricted. The HIV-associated CNAR does not affect other virus families. It is mediated, at least in part, by a CD8⁺ T cell antiviral factor (CAF) that blocks HIV transcription. A variety of assays used to measure CNAR/CAF and the effects on other retrovirus infections are described. Notably, CD8⁺ T cell noncytotoxic antiviral responses have now been observed with other virus families but are mediated by different cytokines. Characterizing the protein structure of CAF has been challenging despite many biologic, immunologic, and molecular studies. It represents a low-abundance protein that may be identified by future next-generation sequencing approaches. Since CNAR/CAF is a natural noncytotoxic activity, it could provide promising strategies for HIV/AIDS therapy, cure, and prevention.

Keywords: CD8+ T cells; HIV transcription; elite controllers; human immunodeficiency virus; innate immunity; noncytotoxic antiviral activity; soluble antiviral factor.

FIG 1

Anti-HIV activities of CD8⁺ T cells. To control HIV replication, CD8⁺ T cells can use the following activity: (A) the CD8⁺ T cell noncytotoxic antiviral response (CNAR), which requires cell-cell contact, is improved by the interaction of adhesion molecules to their ligands, and is mediated by a soluble factor; (B) secretion of the CD8⁺ T cell antiviral factor (CAF), which can pass through a semipermeable membrane (transwell) or is found in the culture supernatants of stimulated CD8⁺ T cells; or (C) killing of the infected target cells by cytotoxic CD8⁺ T lymphocytes (CTL) via perforin and granzyme release, which requires degranulation (cell-surface CD107a expression). As opposed to the latter case, CNAR/CAF does not result in target cell death, and instead, HIV transcription is blocked by the CD8⁺ T cells.

FIG 2

Memory phenotype and maturation stages of CD8⁺ T cell subsets. The successive differentiation stages of peripheral CD8⁺ T cells from T_N (naive) are T_CM (central memory), T_TM (transitional memory), T_EM (effector memory), and T_EMRA (effector memory reexpressing CD45RA). In addition, T_RM (resident memory) cells are CD8⁺ T cells found in some tissues. These various CD8⁺ T cell subsets have different expression profiles for the cell surface and intracellular markers listed. In addition, the major CD8⁺ T cell subsets mediating CNAR or CTL are specified.

FIG 3

Tripartite immune cell interactions can induce CNAR/CAF activity. Infected CD4⁺ cells (below)release HIV particles containing conserved retroviral motifs that can be sensed by germ line-encoded invariant pathogen pattern receptors (e.g., TLRs) on accessory cells (e.g., DCs and macrophages). This recognition can (i) upregulate the expression of costimulatory ligands at the surface of the accessory cells (e.g., CD40, and CD80/CD86), and/or (ii) stimulate cytokine secretion (e.g., IL-2, IL-15, and IL-21). These cell surface or secreted molecules can, in turn, activate the subset of CD8⁺ T cells that can mediate CNAR activity by binding to costimulatory receptors (e.g., CD154 and CD28) or cytokine receptors (e.g., CD25, CD215, and CD360). This action would induce or enhance CAF production. CNAR/CAF activity then inhibits HIV transcription, resulting in fewer virus particles and lowered accessory cell (and CD8⁺ T cell) activation. Similar to the effect of ART treatment (31, 162), this action decreases CNAR/CAF activity over time, for as long as HIV is kept in a reduced and/or latent state.

FIG 4

Effect of different (T_h1 and T_h2) cytokines on HIV replication. To determine the effect of cytokines in the CD8⁺ T cell noncytotoxic anti-HIV response (CNAR), CD8⁺ T cells were stimulated in the presence of type 1 (IL-2) or type 2 (IL-4, IL-10) cytokines for 3 days. After being washed, these cells were tested for their ability to suppress HIV replication in acutely infected CD4⁺ lymphocytes. (Modified from references and with permission.)

FIG 5

Level of anti-HIV activity of CAF. To measure the anti-HIV activity of CAF, CD4⁺ cells acutely infected with HIV-1_SF2 were cultured in the presence of various dilutions of CD8⁺ T cell culture supernatants (changed every 2 days) and monitored for viral reverse transcriptase (RT) activity. Examples from one of several experiments are shown. Medium, control; AIDS patients (pts), CD8⁺ T cell culture fluid prepared from an AIDS patient diluted 50%; Asympt, CD8⁺ T cell culture fluid prepared from an asymptomatic HIV seropositive subject diluted 10%, 25%, and 50%. Modified from reference with permission from Elsevier.

FIG 6

Cleavage of a CAF precursor by a protease may be needed for CAF activity. During production of the CD8⁺ T cell antiviral factor (CAF), it is proposed that noncytotoxic CD8⁺ T cells produce a serine protease that cleaves a precursor to CAF into an active moiety. This activated antiviral protein interacts with the HIV-infected CD4⁺ cell surface to induce an anti-HIV state; or, it could enter the cell to produce this response. If the CNAR/CAF-like activity is not blocked by a protease inhibitor (193), the CAF precursor could have already been cleaved into an active anti-HIV protein. (Modified from reference with permission from Elsevier.)

FIG 7

Effect of CNAR and CAF on parameters of HIV replication. The CD8⁺ T cell noncytotoxic antiviral response blocks viral replication, as indicated by decreased RT activity, viral protein expression measured by immunofluorescent antibody (IFA) techniques, and in situ RNA production. This activity has no effect on the number of infected cells in the culture. The antiviral effect is observed as well by a reduction in unspliced (us), single-spliced (ss), and double-spliced (ds) HIV RNA levels compared to a normal expression of β-actin RNA. Finally, the suppressing effect of CD8⁺ T cells or CAF does not affect the basal-level expression of HIV LTR-driven transcription but blocks induction of this transcription by HIV, simian virus 40 tat expression, or phorbol myristate acetate (PMA) using cells in which the HIV LTR has been linked to a reporter gene (section VI). (Modified from reference with permission.)

FIG 8

Effect of CNAR/CAF on transcription elements of the HIV LTR. This schematic representation of the main transcription elements of the HIV LTR shows (i) the relative position of these elements in the HIV promoter; (ii) which transcription elements are present in the HIV-1_SF2 promoter cloned and transduced into the 1G5 cell line used in many studies discussed in this review; and (iii) the potential role of these various transcription elements in CNAR/CAF activity (section VI). References are as follows: Copeland, 1996 (210); Mackewicz, 1995 (44); Bonneau, 2008 (207); Copeland, 1995 (208); Locher, 2001 (209); Shridhar, 2014 (211); Chen, 1993 (204); Powell, 1993 (112).