CD8+ T cells respond to persistent stimulation during chronic viral infection by stably expressing co-inhibitory receptors and other exhaustion-related molecules. Here we addressed how memory-like CD8+ T (TML) cells, which sustain the immune response to chronic infection thanks to their stem-like properties, adapt to chronic stimulation when they cannot express the co-inhibitory receptor PD-1. We found an increased initial generation and stable long-term persistence of TML cells lacking PD-1 during chronic viral infection. However, these cells had a reduced ability regenerate upon acute restimulation in the context of a recall response. Mechanistically, the lack of PD-1-mediated inhibition was not compensated by an increased expression of other co-inhibitory receptors or exhaustion related molecules. Rather, the absence of PD-1 resulted in a reduced capacity of the TCR to activate TML cells and to express stemness genes including Myb and Klf4. Similar albeit weaker effects on TML cells were noted when PD-1 engagement was transiently interrupted due to anti-PD-L1 treatment. Thus, stem-like CD8+ T cells responding to chronic viral infection adapt to the absence of PD-1-dependent co-inhibitory signals by further reducing TCR-mediated activation signaling, likely to prevent excessive or prolonged stimulation of these cells.

T cell exhaustion is a major obstacle to effective immunotherapy in cancer and chronic infection. Here, we identify the transcription factor IRF8 as a tumor-specific regulator of CD8+ T cell exhaustion. IRF8 is strongly expressed in tumor-reactive CD8+ T cells but not during chronic viral infection. Its expression is induced by TCR signaling and can be suppressed by type I IFN (IFN-I). Sustained IFN-I signaling, a hallmark of chronic infection, correlates with reduced chromatin accessibility at the Irf8 locus and progressive repression of Irf8 expression. In tumor-specific CD8+ T cells, IRF8 overexpression enhanced TOX expression while reducing IFNγ, granzyme B, and TNF production. Conversely, Irf8 deficiency diminished exhaustion, restored effector functions, and improved tumor control. Mechanistically, IRF8 directly binds the Tox locus and promotes its transcription. We further show that additional IRF-family transcription factors contribute similarly to the exhausted T cell program, identifying this transcriptional network as a key regulator of tumor-associated T cell dysfunction.

Brain metastases are the most common brain tumors in patients and are associated with poor prognosis. Investigating the colonization and outgrowth of brain metastases is challenging given the complexity of the organ, tissue sampling difficulty, and limited experimental models. To address this challenge, we empenetrating mCherry tag from labeled tumor cells to neighboring niche cells, using different brain metastasis mouse models. We found that CD206+ macrophages were the most abundant cells taking up the phages uptake and retain the canonical form of mCherry, even without the cell-penetrating portion of the tag. These results identify a specific macrophage subset in the brain that retains tumor-supplied fluo

The formation of memory T cells is a fundamental feature of adaptative immunity, allowing the establishment of long-term protection against pathogens. Although emerging evidence suggests that metabolic reprogramming is crucial for memory T cell differentiation and survival, the underlying mechanisms that drive metabolic rewiring in memory T cells remain unclear. Here, we found that up-regulation of the nuclear receptor peroxisome proliferator–activated receptor β/δ (PPARβ/δ) instructs the metabolic reprogramming that occurs during the establishment of central memory CD8 + T cells. PPARβ/δ-regulated changes included suppression of aerobic glycolysis and enhancement of oxidative metabolism and fatty acid oxidation. Mechanistically, exposure to interleukin-15 and expression of T cell factor 1 facilitated activation of the PPARβ/δ pathway, counteracting apoptosis induced by antigen clearance and metabolic stress. Together, our findings indicate that PPARβ/δ is a master metabolic regulator orchestrating a metabolic switch that may be favorable for T cell longevity.

Current cancer immunotherapy predominately focuses on eliciting type 1 immune responses fighting cancer; however, long-term complete remission remains uncommon(1,2). A pivotal question arises as to whether type 2 immunity can be orchestrated alongside type 1-centric immunotherapy to achieve enduring response against cancer(3,4). Here we show that an interleukin-4 fusion protein (Fc-IL-4), a typical type 2 cytokine, directly acts on CD8+ T cells and enriches functional terminally exhausted CD8+ T (CD8+ T-TE) cells in the tumour. Consequently, Fc-IL-4 enhances antitumour efficacy of type 1 immunity-centric adoptive T cell transfer or immune checkpoint blockade therapies and induces durable remission across several syngeneic and xenograft tumour models. Mechanistically, we discovered that Fc-IL-4 signals through both signal transducer and activator of transcription 6 (STAT6) and mammalian target of rapamycin (mTOR) pathways, augmenting the glycolytic metabolism and the nicotinamide adenine dinucleotide (NAD) concentration of CD8+ T-TE cells in a lactate dehydrogenase A-dependent manner. The metabolic modulation mediated by Fc-IL-4 is indispensable for reinvigorating intratumoural CD8+ T-TE cells. These findings underscore Fc-IL-4 as a potent type 2 cytokine-based immunotherapy that synergizes effectively with type 1 immunity to elicit long-lasting responses against cancer. Our study not only sheds light on the synergy between these two types of immune responses, but also unveils an innovative strategy for advancing next-generation cancer immunotherapy by integrating type 2 immune factors.

Aging compromises hematopoietic and immune system functions, making older adults especially susceptible to hematopoietic failure, infections and tumor development, and thus representing an important medical target for a broad range of diseases. During aging, hematopoietic stem cells (HSCs) lose their blood reconstitution capability and commit preferentially toward the myeloid lineage (myeloid bias)1,2. These processes are accompanied by an aberrant accumulation of mitochondria in HSCs3. The administration of the mitochondrial modulator urolithin A corrects mitochondrial function in HSCs and completely restores the blood reconstitution capability of 'old' HSCs. Moreover, urolithin A-supplemented food restores lymphoid compartments, boosts HSC function and improves the immune response against viral infection in old mice. Altogether our results demonstrate that boosting mitochondrial recycling reverts the aging phenotype in the hematopoietic and immune systems.

It is with deep sadness that we report the passing away of Hugh Robson (Rob) MacDonald on March 16th, 2023, aged 76, following a complicated recovery from a fall last December. Rob was born on September 16th, 1946, and grew up in Willowdale, a neighborhood in Toronto, Canada. After graduating from the University of Toronto (B.Sc. 1968 in Astrophysics), he went on to earn a Ph.D. in Medical Biophysics (1972) with a dissertation on the response of murine lymphocytes to histocompatibility antigens under the direction of Richard G. Miller. He then joined the Swiss Institute for Experimental Cancer Research in Lausanne, Switzerland, where he performed postdoctoral work with Theodore (Teddy) Brunner on the differentiation of cytolytic T cells. Upon his return to Canada in 1975, he took up a joint assistant professorship appointment at the Departments of Bacteriology and Immunology and Therapeutic Radiology of the University of Western Ontario, London, Canada, and performed experimental work in immune-oncology at the Ontario Cancer Treatment and Research Foundation, London, Canada. In 1977, Rob joined the Lausanne Branch of the Ludwig Institute for Cancer Research in Epalinges, Switzerland, where he had a distinguished research career as a developmental T cell immunologist until his retirement in 2015. He became the Associate Director from 1989 and was the Director from 2007–2012. Rob was a pioneer in driving our understanding of how the immune system ensures that T cells are useful and not harmful. He made numerous fundamental discoveries on T cell biology, including T cell development, lineage commitment, repertoire selection, tolerance, and immune memory. He authored more than 430 scientific articles published in peer-reviewed journals that had over 34,000 citations as of 2018. In 1989, he was presented with the prestigious Swiss Max Cloetta Award. In 2001, he was recognized as a Highly Cited Researcher by the Institute for Scientific Information. Some of the key biological insights during his tenure at the Lausanne Branch of the Ludwig Institute came at a time when T cells were defined as thymus-dependent lymphocytes whose function was hard to measure, that were difficult to maintain in vitro, and whose antigen receptors were unknown. By the early 1970s, Jean-Charles Cerottini and Teddy Brunner had devised and painstakingly refined a functional assay to measure T cell-mediated killing using the release of 51chromium by tumor targets. Rob's seminal contributions to further characterize and understand cytolytic T lymphocytes included the long-term growth in vitro of these cells using T cell growth factor (now known as interleukin-2 [IL-2], which was contained in homemade culture supernatants of recently activated T cells), the cloning of cytolytic T lymphocytes, the determination of their precursor frequency, and the demonstration that cytolytic activity was a function of T cells expressing Lyt2 (now known as CD8) (summarized in1MacDonald H.R. Cerottini J.C. Ryser J.E. Maryanski J.L. Taswell C. Widmer M.B. Brunner K. T. Quantitation and cloning of cytolytic T lymphocytes and their precursors. Immunol.Rev. 1980; 51: 93-123https://doi.org/10.1111/j.1600-065x.1980.tb00318.xCrossref Google Scholar). Rob's central break-through discovery in developmental immunology was made possible thanks to his visionary implementation of the technology of fluorescence flow cytometry in Europe, which had just been developed by the Herzenberg lab in Stanford University. He liked to stress how his training in biophysics allowed him to have a good understanding of the principles and mechanics involved in the early bulky fluorescence-actived cell sorting (FACS) instruments. This, together with key reagents generated by Hans Acha-Orbea (Ludwig Institute, Lausanne) and Hans Hengartner (University of Zurich), allowed him to demonstrate that the absence of T cell reactivity to self-antigens (immunological tolerance) is ensured by the physical elimination of self-reactive T cells in the thymus, a process now termed "negative selection."2MacDonald H.R. Schneider R. Lees R.K. Howe R.C. Acha-Orbea H. Festenstein H. Zinkernagel R.M. Hengartner H. T-cell receptor V beta use predicts reactivity and tolerance to Mlsa-encoded antigens. Nature. 1988; 332: 40-50https://doi.org/10.1038/332040a0Crossref Scopus (857) Google Scholar Moreover, he showed that the proper maturation of T cells in the thymus depends on the interaction of their specific T cell receptor with major histocompatibility complex (MHC) antigens expressed by the thymic stroma, a process referred to as "positive selection."3MacDonald H.R. Lees R.K. Schneider R. Zinkernagel R.M. Hengartner H. Positive selection of CD4+ thymocytes controlled by MHC class II gene products.Nature. 1988; 336: 471-473https://doi.org/10.1038/336471a0Crossref PubMed Scopus (157) Google Scholar These seminal findings are part of the foundation of modern immunology. The above discoveries relied on following the fate of T cells with reactivity toward the mysterious minor lymphocyte-stimulating (Mls) antigens. Further work in collaboration with virologist Heidi Diggelmann (Swiss Institute for Cancer Research, ISREC, Lausanne) molecularly identified Mls antigens as mouse mammary tumor virus (MMTV)-derived superantigens and defined the importance of superantigens in MMTV infection. His long-standing interest in T cell development further led to the discovery of unconventional T cells that are characterized by a very restricted T cell receptor repertoire (with Ralph Budd)4Budd R.C. Miescher G.C. Howe R.C. Lees R.K. Bron C. MacDonald H.R. Developmentally regulated expression of T cell receptor beta chain variable domains in immature thymocytes..J Exp Med. 1987; 166: 577-582https://doi.org/10.1084/jem.166.2.577Crossref PubMed Scopus (202) Google Scholar and whose maturation in the thymus requires interactions with hematopoietic rather than stromal MHCs (with T. Ohteki). These unconventional T cells are now known as invariant natural killer T (NKT) cells and understood to play important roles in bridging innate and adaptive immune responses. Finally, to come back to developmental immunology and lineage commitment, together with Freddy Radtke, Anne Wilson, and Michel Aguet, Rob identified the Notch dependent signaling pathway that commits very early thymic precursor cells to the T cell lineage.5Radtke F. Wilson A. Stark G. Bauer M. van Meerwijk J. MacDonald H.R. Aguet M. Deficient T cell fate specification in mice with an induced inactivation of Notch1. Immunity. 1999; 10: 547-558https://doi.org/10.1016/s1074-7613(00)80054-0Abstract Full Text Full Text PDF Google Scholar Rob was loved by the scientific community for his bright intellect, phenomenal memory, great sense of humor, eclectic spirit, insatiable curiosity, and his trademark humility. Rob's sharp and critical thinking about results and manuscripts alike was invariably delivered to trainees and peers in measured, nuanced, and balanced critiques. He was always engaged in supporting his trainees and collaborators. He both fostered and relied on the quiet and diligent laboratory management and unfailing experimental support of Rosemary Lees throughout his tenure at the Ludwig Institute. He leaves a large alumni family, many of whom are leading research in immunology at centers of academic excellence worldwide. He is remembered by his beloved wife, Lana; his sisters, Chris and Kathy; his brothers-in-law, John and Peter; his sister-in-law, Rita; and his many nieces and nephews and their families.

Immunogenic non-replicative recombinant modified vaccinia virus Ankara (rMVA) has been developed to turn "cold" tumors into "hot" tumors. MVAΔE5R-hFlt3L-mOX40L is our first-generation rMVA by deleting a cGAS (cyclic GMP-AMP synthase) inhibitor encoded by the vaccinia E5R gene and inserting human Flt3L (a growth factor for dendritic cells) and murine OX40L (a co-stimulatory molecule for T cells). In this study, we used a combined single-cell RNA sequencing (scRNA-seq) and single-cell TCR sequencing (scTCR-seq) strategy to elucidate how IT rMVA affects the transcriptome and proliferation of CD8+ T cells in the injected tumors. We performed scRNA-seq and scTCR-seq of sorted CD8+ T cells from the B16-F10 murine melanomas treated with IT rMVA from C57BL/6J mice and identified six clusters of intratumoral CD8+ T cells, including Tcf7+CD62L+ (C0), Tcf7+CD62L- (C1), Proliferating Toxint (C2), effector-like Toxlow (C3), effector-like Toxint (C4), and exhausted Toxhi (C5). IT rMVA resulted in the reduction of stem-like CD8+ T cells (C0 and C1) and exhausted CD8+ T cells (C5) clusters and the expansion of proliferating and effector-like CD8+ T cells (C2, C3, and C4). scTCR-seq of CD8+ T cells revealed that in the PBS mock-treated tumors, the most expanded TCR clones were in C5, whereas in rMVA-treated tumors, C2 and C4 had the highest TCR clonal expansion. Depleting Tcf7+ stem-like T cells resulted in the reduction of intratumoral CD8+ T cells and antitumor effects in response to rMVA treatment. These results demonstrate that IT rMVA promotes tumor-infiltrating Tcf7+ stem-like CD8+T cell differentiation, expansion, and activation.

Persistent exposure to antigen during chronic infection or cancer renders T cells dysfunctional. The molecular mechanisms regulating this state of exhaustion are thought to be common in infection and cancer, despite obvious differences in their microenvironments. Here we found that NFAT5, an NFAT family transcription factor that lacks an AP-1 docking site, was highly expressed in exhausted CD8+ T cells in the context of chronic infections and tumors but was selectively required in tumor-induced CD8+ T cell exhaustion. Overexpression of NFAT5 in CD8+ T cells reduced tumor control, while deletion of NFAT5 improved tumor control by promoting the accumulation of tumor-specific CD8+ T cells that had reduced expression of the exhaustion-associated proteins TOX and PD-1 and produced more cytokines, such as IFNɣ and TNF, than cells with wild-type levels of NFAT5, specifically in the precursor exhausted PD–1+TCF1+TIM–3–CD8+ T cell population. NFAT5 did not promote T cell exhaustion during chronic infection with clone 13 of lymphocytic choriomeningitis virus. Expression of NFAT5 was induced by TCR triggering, but its transcriptional activity was specific to the tumor microenvironment and required hyperosmolarity. Thus, NFAT5 promoted the exhaustion of CD8+ T cells in a tumor-selective fashion. Verdeil and colleagues show that the transcription factor NFAT5 is selectively required in tumor-induced, but not chronic infection-induced, CD8+ T cell exhaustion, possibly due to the modulation of NFAT5 activation by hyperosmolarity in the tumor environment.

In response to infection, naïve CD8+ T (TN) cells yield a large pool of short-lived terminal effector (TTE) cells that eliminate infected host cells. In parallel, a minor population of stem cell-like central memory (TCM) cells forms, which has the capacity to maintain immunity after pathogen clearance. It has remained uncertain whether stem-like TCM cells arise by dedifferentiation from a subset of cytolytic TTE cells or whether priming generates stem-like cells capable of seeding the TCM compartment and, if so, when cytolytic TTE cells branch off. Here, we show that CD8+ T cells with stem-like properties, which are identified by the expression of TCF1 (encoded by Tcf7), are present across the primary response to infection. Priming programs TN cells to undergo multiple cell divisions, over the course of which TCF1 expression is maintained. These TCF1+ cells further expand relatively independently of systemic inflammation, antigen dose, or affinity, and they quantitatively yield TCF1+ TCM cells after pathogen clearance. Inflammatory signals suppress TCF1 expression in early divided TCF1+ cells. TCF1 down-regulation is associated with the irreversible loss of self-renewal capacity and the silencing of stem/memory genes, which precedes the stable acquisition of a TTE state. TCF1 expression restrains cell cycling, explaining in part the limited expansion of TCF1+ relative to TCF1- cells during the primary response. Thus, our data are consistent with terminal differentiation of effector cells being a step-wise process that is initiated by inflammation in primed stem-like cells, which would otherwise become central memory cells by default.

In allogeneic hematopoietic stem cell transplantation, donor αβ T cells attack recipient tissues, causing graft-versus-host disease (GVHD), a major cause of morbidity and mortality. A central question has been how GVHD is sustained despite T cell exhaustion from chronic antigen stimulation. The current model for GVHD holds that disease is maintained through the continued recruitment of alloreactive effectors from blood into affected tissues. Here, we show, using multiple approaches including parabiosis of mice with GVHD, that GVHD is instead primarily maintained locally within diseased tissues. By tracking 1,203 alloreactive T cell clones, we fitted a mathematical model predicting that within each tissue a small number of progenitor T cells maintain a larger effector pool. Consistent with this, we identified a tissue-resident TCF-1+ subpopulation that preferentially engrafted, expanded, and differentiated into effectors upon adoptive transfer. These results suggest that therapies targeting affected tissues and progenitor T cells within them would be effective.

Background: Hematopoietic stem cells (HSCs) generate all blood lineages and ensure the correct homeostasis between myeloid and lymphoid lineages during the entire life of an organism. However, HSCs decrease their regenerative potential and undergo toward a myeloid-biased fate skewing with age. This phenomenon leads to a reduction of B and T cell compartments and immune dysfunction. Consequently, elderly patients are more susceptible to severe viral and fungal infections. It was reported that HSC aging is associated with defective autophagy and accumulation of damaged mitochondria. And emerging studies showed that dysregulation of mitochondria metabolism correlates closely with hematopoietic aging process. However, the clear causal relationship between mitochondrial metabolism and HSC aging remains unknown. Aims: We were wondering if metabolic modulator can rejuvenate aged HSCs, rescue loss-of-function of aged HSCs and improve overall immune response. Methods: We used mito-QC reporter mice and mitochondrial stain to analyze mitochondrial network upon in vitro treatment of metabolic modulator. In vivo long-term blood reconstitution assays and LCMV infection model were used to estimate the effect of metabolic modulator on HSC functionality. Results: Here we reported that interventions aimed to modulate mitochondria recycling and metabolism can rejuvenate aged HSCs and thus preserving the lymphoid compartments and immune function. In vitro administration of the mitophagy inducer Urolithin A on aged HSCs restore their blood and immune reconstitution capability. Moreover, Urolithin A supplemented in the mouse diet, improves bone marrow functionality, expands the lymphoid compartment and boost immune response against viral infection in old mice. Finally, we discovered that HSCs rejuvenation is associated with mitophagy induction and re-establishment of mitochondria homeostasis and fitness in aged HSCs. Summary/Conclusion: In summary, we identified mitophagy as potent regulator of aging process and as valuable target for rejuvenating approaches in HSCs.

Tumor-infiltrated T cells with stem-cell-like properties are important for determining the immunotherapy response. In this issue of Cancer Cell, Asrir and colleagues show that their entry requires specialized tumor-associated endothelial cells that resemble immature and inflamed lymph node vessels and that immunotherapy enhances the recruitment capacity of these endothelial cells.

Ankylosing Spondylitis is a debilitating chronic arthropathy that affects multiple joints. Anaesthesiologists face significant challenges when dealing with the airway implications of this disease, especially when it is unanticipated that a difficult airway may be encountered. This case describes a 42-year-old trauma victim who required an emergency denitive trachea at his ward. Ankylosing Spondylitis and complex airway anatomy led to his intubation failure. He was eventually given an emergency surgical tracheostomy. The successful management of a difficult airway was possible thanks to the appropriate use of modern airway adjuncts as well as workplace soft skills.

Persistent exposure to antigen during chronic infection or cancer renders T cells dysfunctional. The molecular mechanisms regulating this state of exhaustion are thought to be common in infection and cancer, despite obvious differences in their microenvironments. We discovered that NFAT5, an NFAT family member lacking an AP-1 docking site, is highly expressed in exhausted T cells from murine and human tumors and is a central player in tumor-induced exhaustion. While NFAT5 overexpression reduced tumor control, NFAT5 deletion improved tumor control by promoting the accumulation of tumor-specific CD8+ T cells that expressed less TOX and PD-1 and produced more cytokines particularly among precursor exhausted cells. Conversely, NFAT5 had no effect on chronic infection-induced T cell exhaustion. Mechanistically we found that TCR triggering induced NFAT5 expression and that hyperosmolarity stimulated transcriptional activity of NFAT5. We propose that NFAT5 takes over NFAT1/2 to promote exhaustion specifically in tumor-infiltrating CD8+ T cells.

T cell exhaustion presents one of the major hurdles to cancer immunotherapy. Among exhausted CD8+ tumor-infiltrating lymphocytes, the terminally exhausted subset contributes directly to tumor cell killing owing to its cytotoxic effector function. However, this subset does not respond to immune checkpoint blockades and is difficult to be reinvigorated with restored proliferative capacity. Here, we show that a half-life-extended interleukin-10–Fc fusion protein directly and potently enhanced expansion and effector function of terminally exhausted CD8+ tumor-infiltrating lymphocytes by promoting oxidative phosphorylation, a process that was independent of the progenitor exhausted T cells. Interleukin-10–Fc was a safe and highly efficient metabolic intervention that synergized with adoptive T cell transfer immunotherapy, leading to eradication of established solid tumors and durable cures in the majority of treated mice. These findings show that metabolic reprogramming by upregulating mitochondrial pyruvate carrier-dependent oxidative phosphorylation can revitalize terminally exhausted T cells and enhance the response to cancer immunotherapy. Tang and colleagues show that a half-life-extended IL-10–Fc fusion protein acts directly on terminally exhausted PD1+TIM-3+CD8+ T cells to enhance their proliferation and effector function by reprogramming the cellular metabolism to oxidative phosphorylation in a mitochondrial pyruvate carrier–dependent manner. Treatment of tumor-bearing mice with IL-10–Fc and adoptive T cell therapy led to eradication of their established solid tumors and durable cures.

Adoptive cell immunotherapy using in vitro expanded autologous tumor-infiltrating lymphocytes has the potential to mediate durable remission of certain types of cancer. A recent paper in Science shows that complete and durable control of metastatic melanoma requires the infusion of tumor-specific CD8(+) T cells that have stem-cell-like properties.

Virus-specific PD1(+) Tcf1(+) memory-like CD8(+) T cells (TMLs) maintain the CD8(+) T cell response during chronic viral infection. However, the fate of these cells following cessation of persistent antigen exposure has been unclear. Here, we find that TMLs persist upon transfer into antigen-free hosts and form memory following recall stimulation. Phenotypic, functional, and transcriptome analyses show that TML-derived memory cells resemble those arising in response to acute, resolved infection, but they retain features of chronically stimulated cells, including elevated PD-1 and Tox and reduced cytokine expression. This chronic infection imprint is largely accounted for by constitutive Tox expression. Virus-specific Tcf1(+) CD8(+) T cells that persist after clearance of systemic infection also display a chronic infection imprint. Notwithstanding, renewed virus exposure induces a recall response, which controls virus infection in part. Thus, cessation of chronic antigen exposure yields a memory CD8(+) T cell compartment that reflects prior stimulation.

Cytomegalovirus-based vaccine vectors offer interesting opportunities for T cell-based vaccination purposes as CMV infection induces large numbers of functional effector-like cells that accumulate in peripheral tissues, a process termed memory inflation. Maintenance of high numbers of peripheral CD8 T cells requires continuous replenishment of the inflationary T cell pool. Here, we show that the inflationary T cell population contains a small subset of cells expressing the transcription factor Tcf1. These Tcf1 + cells resemble central memory T cells and are proliferation competent. Upon sensing viral reactivation events, Tcf1 + cells feed into the pool of peripheral Tcf1 − cells and depletion of Tcf1 + cells hampers memory inflation. TCR repertoires of Tcf1 + and Tcf1 − populations largely overlap, with the Tcf1 + population showing higher clonal diversity. These data show that Tcf1 + cells are necessary for sustaining the inflationary T cell response, and upholding this subset is likely critical for the success of CMV-based vaccination approaches.