Epigenetic memory of coronavirus infection in innate immune cells and their progenitors.

Inflammation can trigger lasting phenotypes in immune and non-immune cells. Whether and how human infections and associated inflammation can form innate immune memory in hematopoietic stem and progenitor cells (HSPC) has remained unclear. We found that circulating HSPC, enriched from peripheral blood, captured the diversity of bone marrow HSPC, enabling investigation of their epigenomic reprogramming following coronavirus disease 2019 (COVID-19). Alterations in innate immune phenotypes and epigenetic programs of HSPC persisted for months to 1 year following severe COVID-19 and were associated with distinct transcription factor (TF) activities, altered regulation of inflammatory programs, and durable increases in myelopoiesis. HSPC epigenomic alterations were conveyed, through differentiation, to progeny innate immune cells. Early activity of IL-6 contributed to these persistent phenotypes in human COVID-19 and a mouse coronavirus infection model. Epigenetic reprogramming of HSPC may underlie altered immune function following infection and be broadly relevant, especially for millions of COVID-19 survivors.

Lrp1 is a host entry factor for Rift Valley fever virus.

Rift Valley fever virus (RVFV) is a zoonotic pathogen with pandemic potential. RVFV entry is mediated by the viral glycoprotein (Gn), but host entry factors remain poorly defined. Our genome-wide CRISPR screen identified low-density lipoprotein receptor-related protein 1 (mouse Lrp1/human LRP1), heat shock protein (Grp94), and receptor-associated protein (RAP) as critical host factors for RVFV infection. RVFV Gn directly binds to specific Lrp1 clusters and is glycosylation independent. Exogenous addition of murine RAP domain 3 (mRAPD3) and anti-Lrp1 antibodies neutralizes RVFV infection in taxonomically diverse cell lines. Mice treated with mRAPD3 and infected with pathogenic RVFV are protected from disease and death. A mutant mRAPD3 that binds Lrp1 weakly failed to protect from RVFV infection. Together, these data support Lrp1 as a host entry factor for RVFV infection and define a new target to limit RVFV infections.

Human circulating and tissue-resident memory CD8+ T cells.

Our current knowledge of human memory CD8+ T cells is derived largely from studies of the intravascular space. However, emerging data are starting to challenge some of the dogmas based on this work, suggesting that a conceptual revision may be necessary. In this review, we provide a brief history of the field and summarize the biology of circulating and tissue-resident memory CD8+ T cells, which are ultimately responsible for effective immune surveillance. We also incorporate recent findings into a biologically integrated model of human memory CD8+ T cell differentiation. Finally, we address how future innovative human studies could improve our understanding of anatomically localized CD8+ T cells to inform the development of more effective immunotherapies and vaccines, the need for which has been emphasized by the global struggle to contain severe acute respiratory syndrome coronavirus 2.

SARS-CoV-2 reservoir in post-acute sequelae of COVID-19 (PASC).

Millions of people are suffering from Long COVID or post-acute sequelae of COVID-19 (PASC). Several biological factors have emerged as potential drivers of PASC pathology. Some individuals with PASC may not fully clear the coronavirus SARS-CoV-2 after acute infection. Instead, replicating virus and/or viral RNA-potentially capable of being translated to produce viral proteins-persist in tissue as a 'reservoir'. This reservoir could modulate host immune responses or release viral proteins into the circulation. Here we review studies that have identified SARS-CoV-2 RNA/protein or immune responses indicative of a SARS-CoV-2 reservoir in PASC samples. Mechanisms by which a SARS-CoV-2 reservoir may contribute to PASC pathology, including coagulation, microbiome and neuroimmune abnormalities, are delineated. We identify research priorities to guide the further study of a SARS-CoV-2 reservoir in PASC, with the goal that clinical trials of antivirals or other therapeutics with potential to clear a SARS-CoV-2 reservoir are accelerated.

Robust T Cell Immunity in Convalescent Individuals with Asymptomatic or Mild COVID-19.

SARS-CoV-2-specific memory T cells will likely prove critical for long-term immune protection against COVID-19. Here, we systematically mapped the functional and phenotypic landscape of SARS-CoV-2-specific T cell responses in unexposed individuals, exposed family members, and individuals with acute or convalescent COVID-19. Acute-phase SARS-CoV-2-specific T cells displayed a highly activated cytotoxic phenotype that correlated with various clinical markers of disease severity, whereas convalescent-phase SARS-CoV-2-specific T cells were polyfunctional and displayed a stem-like memory phenotype. Importantly, SARS-CoV-2-specific T cells were detectable in antibody-seronegative exposed family members and convalescent individuals with a history of asymptomatic and mild COVID-19. Our collective dataset shows that SARS-CoV-2 elicits broadly directed and functionally replete memory T cell responses, suggesting that natural exposure or infection may prevent recurrent episodes of severe COVID-19.

The Identity of Human Tissue-Emigrant CD8+ T Cells.

Lymphocyte migration is essential for adaptive immune surveillance. However, our current understanding of this process is rudimentary, because most human studies have been restricted to immunological analyses of blood and various tissues. To address this knowledge gap, we used an integrated approach to characterize tissue-emigrant lineages in thoracic duct lymph (TDL). The most prevalent immune cells in human and non-human primate efferent lymph were T cells. Cytolytic CD8+ T cell subsets with effector-like epigenetic and transcriptional signatures were clonotypically skewed and selectively confined to the intravascular circulation, whereas non-cytolytic CD8+ T cell subsets with stem-like epigenetic and transcriptional signatures predominated in tissues and TDL. Moreover, these anatomically distinct gene expression profiles were recapitulated within individual clonotypes, suggesting parallel differentiation programs independent of the expressed antigen receptor. Our collective dataset provides an atlas of the migratory immune system and defines the nature of tissue-emigrant CD8+ T cells that recirculate via TDL.

Chronic Inflammation Permanently Reshapes Tissue-Resident Immunity in Celiac Disease.

Tissue-resident lymphocytes play a key role in immune surveillance, but it remains unclear how these inherently stable cell populations respond to chronic inflammation. In the setting of celiac disease (CeD), where exposure to dietary antigen can be controlled, gluten-induced inflammation triggered a profound depletion of naturally occurring Vγ4+/Vδ1+ intraepithelial lymphocytes (IELs) with innate cytolytic properties and specificity for the butyrophilin-like (BTNL) molecules BTNL3/BTNL8. Creation of a new niche with reduced expression of BTNL8 and loss of Vγ4+/Vδ1+ IELs was accompanied by the expansion of gluten-sensitive, interferon-γ-producing Vδ1+ IELs bearing T cell receptors (TCRs) with a shared non-germline-encoded motif that failed to recognize BTNL3/BTNL8. Exclusion of dietary gluten restored BTNL8 expression but was insufficient to reconstitute the physiological Vγ4+/Vδ1+ subset among TCRγδ+ IELs. Collectively, these data show that chronic inflammation permanently reconfigures the tissue-resident TCRγδ+ IEL compartment in CeD. VIDEO ABSTRACT.

Memory-like HCV-specific CD8+ T cells retain a molecular scar after cure of chronic HCV infection.

In chronic hepatitis C virus (HCV) infection, exhausted HCV-specific CD8+ T cells comprise memory-like and terminally exhausted subsets. However, little is known about the molecular profile and fate of these two subsets after the elimination of chronic antigen stimulation by direct-acting antiviral (DAA) therapy. Here, we report a progenitor-progeny relationship between memory-like and terminally exhausted HCV-specific CD8+ T cells via an intermediate subset. Single-cell transcriptomics implicated that memory-like cells are maintained and terminally exhausted cells are lost after DAA-mediated cure, resulting in a memory polarization of the overall HCV-specific CD8+ T cell response. However, an exhausted core signature of memory-like CD8+ T cells was still detectable, including, to a smaller extent, in HCV-specific CD8+ T cells targeting variant epitopes. These results identify a molecular signature of T cell exhaustion that is maintained as a chronic scar in HCV-specific CD8+ T cells even after the cessation of chronic antigen stimulation.

Two subsets of stem-like CD8+ memory T cell progenitors with distinct fate commitments in humans.

T cell memory relies on the generation of antigen-specific progenitors with stem-like properties. However, the identity of these progenitors has remained unclear, precluding a full understanding of the differentiation trajectories that underpin the heterogeneity of antigen-experienced T cells. We used a systematic approach guided by single-cell RNA-sequencing data to map the organizational structure of the human CD8+ memory T cell pool under physiological conditions. We identified two previously unrecognized subsets of clonally, epigenetically, functionally, phenotypically and transcriptionally distinct stem-like CD8+ memory T cells. Progenitors lacking the inhibitory receptors programmed death-1 (PD-1) and T cell immunoreceptor with Ig and ITIM domains (TIGIT) were committed to a functional lineage, whereas progenitors expressing PD-1 and TIGIT were committed to a dysfunctional, exhausted-like lineage. Collectively, these data reveal the existence of parallel differentiation programs in the human CD8+ memory T cell pool, with potentially broad implications for the development of immunotherapies and vaccines.

CD4+ T cell-mediated recognition of a conserved cholesterol-dependent cytolysin epitope generates broad antibacterial immunity.

CD4+ T cell-mediated immunity against Streptococcus pneumoniae (pneumococcus) can protect against recurrent bacterial colonization and invasive pneumococcal diseases (IPDs). Although such immune responses are common, the pertinent antigens have remained elusive. We identified an immunodominant CD4+ T cell epitope derived from pneumolysin (Ply), a member of the bacterial cholesterol-dependent cytolysins (CDCs). This epitope was broadly immunogenic as a consequence of presentation by the pervasive human leukocyte antigen (HLA) allotypes DPB1∗02 and DPB1∗04 and recognition via architecturally diverse T cell receptors (TCRs). Moreover, the immunogenicity of Ply427-444 was underpinned by core residues in the conserved undecapeptide region (ECTGLAWEWWR), enabling cross-recognition of heterologous bacterial pathogens expressing CDCs. Molecular studies further showed that HLA-DP4-Ply427-441 was engaged similarly by private and public TCRs. Collectively, these findings reveal the mechanistic determinants of near-global immune focusing on a trans-phyla bacterial epitope, which could inform ancillary strategies to combat various life-threatening infectious diseases, including IPDs.

IDR-targeting compounds suppress HPV genome replication via disruption of phospho-BRD4 association with DNA damage response factors.

Compounds binding to the bromodomains of bromodomain and extra-terminal (BET) family proteins, particularly BRD4, are promising anticancer agents. Nevertheless, side effects and drug resistance pose significant obstacles in BET-based therapeutics development. Using high-throughput screening of a 200,000-compound library, we identified small molecules targeting a phosphorylated intrinsically disordered region (IDR) of BRD4 that inhibit phospho-BRD4 (pBRD4)-dependent human papillomavirus (HPV) genome replication in HPV-containing keratinocytes. Proteomic profiling identified two DNA damage response factors-53BP1 and BARD1-crucial for differentiation-associated HPV genome amplification. pBRD4-mediated recruitment of 53BP1 and BARD1 to the HPV origin of replication occurs in a spatiotemporal and BRD4 long (BRD4-L) and short (BRD4-S) isoform-specific manner. This recruitment is disrupted by phospho-IDR-targeting compounds with little perturbation of the global transcriptome and BRD4 chromatin landscape. The discovery of these protein-protein interaction inhibitors (PPIi) not only demonstrates the feasibility of developing PPIi against phospho-IDRs but also uncovers antiviral agents targeting an epigenetic regulator essential for virus-host interaction and cancer development.

Memory CD4+ T cells are generated in the human fetal intestine.

The fetus is thought to be protected from exposure to foreign antigens, yet CD45RO+ T cells reside in the fetal intestine. Here we combined functional assays with mass cytometry, single-cell RNA sequencing and high-throughput T cell antigen receptor (TCR) sequencing to characterize the CD4+ T cell compartment in the human fetal intestine. We identified 22 CD4+ T cell clusters, including naive-like, regulatory-like and memory-like subpopulations, which were confirmed and further characterized at the transcriptional level. Memory-like CD4+ T cells had high expression of Ki-67, indicative of cell division, and CD5, a surrogate marker of TCR avidity, and produced the cytokines IFN-γ and IL-2. Pathway analysis revealed a differentiation trajectory associated with cellular activation and proinflammatory effector functions, and TCR repertoire analysis indicated clonal expansions, distinct repertoire characteristics and interconnections between subpopulations of memory-like CD4+ T cells. Imaging mass cytometry indicated that memory-like CD4+ T cells colocalized with antigen-presenting cells. Collectively, these results provide evidence for the generation of memory-like CD4+ T cells in the human fetal intestine that is consistent with exposure to foreign antigens.

Mitochondria in health, disease, and aging.

Mitochondria are well known as organelles responsible for the maintenance of cellular bioenergetics through the production of ATP. Although oxidative phosphorylation may be their most important function, mitochondria are also integral for the synthesis of metabolic precursors, calcium regulation, the production of reactive oxygen species, immune signaling, and apoptosis. Considering the breadth of their responsibilities, mitochondria are fundamental for cellular metabolism and homeostasis. Appreciating this significance, translational medicine has begun to investigate how mitochondrial dysfunction can represent a harbinger of disease. In this review, we provide a detailed overview of mitochondrial metabolism, cellular bioenergetics, mitochondrial dynamics, autophagy, mitochondrial damage-associated molecular patterns, mitochondria-mediated cell death pathways, and how mitochondrial dysfunction at any of these levels is associated with disease pathogenesis. Mitochondria-dependent pathways may thereby represent an attractive therapeutic target for ameliorating human disease.

T cell autoreactivity directed toward CD1c itself rather than toward carried self lipids.

The hallmark function of αß T cell antigen receptors (TCRs) involves the highly specific co-recognition of a major histocompatibility complex molecule and its carried peptide. However, the molecular basis of the interactions of TCRs with the lipid antigen-presenting molecule CD1c is unknown. We identified frequent staining of human T cells with CD1c tetramers across numerous subjects. Whereas TCRs typically show high specificity for antigen, both tetramer binding and autoreactivity occurred with CD1c in complex with numerous, chemically diverse self lipids. Such extreme polyspecificity was attributable to binding of the TCR over the closed surface of CD1c, with the TCR covering the portal where lipids normally protrude. The TCR essentially failed to contact lipids because they were fully seated within CD1c. These data demonstrate the sequestration of lipids within CD1c as a mechanism of autoreactivity and point to small lipid size as a determinant of autoreactive T cell responses.

MYC assembles and stimulates topoisomerases 1 and 2 in a "topoisome".

High-intensity transcription and replication supercoil DNA to levels that can impede or halt these processes. As a potent transcription amplifier and replication accelerator, the proto-oncogene MYC must manage this interfering torsional stress. By comparing gene expression with the recruitment of topoisomerases and MYC to promoters, we surmised a direct association of MYC with topoisomerase 1 (TOP1) and TOP2 that was confirmed in vitro and in cells. Beyond recruiting topoisomerases, MYC directly stimulates their activities. We identify a MYC-nucleated "topoisome" complex that unites TOP1 and TOP2 and increases their levels and activities at promoters, gene bodies, and enhancers. Whether TOP2A or TOP2B is included in the topoisome is dictated by the presence of MYC versus MYCN, respectively. Thus, in vitro and in cells, MYC assembles tools that simplify DNA topology and promote genome function under high output conditions.

Germline bias dictates cross-serotype reactivity in a common dengue-virus-specific CD8+ T cell response.

Adaptive immune responses protect against infection with dengue virus (DENV), yet cross-reactivity with distinct serotypes can precipitate life-threatening clinical disease. We found that clonotypes expressing the T cell antigen receptor (TCR) ß-chain variable region 11 (TRBV11-2) were 'preferentially' activated and mobilized within immunodominant human-leukocyte-antigen-(HLA)-A*11:01-restricted CD8+ T cell populations specific for variants of the nonstructural protein epitope NS3133 that characterize the serotypes DENV1, DENV3 and DENV4. In contrast, the NS3133-DENV2-specific repertoire was largely devoid of such TCRs. Structural analysis of a representative TRBV11-2+ TCR demonstrated that cross-serotype reactivity was governed by unique interplay between the variable antigenic determinant and germline-encoded residues in the second ß-chain complementarity-determining region (CDR2ß). Extensive mutagenesis studies of three distinct TRBV11-2+ TCRs further confirmed that antigen recognition was dependent on key contacts between the serotype-defined peptide and discrete residues in the CDR2ß loop. Collectively, these data reveal an innate-like mode of epitope recognition with potential implications for the outcome of sequential exposure to heterologous DENVs.

RNA polymerase II elongation control.

Regulation of the elongation phase of transcription by RNA polymerase II (Pol II) is utilized extensively to generate the pattern of mRNAs needed to specify cell types and to respond to environmental changes. After Pol II initiates, negative elongation factors cause it to pause in a promoter proximal position. These polymerases are poised to respond to the positive transcription elongation factor P-TEFb, and then enter productive elongation only under the appropriate set of signals to generate full-length properly processed mRNAs. Recent global analyses of Pol II and elongation factors, mechanisms that regulate P-TEFb involving the 7SK small nuclear ribonucleoprotein (snRNP), factors that control both the negative and positive elongation properties of Pol II, and the mRNA processing events that are coupled with elongation are discussed.

A Razor's Edge: Vascular Responses to Acute Inflammatory Lung Injury/Acute Respiratory Distress Syndrome.

Historically considered a metabolically inert cellular layer separating the blood from the underlying tissue, the endothelium is now recognized as a highly dynamic, metabolically active tissue that is critical to organ homeostasis. Under homeostatic conditions, lung endothelial cells (ECs) in healthy subjects are quiescent, promoting vasodilation, platelet disaggregation, and anti-inflammatory mechanisms. In contrast, lung ECs are essential contributors to the pathobiology of acute respiratory distress syndrome (ARDS), as the quiescent endothelium is rapidly and radically altered upon exposure to environmental stressors, infectious pathogens, or endogenous danger signals into an effective and formidable regulator of innate and adaptive immunity. These dramatic perturbations, produced in a tsunami of inflammatory cascade activation, result in paracellular gap formation between lung ECs, sustained lung edema, and multi-organ dysfunction that drives ARDS mortality. The astonishing plasticity of the lung endothelium in negotiating this inflammatory environment and efforts to therapeutically target the aberrant ARDS endothelium are examined in further detail in this review.