During a microbial infection, responding CD8+ T cells give rise to effector cells that provide acute host defense and memory cells that provide sustained protection. An alternative outcome is exhaustion, a state of T cell dysfunction that occurs in the context of chronic infections and cancer. Although it is evident that exhausted CD8+ T (TEX) cells are phenotypically and molecularly distinct from effector and memory CD8+ T cells, the factors regulating the earliest events in the differentiation process of TEX cells remain incompletely understood. Here, we performed single-cell RNA-sequencing and single-cell ATAC-sequencing of CD8+ T cells responding to LCMV-Armstrong (LCMV-Arm) or LCMV-Clone 13 (LCMV-Cl13), which result in acute or chronic infections, respectively. Compared to CD8+ T cells that had undergone their first division in response to LCMV-Arm (Div1ARM) cells, CD8+ T cells that had undergone their first division in response to LCMV-Cl13 (Div1CL13) expressed higher levels of genes encoding transcription factors previously associated with exhaustion, along with higher levels of Ezh2, the catalytic component of the Polycomb Repressive Complex 2 (PRC2) complex, which mediates epigenetic silencing. Modulation of Ezh2 resulted in altered expression of exhaustion-associated molecules by CD8+ T cells responding to LCMV-Cl13, though the specific cellular and infectious contexts, rather than simply the level of Ezh2 expression, likely determine the eventual outcome. Taken together, these findings suggest that the differentiation paths of CD8+ T cells responding to acute versus chronic infections may diverge earlier than previously appreciated.

Inflammatory bowel disease (IBD) encompasses a spectrum of gastrointestinal disorders driven by dysregulated immune responses against gut microbiota. We integrated single-cell RNA and antigen receptor sequencing to elucidate key components, cellular states, and clonal relationships of the peripheral and gastrointestinal mucosal immune systems in health and ulcerative colitis (UC). UC was associated with an increase in IgG1+ plasma cells in colonic tissue, increased colonic regulatory T cells characterized by elevated expression of the transcription factor ZEB2, and an enrichment of a γδ T cell subset in the peripheral blood. Moreover, we observed heterogeneity in CD8+ tissue-resident memory T (TRM) cells in colonic tissue, with four transcriptionally distinct states of differentiation observed across health and disease. In the setting of UC, there was a marked shift of clonally related CD8+ TRM cells toward an inflammatory state, mediated, in part, by increased expression of the T-box transcription factor Eomesodermin. Together, these results provide a detailed atlas of transcriptional changes occurring in adaptive immune cells in the context of UC and suggest a role for CD8+ TRM cells in IBD.

Unremitting defense against diverse pathogens and malignancies requires a dynamic and durable immune response. Tissue-resident memory CD8 + T cells (T rm ) afford robust protection against infection and cancer progression through continuous surveillance of non-lymphoid tissues. Here, we provide insight into how T rm confer potent and persistent immunity through partitioning of distinct cellular subsets differing in longevity, effector function, and multipotency. Antigen-specific CD8 + T cells localized to the epithelium of the small intestine are primarily comprised of a shorter-lived effector population most prominent early following both acute viral and bacterial infections, and a longer-lived Id3 hi T rm population that subsequently accumulates at later memory timepoints. We define regulatory gene-programs driving these distinct T rm states, and further clarify roles for Blimp1, T-bet, Id2, and Id3 in supporting and maintaining intestinal T rm heterogeneity during infection. Further, through single-cell RNAseq analysis we demonstrate that tumor-infiltrating lymphocytes broadly differentiate into discrete populations of short-lived and long-lived T rm -like subsets, which share qualities with terminally-exhausted and progenitor-exhausted cells, respectively. As the clinical relevance of T rm continues to widen from acute infections to settings of chronic inflammation and malignancy, clarification of the spectrum of phenotypic and functional states exhibited by CD8 + T cells that reside in non-lymphoid tissues will provide a framework for understanding their regulation and identity in diverse pathophysiological contexts.

During an immune response to microbial infection, CD8(+) T cells give rise to distinct classes of cellular progeny that coordinately mediate clearance of the pathogen and provide long-lasting protection against reinfection, including a subset of noncirculating tissue-resident memory (TRM) cells that mediate potent protection within nonlymphoid tissues. Here, we used single-cell RNA sequencing to examine the gene expression patterns of individual CD8(+) T cells in the spleen and small intestine intraepithelial lymphocyte (siIEL) compartment throughout the course of their differentiation in response to viral infection. These analyses revealed previously unknown transcriptional heterogeneity within the siIEL CD8(+) T cell population at several stages of differentiation, representing functionally distinct TRM cell subsets and a subset of TRM cell precursors within the tissue early in infection. Together, these findings may inform strategies to optimize CD8(+) T cell responses to protect against microbial infection and cancer.

Tissue-resident memory CD8+ T cells (Trm) provide host protection through continuous surveillance of non-lymphoid tissues. Using single-cell RNA-sequencing (scRNA-seq) and genetic reporter mice, we identified discrete lineages of intestinal antigen-specific CD8+ T cells, including a Blimp1hiId3lo tissue-resident effector cell population most prominent in the early phase of acute viral and bacterial infections and a molecularly distinct Blimp1loId3hi tissue-resident memory population that subsequently accumulated at later infection time points. These Trm populations exhibited distinct cytokine production, secondary memory potential, and transcriptional programs including differential roles for transcriptional regulators Blimp1, T-bet, Id2, and Id3 in supporting and maintaining intestinal Trm. Extending our analysis to malignant tissue, we also identified discrete populations of effector-like and memory-like CD8+ T cell populations with tissue-resident gene-expression signatures that shared features of terminally exhausted and progenitor-exhausted T cells, respectively. Our findings provide insight into the development and functional heterogeneity of Trm cells, which has implications for enhancing vaccination and immunotherapy approaches.

In the green lynx spider Peucelia viridans (Hentz 1832), the two openings of a mated female's epigynum are often scaled by copulatory plugs, sometimes with the two-pronged distal portion of the paracybium of a male palpus inserted in each opening and embedded in the plugs. The presence of copulatory Plugs and paracymbia may prevent further mating by the female. However, not all mated females exhibit these structures, perhaps allowing some P. viridans females to mate with more than one male, despite the assertion of Whitcomb & Eason (1965) that females only mate once. We investigated this possibility by surveying the extent of multiple paternity in field-collected P. viridans broods from southern California. For adult females and their egg sacs, we determined the aspartate aminotransferase genotype for each mother and her spiderlings using allozyme electrophoresis in order to assess whether the progeny data best fit with a single male its the father. Two broods exhibited clear evidence of multiple paternity, verifying that multiple mating by females is possible in this species. Although most mothers of single paternity broods had one or both epigynal orifices blocked, sonic had no blockage at all, while the two mothers of multiple paternity broods had some kind of blockage to one or both orifices, Suggesting that neither Plugs nor inserted paracymbial processes are associated with a reduction in female remitting.

We developed a system to encode digital information in DNA polymers based on the partial restriction digest (PRD). Our encoding method relies on the length of the fragments obtained by the PRD rather than the actual content of the nucleotide sequence, thus eliminating the need for expensive sequencing machinery. In this letter, we report on the encoding of 12 bits of data in a DNA fragment of 110 nucleotides and the process of recovering the data.