T Cell Responses to SARS-CoV-2.

A large body of evidence generated in the last two and a half years addresses the roles of T cells in SARS-CoV-2 infection and following vaccination. Infection or vaccination induces multi-epitope CD4 and CD8 T cell responses with polyfunctionality. Early T cell responses have been associated with mild COVID-19 outcomes. In concert with animal model data, these results suggest that while antibody responses are key to prevent infection, T cell responses may also play valuable roles in reducing disease severity and controlling infection. T cell memory after vaccination is sustained for at least six months. While neutralizing antibody responses are impacted by SARS-CoV-2 variants, most CD4 and CD8 T cell responses are preserved. This review highlights the extensive progress made, and the data and knowledge gaps that remain, in our understanding of T cell responses to SARS-CoV-2 and COVID-19 vaccines.

Humoral and cellular immune memory to four COVID-19 vaccines.

Multiple COVID-19 vaccines, representing diverse vaccine platforms, successfully protect against symptomatic COVID-19 cases and deaths. Head-to-head comparisons of T cell, B cell, and antibody responses to diverse vaccines in humans are likely to be informative for understanding protective immunity against COVID-19, with particular interest in immune memory. Here, SARS-CoV-2-spike-specific immune responses to Moderna mRNA-1273, Pfizer/BioNTech BNT162b2, Janssen Ad26.COV2.S, and Novavax NVX-CoV2373 were examined longitudinally for 6 months 100% of individuals made memory CD4+ T cells, with cTfh and CD4-CTL highly represented after mRNA or NVX-CoV2373 vaccination. mRNA vaccines and Ad26.COV2.S induced comparable CD8+ T cell frequencies, though only detectable in 60-67% of subjects at 6 months. A differentiating feature of Ad26.COV2.S immunization was a high frequency of CXCR3+ memory B cells. mRNA vaccinees had substantial declines in antibodies, while memory T and B cells were comparatively stable. These results may also be relevant for insights against other pathogens.

SARS-CoV-2 vaccination induces immunological T cell memory able to cross-recognize variants from Alpha to Omicron.

We address whether T cell responses induced by different vaccine platforms (mRNA-1273, BNT162b2, Ad26.COV2.S, and NVX-CoV2373) cross-recognize early SARS-CoV-2 variants. T cell responses to early variants were preserved across vaccine platforms. By contrast, significant overall decreases were observed for memory B cells and neutralizing antibodies. In subjects ∼6 months post-vaccination, 90% (CD4+) and 87% (CD8+) of memory T cell responses were preserved against variants on average by AIM assay, and 84% (CD4+) and 85% (CD8+) preserved against Omicron. Omicron RBD memory B cell recognition was substantially reduced to 42% compared with other variants. T cell epitope repertoire analysis revealed a median of 11 and 10 spike epitopes recognized by CD4+ and CD8+ T cells, with average preservation > 80% for Omicron. Functional preservation of the majority of T cell responses may play an important role as a second-level defense against diverse variants.

Affinity gaps among B cells in germinal centers drive the selection of MPER precursors.

Current prophylactic human immunodeficiency virus 1 (HIV-1) vaccine research aims to elicit broadly neutralizing antibodies (bnAbs). Membrane-proximal external region (MPER)-targeting bnAbs, such as 10E8, provide exceptionally broad neutralization, but some are autoreactive. Here, we generated humanized B cell antigen receptor knock-in mouse models to test whether a series of germline-targeting immunogens could drive MPER-specific precursors toward bnAbs. We found that recruitment of 10E8 precursors to germinal centers (GCs) required a minimum affinity for germline-targeting immunogens, but the GC residency of MPER precursors was brief due to displacement by higher-affinity endogenous B cell competitors. Higher-affinity germline-targeting immunogens extended the GC residency of MPER precursors, but robust long-term GC residency and maturation were only observed for MPER-HuGL18, an MPER precursor clonotype able to close the affinity gap with endogenous B cell competitors in the GC. Thus, germline-targeting immunogens could induce MPER-targeting antibodies, and B cell residency in the GC may be regulated by a precursor-competitor affinity gap.

Adaptive immunity to SARS-CoV-2 and COVID-19.

The adaptive immune system is important for control of most viral infections. The three fundamental components of the adaptive immune system are B cells (the source of antibodies), CD4+ T cells, and CD8+ T cells. The armamentarium of B cells, CD4+ T cells, and CD8+ T cells has differing roles in different viral infections and in vaccines, and thus it is critical to directly study adaptive immunity to SARS-CoV-2 to understand COVID-19. Knowledge is now available on relationships between antigen-specific immune responses and SARS-CoV-2 infection. Although more studies are needed, a picture has begun to emerge that reveals that CD4+ T cells, CD8+ T cells, and neutralizing antibodies all contribute to control of SARS-CoV-2 in both non-hospitalized and hospitalized cases of COVID-19. The specific functions and kinetics of these adaptive immune responses are discussed, as well as their interplay with innate immunity and implications for COVID-19 vaccines and immune memory against re-infection.

Targets of T Cell Responses to SARS-CoV-2 Coronavirus in Humans with COVID-19 Disease and Unexposed Individuals.

Understanding adaptive immunity to SARS-CoV-2 is important for vaccine development, interpreting coronavirus disease 2019 (COVID-19) pathogenesis, and calibration of pandemic control measures. Using HLA class I and II predicted peptide "megapools," circulating SARS-CoV-2-specific CD8+ and CD4+ T cells were identified in ∼70% and 100% of COVID-19 convalescent patients, respectively. CD4+ T cell responses to spike, the main target of most vaccine efforts, were robust and correlated with the magnitude of the anti-SARS-CoV-2 IgG and IgA titers. The M, spike, and N proteins each accounted for 11%-27% of the total CD4+ response, with additional responses commonly targeting nsp3, nsp4, ORF3a, and ORF8, among others. For CD8+ T cells, spike and M were recognized, with at least eight SARS-CoV-2 ORFs targeted. Importantly, we detected SARS-CoV-2-reactive CD4+ T cells in ∼40%-60% of unexposed individuals, suggesting cross-reactive T cell recognition between circulating "common cold" coronaviruses and SARS-CoV-2.

Antigen-Specific Adaptive Immunity to SARS-CoV-2 in Acute COVID-19 and Associations with Age and Disease Severity.

Limited knowledge is available on the relationship between antigen-specific immune responses and COVID-19 disease severity. We completed a combined examination of all three branches of adaptive immunity at the level of SARS-CoV-2-specific CD4+ and CD8+ T cell and neutralizing antibody responses in acute and convalescent subjects. SARS-CoV-2-specific CD4+ and CD8+ T cells were each associated with milder disease. Coordinated SARS-CoV-2-specific adaptive immune responses were associated with milder disease, suggesting roles for both CD4+ and CD8+ T cells in protective immunity in COVID-19. Notably, coordination of SARS-CoV-2 antigen-specific responses was disrupted in individuals ≥ 65 years old. Scarcity of naive T cells was also associated with aging and poor disease outcomes. A parsimonious explanation is that coordinated CD4+ T cell, CD8+ T cell, and antibody responses are protective, but uncoordinated responses frequently fail to control disease, with a connection between aging and impaired adaptive immune responses to SARS-CoV-2.

Slow Delivery Immunization Enhances HIV Neutralizing Antibody and Germinal Center Responses via Modulation of Immunodominance.

Conventional immunization strategies will likely be insufficient for the development of a broadly neutralizing antibody (bnAb) vaccine for HIV or other difficult pathogens because of the immunological hurdles posed, including B cell immunodominance and germinal center (GC) quantity and quality. We found that two independent methods of slow delivery immunization of rhesus monkeys (RMs) resulted in more robust T follicular helper (TFH) cell responses and GC B cells with improved Env-binding, tracked by longitudinal fine needle aspirates. Improved GCs correlated with the development of >20-fold higher titers of autologous nAbs. Using a new RM genomic immunoglobulin locus reference, we identified differential IgV gene use between immunization modalities. Ab mapping demonstrated targeting of immunodominant non-neutralizing epitopes by conventional bolus-immunized animals, whereas slow delivery-immunized animals targeted a more diverse set of epitopes. Thus, alternative immunization strategies can enhance nAb development by altering GCs and modulating the immunodominance of non-neutralizing epitopes.

Follicular helper CD4 T cells (TFH).

T cell help to B cells is a fundamental aspect of adaptive immunity and the generation of immunological memory. Follicular helper CD4 T (T(FH)) cells are the specialized providers of B cell help. T(FH) cells depend on expression of the master regulator transcription factor Bcl6. Distinguishing features of T(FH) cells are the expression of CXCR5, PD-1, SAP (SH2D1A), IL-21, and ICOS, among other molecules, and the absence of Blimp-1 (prdm1). T(FH) cells are important for the formation of germinal centers. Once germinal centers are formed, T(FH) cells are needed to maintain them and to regulate germinal center B cell differentiation into plasma cells and memory B cells. This review covers T(FH) differentiation, T(FH) functions, and human T(FH) cells, discussing recent progress and areas of uncertainty or disagreement in the literature, and it debates the developmental relationship between T(FH) cells and other CD4 T cell subsets (Th1, Th2, Th17, iTreg).

Bcl-6 is the nexus transcription factor of T follicular helper cells via repressor-of-repressor circuits.

T follicular helper (TFH) cells are a distinct type of CD4+ T cells that are essential for most antibody and B lymphocyte responses. TFH cell regulation and dysregulation is involved in a range of diseases. Bcl-6 is the lineage-defining transcription factor of TFH cells and its activity is essential for TFH cell differentiation and function. However, how Bcl-6 controls TFH biology has largely remained unclear, at least in part due to the intrinsic challenges of connecting repressors to gene upregulation in complex cell types with multiple possible differentiation fates. Multiple competing models were tested here by a series of experimental approaches to determine that Bcl-6 exhibits negative autoregulation and controls pleiotropic attributes of TFH differentiation and function, including migration, costimulation, inhibitory receptors and cytokines, via multiple repressor-of-repressor gene circuits.

DNA architectural protein CTCF facilitates subset-specific chromatin interactions to limit the formation of memory CD8+ T cells.

Although the importance of genome organization for transcriptional regulation of cell-fate decisions and function is clear, the changes in chromatin architecture and how these impact effector and memory CD8+ T cell differentiation remain unknown. Using Hi-C, we studied how genome configuration is integrated with CD8+ T cell differentiation during infection and investigated the role of CTCF, a key chromatin remodeler, in modulating CD8+ T cell fates through CTCF knockdown approaches and perturbation of specific CTCF-binding sites. We observed subset-specific changes in chromatin organization and CTCF binding and revealed that weak-affinity CTCF binding promotes terminal differentiation of CD8+ T cells through the regulation of transcriptional programs. Further, patients with de novo CTCF mutations had reduced expression of the terminal-effector genes in peripheral blood lymphocytes. Therefore, in addition to establishing genome architecture, CTCF regulates effector CD8+ T cell heterogeneity through altering interactions that regulate the transcription factor landscape and transcriptome.

Immunogenicity of Stabilized HIV-1 Envelope Trimers with Reduced Exposure of Non-neutralizing Epitopes.

The envelope glycoprotein trimer mediates HIV-1 entry into cells. The trimer is flexible, fluctuating between closed and more open conformations and sometimes sampling the fully open, CD4-bound form. We hypothesized that conformational flexibility and transient exposure of non-neutralizing, immunodominant epitopes could hinder the induction of broadly neutralizing antibodies (bNAbs). We therefore modified soluble Env trimers to stabilize their closed, ground states. The trimer variants were indeed stabilized in the closed conformation, with a reduced ability to undergo receptor-induced conformational changes and a decreased exposure of non-neutralizing V3-directed antibody epitopes. In rabbits, the stabilized trimers induced similar autologous Tier-1B or Tier-2 NAb titers to those elicited by the corresponding wild-type trimers but lower levels of V3-directed Tier-1A NAbs. Stabilized, closed trimers might therefore be useful components of vaccines aimed at inducing bNAbs.

Epigenetic landscapes reveal transcription factors that regulate CD8+ T cell differentiation.

Dynamic changes in the expression of transcription factors (TFs) can influence the specification of distinct CD8+ T cell fates, but the observation of equivalent expression of TFs among differentially fated precursor cells suggests additional underlying mechanisms. Here we profiled the genome-wide histone modifications, open chromatin and gene expression of naive, terminal-effector, memory-precursor and memory CD8+ T cell populations induced during the in vivo response to bacterial infection. Integration of these data suggested that the expression and binding of TFs contributed to the establishment of subset-specific enhancers during differentiation. We developed a new bioinformatics method using the PageRank algorithm to reveal key TFs that influence the generation of effector and memory populations. The TFs YY1 and Nr3c1, both constitutively expressed during CD8+ T cell differentiation, regulated the formation of terminal-effector cell fates and memory-precursor cell fates, respectively. Our data define the epigenetic landscape of differentiation intermediates and facilitate the identification of TFs with previously unappreciated roles in CD8+ T cell differentiation.

Multifaceted Effects of Antigen Valency on B Cell Response Composition and Differentiation In Vivo.

How antigen valency affects B cells in vivo during immune responses is not well understood. Here, using HIV immunogens with defined valencies ranging from 1 to 60, we investigated the role of antigen valency during different phases of B cell responses in vivo. Highly multimerized immunogens preferentially rapidly activated cognate B cells, with little affinity discrimination. This led to strong early induction of the transcription factors IRF4 (interferon regulatory factor 4) and Bcl6, driving both early extrafollicular plasma cell and germinal center responses, in a CD4+ T-cell-dependent manner, involving B cells with a broad range of affinities. Low-valency antigens induced smaller effector B cell responses, with preferential recruitment of high-affinity B cells. Thus, antigen valency has multifaceted effects on B cell responses and can dictate affinity thresholds and competitive landscapes for B cells in vivo, with implications for vaccine design.

Activin A programs the differentiation of human TFH cells.

Follicular helper T cells (TFH cells) are CD4(+) T cells specialized in helping B cells and are associated both with protective antibody responses and autoimmune diseases. The promise of targeting TFH cells therapeutically has been limited by fragmentary understanding of extrinsic signals that regulate the differentiation of human TFH cells. A screen of a human protein library identified activin A as a potent regulator of TFH cell differentiation. Activin A orchestrated the expression of multiple genes associated with the TFH program, independently or in concert with additional signals. TFH cell programming by activin A was antagonized by the cytokine IL-2. Activin A's ability to drive TFH cell differentiation in vitro was conserved in non-human primates but not in mice. Finally, activin-A-induced TFH programming was dependent on signaling via SMAD2 and SMAD3 and was blocked by pharmacological inhibitors.

A TRAF-like motif of the inducible costimulator ICOS controls development of germinal center TFH cells via the kinase TBK1.

Signaling via the inducible costimulator ICOS fuels the stepwise development of follicular helper T cells (TFH cells). However, a signaling pathway unique to ICOS has not been identified. We found here that the kinase TBK1 associated with ICOS via a conserved motif, IProx, that shares homology with the tumor-necrosis-factor receptor (TNFR)-associated factors TRAF2 and TRAF3. Disruption of this motif abolished the association of TBK1 with ICOS, TRAF2 and TRAF3, which identified a TBK1-binding consensus. Alteration of this motif in ICOS or depletion of TBK1 in T cells severely impaired the differentiation of germinal center (GC) TFH cells and the development of GCs, interfered with B cell differentiation and disrupted the development of antibody responses, but the IProx motif and TBK1 were dispensable for the early differentiation of TFH cells. These results reveal a previously unknown ICOS-TBK1 signaling pathway that specifies the commitment of GC TFH cells.

Id2 reinforces TH1 differentiation and inhibits E2A to repress TFH differentiation.

The differentiation of helper T cells into effector subsets is critical to host protection. Transcription factors of the E-protein and Id families are important arbiters of T cell development, but their role in the differentiation of the TH1 and TFH subsets of helper T cells is not well understood. Here, TH1 cells showed more robust Id2 expression than that of TFH cells, and depletion of Id2 via RNA-mediated interference increased the frequency of TFH cells. Furthermore, TH1 differentiation was blocked by Id2 deficiency, which led to E-protein-dependent accumulation of effector cells with mixed characteristics during viral infection and severely impaired the generation of TH1 cells following infection with Toxoplasma gondii. The TFH cell-defining transcriptional repressor Bcl6 bound the Id2 locus, which provides a mechanism for the bimodal Id2 expression and reciprocal development of TH1 cells and TFH cells.