Polyomavirus Wakes Up and Chooses Neurovirulence.

JC polyomavirus (JCPyV) is a human-specific polyomavirus that establishes a silent lifelong infection in multiple peripheral organs, predominantly those of the urinary tract, of immunocompetent individuals. In immunocompromised settings, however, JCPyV can infiltrate the central nervous system (CNS), where it causes several encephalopathies of high morbidity and mortality. JCPyV-induced progressive multifocal leukoencephalopathy (PML), a devastating demyelinating brain disease, was an AIDS-defining illness before antiretroviral therapy that has "reemerged" as a complication of immunomodulating and chemotherapeutic agents. No effective anti-polyomavirus therapeutics are currently available. How depressed immune status sets the stage for JCPyV resurgence in the urinary tract, how the virus evades pre-existing antiviral antibodies to become viremic, and where/how it enters the CNS are incompletely understood. Addressing these questions requires a tractable animal model of JCPyV CNS infection. Although no animal model can replicate all aspects of any human disease, mouse polyomavirus (MuPyV) in mice and JCPyV in humans share key features of peripheral and CNS infection and antiviral immunity. In this review, we discuss the evidence suggesting how JCPyV migrates from the periphery to the CNS, innate and adaptive immune responses to polyomavirus infection, and how the MuPyV-mouse model provides insights into the pathogenesis of JCPyV CNS disease.

Merkel Cell Polyomavirus Large T Antigen Induces Cellular Senescence for Host Growth Arrest and Viral Genome Persistence through Its Unique Domain.

Senescent cells accumulate in the host during the aging process and are associated with age-related pathogeneses, including cancer. Although persistent senescence seems to contribute to many aspects of cellular pathways and homeostasis, the role of senescence in virus-induced human cancer is not well understood. Merkel cell carcinoma (MCC) is an aggressive skin cancer induced by a life-long human infection of Merkel cell polyomavirus (MCPyV). Here, we show that MCPyV large T (LT) antigen expression in human skin fibroblasts causes a novel nucleolar stress response, followed by p21-dependent senescence and senescence-associated secretory phenotypes (SASPs), which are required for MCPyV genome maintenance. Senolytic and navitoclax treatments result in decreased senescence and MCPyV genome levels, suggesting a potential therapeutic for MCC prevention. Our results uncover the mechanism of a host stress response regulating human polyomavirus genome maintenance in viral persistency, which may lead to targeted intervention for MCC.

T cell deficiency precipitates antibody evasion and emergence of neurovirulent polyomavirus.

JC polyomavirus (JCPyV) causes progressive multifocal leukoencephalopathy (PML), a life-threatening brain disease in immunocompromised patients. Inherited and acquired T cell deficiencies are associated with PML. The incidence of PML is increasing with the introduction of new immunomodulatory agents, several of which target T cells or B cells. PML patients often carry mutations in the JCPyV VP1 capsid protein, which confer resistance to neutralizing VP1 antibodies (Ab). Polyomaviruses (PyV) are tightly species-specific; the absence of tractable animal models has handicapped understanding PyV pathogenesis. Using mouse polyomavirus (MuPyV), we found that T cell deficiency during persistent infection, in the setting of monospecific VP1 Ab, was required for outgrowth of VP1 Ab-escape viral variants. CD4 T cells were primarily responsible for limiting polyomavirus infection in the kidney, a major reservoir of persistent infection by both JCPyV and MuPyV, and checking emergence of these mutant viruses. T cells also provided a second line of defense by controlling the outgrowth of VP1 mutant viruses that evaded Ab neutralization. A virus with two capsid mutations, one conferring Ab-escape yet impaired infectivity and a second compensatory mutation, yielded a highly neurovirulent variant. These findings link T cell deficiency and evolution of Ab-escape polyomavirus VP1 variants with neuropathogenicity.

Understanding polyomavirus CNS disease - a perspective from mouse models.

JC polyomavirus (JCPyV), a ubiquitous human pathogen, causes several devastating brain diseases in immune-compromised individuals. The most notable of these JCPyV-associated CNS diseases is the frequently fatal demyelinating brain disease progressive multifocal leukoencephalopathy (PML). PML, an AIDS-defining disease in the pre-cART epoch, has emerged as a life-threatening complication in patients receiving immunomodulatory agents for autoimmune and inflammatory disorders and treatment for certain hematological malignancies. Among the rapidly expanding list of PML-associated biologics, natalizumab (Tysabri®) has the highest incidence and is an ominous sequela for multiple sclerosis (MS) patients who otherwise benefit from dramatic reductions in relapses using this immunomodulatory agent. Drug withdrawal, the only therapeutic option for PML, is often complicated by a high-mortality cerebral inflammatory reaction. No anti-JCPyV agents are available. Lack of a tractable animal model of polyomavirus-induced central nervous system (CNS) disease is an acknowledged bottleneck to elucidating PML pathogenesis, immunological mechanisms that control JCPyV, in vivo evaluation of agents that inhibit polyomavirus replication in tissue culture, and uncovering early events that presage JCPyV-associated neuropathology. The natural virus-host mouse polyomavirus (MuPyV) model has recently been developed to explore mechanisms of polyomavirus-associated CNS disease. In this review, we will cover the benefits of using the MuPyV model to answer fundamental questions about innate and adaptive immune control of JCPyV, the impact of immunomodulation on JCPyV pathogenesis, and how this MuPyV CNS infection model will help improve criteria for identifying patients at risk for JCPyV-associated CNS diseases before the development of irreversible lesions.

New developments implicating IL-21 in autoimmune disease.

Elevated interleukin (IL)-21 is a common finding in the tissues and/or sera of patients with autoimmune disease. CD4 T cells are the primary producers of IL-21; often the IL-21 producing CD4 T cells will express molecules associated with follicular helper cells (TFH). Recent work has shown that the CD4 T cell-derived IL-21 is able to promote effector functions and memory differentiation of CD8 T cells in chronic infections and cancer. Autoimmunity has similarities to chronic infections and cancer. However, CD4 T cell-derived IL-21:IL21R signaling in CD8 T cells has not been fully appreciated in the context of autoimmunity. In this review, we assess the current knowledge regarding CD4 T cell-derived IL-21 and IL21R signaling within CD8 T cells and evaluate what implications it has within several autoimmune diseases including systemic lupus erythematous, rheumatoid arthritis, juvenile idiopathic arthritis, type 1 diabetes mellitus, psoriasis, Sjögren's syndrome, vitiligo, antiphospholipid syndrome, pemphigus, and giant cell arteritis.

JCPyV VP1 Mutations in Progressive MultifocalLeukoencephalopathy: Altering Tropismor Mediating Immune Evasion?

Polyomaviruses are ubiquitous human pathogens that cause lifelong, asymptomatic infections in healthy individuals. Although these viruses are restrained by an intact immune system, immunocompromised individuals are at risk for developing severe diseases driven by resurgent viral replication. In particular, loss of immune control over JC polyomavirus can lead to the development of the demyelinating brain disease progressive multifocal leukoencephalopathy (PML). Viral isolates from PML patients frequently carry point mutations in the major capsid protein, VP1, which mediates virion binding to cellular glycan receptors. Because polyomaviruses are non-enveloped, VP1 is also the target of the host's neutralizing antibody response. Thus, VP1 mutations could affect tropism and/or recognition by polyomavirus-specific antibodies. How these mutations predispose susceptible individuals to PML and other JCPyV-associated CNS diseases remains to be fully elucidated. Here, we review the current understanding of polyomavirus capsid mutations and their effects on viral tropism, immune evasion, and virulence.

IL-21 in Homeostasis of Resident Memory and Exhausted CD8 T Cells during Persistent Infection.

CD4 T cells guide the development of CD8 T cells into memory by elaborating mitogenic and differentiation factors and by licensing professional antigen-presenting cells. CD4 T cells also act to stave off CD8 T cell dysfunction during repetitive antigen stimulation in persistent infection and cancer by mitigating generation of exhausted T cells (TEX). CD4 T cell help is also required for establishing and maintaining tissue-resident memory T cells (TRM), the nonrecirculating memory T cell subset parked in nonlymphoid tissues to provide frontline defense against reinvading pathogens. Interleukin (IL)-21 is the signature cytokine secreted by follicular helper CD4 T cells (TFH) to drive B cell expansion and differentiation in germinal centers to mount high-affinity, isotype class-switched antibodies. In several infection models, IL-21 has been identified as the CD4 T help needed for formation and survival of TRM and TEX. In this review, we will explore the different memory subsets of CD8 T cells in persistent infections, the metabolic profiles associated with each, and evidence documenting the importance of CD4 T cell-derived IL-21 in regulating CD8 TRM and TEX development, homeostasis, and function.

Antibody escape by polyomavirus capsid mutation facilitates neurovirulence.

JCPyV polyomavirus, a member of the human virome, causes progressive multifocal leukoencephalopathy (PML), an oft-fatal demyelinating brain disease in individuals receiving immunomodulatory therapies. Mutations in the major viral capsid protein, VP1, are common in JCPyV from PML patients (JCPyV-PML) but whether they confer neurovirulence or escape from virus-neutralizing antibody (nAb) in vivo is unknown. A mouse polyomavirus (MuPyV) with a sequence-equivalent JCPyV-PML VP1 mutation replicated poorly in the kidney, a major reservoir for JCPyV persistence, but retained the CNS infectivity, cell tropism, and neuropathology of the parental virus. This mutation rendered MuPyV resistant to a monoclonal Ab (mAb), whose specificity overlapped the endogenous anti-VP1 response. Using cryo-EM and a custom sub-particle refinement approach, we resolved an MuPyV:Fab complex map to 3.2 Å resolution. The structure revealed the mechanism of mAb evasion. Our findings demonstrate convergence between nAb evasion and CNS neurovirulence in vivo by a frequent JCPyV-PML VP1 mutation.

IL-21 from high-affinity CD4 T cells drives differentiation of brain-resident CD8 T cells during persistent viral infection.

Development of tissue-resident memory (TRM) CD8 T cells depends on CD4 T cells. In polyomavirus central nervous system infection, brain CXCR5hi PD-1hi CD4 T cells produce interleukin-21 (IL-21), and CD8 T cells lacking IL-21 receptors (IL21R-/-) fail to become bTRM IL-21+ CD4 T cells exhibit elevated T cell receptor (TCR) affinity and higher TCR density. IL21R-/- brain CD8 T cells do not express CD103, depend on vascular CD8 T cells for maintenance, are antigen recall defective, and lack TRM core signature genes. CD4 T cell-deficient and IL21R-/- brain CD8 T cells show similar deficiencies in expression of genes for oxidative metabolism, and intrathecal delivery of IL-21 to CD4 T cell-depleted mice restores expression of electron transport genes in CD8 T cells to wild-type levels. Thus, high-affinity CXCR5hi PD-1hi CD4 T cells in the brain produce IL-21, which drives CD8 bTRM differentiation in response to a persistent viral infection.

Serine Phosphorylation of the STAT1 Transactivation Domain Promotes Autoreactive B Cell and Systemic Autoimmunity Development.

Although STAT1 tyrosine-701 phosphorylation (designated STAT1-pY701) is indispensable for STAT1 function, the requirement for STAT1 serine-727 phosphorylation (designated STAT1-pS727) during systemic autoimmune and antipathogen responses remains unclear. Using autoimmune-prone B6.Sle1b mice expressing a STAT1-S727A mutant in which serine is replaced by alanine, we report in this study that STAT1-pS727 promotes autoimmune Ab-forming cell (AFC) and germinal center (GC) responses, driving autoantibody production and systemic lupus erythematosus (SLE) development. In contrast, STAT1-pS727 is not required for GC, T follicular helper cell (Tfh), and Ab responses to various foreign Ags, including pathogens. STAT1-pS727 is also not required for gut microbiota and dietary Ag-driven GC and Tfh responses in B6.Sle1b mice. By generating B cell-specific bone marrow chimeras, we demonstrate that STAT1-pS727 plays an important B cell-intrinsic role in promoting autoimmune AFC, GC, and Tfh responses, leading to SLE-associated autoantibody production. Our analysis of the TLR7-accelerated B6.Sle1b.Yaa SLE disease model expressing a STAT1-S727A mutant reveals STAT1-pS727-mediated regulation of autoimmune AFC and GC responses and lupus nephritis development. Together, we identify previously unrecognized differential regulation of systemic autoimmune and antipathogen responses by STAT1-pS727. Our data implicate STAT1-pS727 as a therapeutic target for SLE without overtly affecting STAT1-mediated protection against pathogenic infections.

CD8 T Cells and STAT1 Signaling Are Essential Codeterminants in Protection from Polyomavirus Encephalopathy.

JC polyomavirus (JCPyV), a human-specific virus, causes the aggressive brain-demyelinating disease progressive multifocal leukoencephalopathy (PML) in individuals with depressed immune status. The increasing incidence of PML in patients receiving immunotherapeutic and chemotherapeutic agents creates a pressing clinical need to define biomarkers to stratify PML risk and develop anti-JCPyV interventions. Mouse polyomavirus (MuPyV) CNS infection causes encephalopathology and may provide insight into JCPyV-PML pathogenesis. Type I, II, and III interferons (IFNs), which all signal via the STAT1 transcription factor, mediate innate and adaptive immune defense against a variety of viral infections. We previously reported that type I and II IFNs control MuPyV infection in non-central nervous system (CNS) organs, but their relative contributions to MuPyV control in the brain remain unknown. To this end, mice deficient in type I, II, or III IFN receptors or STAT1 were infected intracerebrally with MuPyV. We found that STAT1, but not type I, II, or III IFNs, mediated viral control during acute and persistent MuPyV encephalitis. Mice deficient in STAT1 also developed severe hydrocephalus, blood-brain barrier permeability, and increased brain infiltration by myeloid cells. CD8 T cell deficiency alone did not increase MuPyV infection and pathology in the brain. In the absence of STAT1 signaling, however, depletion of CD8 T cells resulted in lytic infection of the choroid plexus and ependymal lining, marked meningitis, and 100% mortality within 2 weeks postinfection. Collectively, these findings indicate that STAT1 signaling and CD8 T cells cocontribute to controlling MuPyV infection in the brain and CNS injury.IMPORTANCE A comprehensive understanding of JCPyV-induced PML pathogenesis is needed to define determinants that predispose patients to PML, a goal whose urgency is heightened by the lack of anti-JCPyV agents. A handicap to achieving this goal is the lack of a tractable animal model to study PML pathogenesis. Using intracerebral inoculation with MuPyV, we found that MuPyV encephalitis in wild-type mice causes an encephalopathy, which is markedly exacerbated in mice deficient in STAT1, a molecule involved in transducing signals from type I, II, and III IFN receptors. CD8 T cell deficiency compounded the severity of MuPyV neuropathology and resulted in dramatically elevated virus levels in the CNS. These findings demonstrate that STAT1 signaling and CD8 T cells concomitantly act to mitigate MuPyV-encephalopathy and control viral infection.

Balancing Inflammation and Central Nervous System Homeostasis: T Cell Receptor Signaling in Antiviral Brain TRM Formation and Function.

Tissue-resident memory (TRM) CD8 T cells provide early frontline defense against regional pathogen reencounter. CD8 TRM are predominantly parked in nonlymphoid tissues and do not circulate. In addition to this anatomic difference, TRM are transcriptionally and phenotypically distinct from central-memory T cells (TCM) and effector-memory T cells (TEM). Moreover, TRM differ phenotypically, functionally, and transcriptionally across barrier tissues (e.g., gastrointestinal tract, respiratory tract, urogenital tract, and skin) and in non-barrier organs (e.g., brain, liver, kidney). In the brain, TRM are governed by a contextual milieu that balances TRM activation and preservation of essential post-mitotic neurons. Factors contributing to the development and maintenance of brain TRM, of which T cell receptor (TCR) signal strength and duration is a central determinant, vary depending on the infectious agent and modulation of TCR signaling by inhibitory markers that quell potentially pathogenic inflammation. This review will explore our current understanding of the context-dependent factors that drive the acquisition of brain (b)TRM phenotype and function, and discuss the contribution of TRM to promoting protective immune responses in situ while maintaining tissue homeostasis.

To Go or Stay: The Development, Benefit, and Detriment of Tissue-Resident Memory CD8 T Cells during Central Nervous System Viral Infections.

CD8 T cells coordinate immune defenses against viral infections of the central nervous system (CNS). Virus-specific CD8 T cells infiltrate the CNS and differentiate into brain-resident memory CD8 T cells (CD8 bTRM). CD8 bTRM are characterized by a lack of recirculation and expression of phenotypes and transcriptomes distinct from other CD8 T cell memory subsets. CD8 bTRM have been shown to provide durable, autonomous protection against viral reinfection and the resurgence of latent viral infections. CD8 T cells have also been implicated in the development of neural damage following viral infection, which demonstrates that the infiltration of CD8 T cells into the brain can also be pathogenic. In this review, we will explore the residency and maintenance requirements for CD8 bTRM and discuss their roles in controlling viral infections of the brain.

PD-1 Dynamically Regulates Inflammation and Development of Brain-Resident Memory CD8 T Cells During Persistent Viral Encephalitis.

Programmed cell death-1 (PD-1) receptor signaling dampens the functionality of T cells faced with repetitive antigenic stimulation from chronic infections or tumors. Using intracerebral (i.c.) inoculation with mouse polyomavirus (MuPyV), we have shown that CD8 T cells establish a PD-1hi, tissue-resident memory population in the brains (bTRM) of mice with a low-level persistent infection. In MuPyV encephalitis, PD-L1 was expressed on infiltrating myeloid cells, microglia and astrocytes, but not on oligodendrocytes. Engagement of PD-1 on anti-MuPyV CD8 T cells limited their effector activity. NanoString gene expression analysis showed that neuroinflammation was higher in PD-L1-/- than wild type mice at day 8 post-infection, the peak of the MuPyV-specific CD8 response. During the persistent phase of infection, however, the absence of PD-1 signaling was found to be associated with a lower inflammatory response than in wild type mice. Genetic disruption and intracerebroventricular blockade of PD-1 signaling resulted in an increase in number of MuPyV-specific CD8 bTRM and the fraction of these cells expressing CD103, the αE integrin commonly used to define tissue-resident T cells. However, PD-L1-/- mice persistently infected with MuPyV showed impaired virus control upon i.c. re-infection with MuPyV. Collectively, these data reveal a temporal duality in PD-1-mediated regulation of MuPyV-associated neuroinflammation. PD-1 signaling limited the severity of neuroinflammation during acute infection but sustained a level of inflammation during persistent infection for maintaining control of virus re-infection.

CD4 T cells control development and maintenance of brain-resident CD8 T cells during polyomavirus infection.

Tissue-resident memory CD8 T (TRM) cells defend against microbial reinfections at mucosal barriers; determinants driving durable TRM cell responses in non-mucosal tissues, which often harbor opportunistic persistent pathogens, are unknown. JC polyomavirus (JCPyV) is a ubiquitous constituent of the human virome. With altered immunological status, JCPyV can cause the oft-fatal brain demyelinating disease progressive multifocal leukoencephalopathy (PML). JCPyV is a human-only pathogen. Using the mouse polyomavirus (MuPyV) encephalitis model, we demonstrate that CD4 T cells regulate development of functional antiviral brain-resident CD8 T cells (bTRM) and renders their maintenance refractory to systemic CD8 T cell depletion. Acquired CD4 T cell deficiency, modeled by delaying systemic CD4 T cell depletion until MuPyV-specific CD8 T cells have infiltrated the brain, impacted the stability of CD8 bTRM, impaired their effector response to reinfection, and rendered their maintenance dependent on circulating CD8 T cells. This dependence of CD8 bTRM differentiation on CD4 T cells was found to extend to encephalitis caused by vesicular stomatitis virus. Together, these findings reveal an intimate association between CD4 T cells and homeostasis of functional bTRM to CNS viral infection.

Combined sublethal irradiation and agonist anti-CD40 enhance donor T cell accumulation and control of autochthonous murine pancreatic tumors.

Tumor-reactive T lymphocytes can promote the regression of established tumors. However, their efficacy is often limited by immunosuppressive mechanisms that block T cell accumulation or function. ACT provides the opportunity to ameliorate immune suppression prior to transfer of tumor-reactive T cells to improve the therapeutic benefit. We evaluated the combination of lymphodepleting whole body irradiation (WBI) and agonist anti-CD40 (αCD40) antibody on control of established autochthonous murine neuroendocrine pancreatic tumors following the transfer of naïve tumor-specific CD8 T cells. Sublethal WBI had little impact on disease outcome but did promote T cell persistence in the lymphoid organs. Host conditioning with αCD40, an approach known to enhance APC function and T cell expansion, transiently increased donor T cell accumulation in the lymphoid organs and pancreas, but failed to control tumor progression. In contrast, combined WBI and αCD40 prolonged T cell proliferation and dramatically enhanced accumulation of donor T cells in both the lymphoid organs and pancreas. This dual conditioning approach also promoted high levels of inflammation in the pancreas and tumor, induced histological regression of established tumors, and extended the lifespan of treated mice. Prolonged survival was entirely dependent upon adoptive transfer, but only partially dependent upon IFNγ production by donor T cells. Our results identify the novel combination of two clinically relevant host conditioning approaches that synergize to overcome immune suppression and drive strong tumor-specific T cell accumulation within well-established tumors.

Inhibition of Retrograde Transport Limits Polyomavirus Infection In Vivo.

Polyomaviruses (PyVs) silently infect most humans, but they can cause life-threatening diseases in immunocompromised individuals. The JC polyomavirus (JCPyV) induces progressive multifocal leukoencephalopathy, a severe demyelinating disease in multiple sclerosis patients receiving immunomodulatory therapy, and BK polyomavirus (BKPyV)-associated nephropathy is a major cause of kidney allograft failure. No effective anti-PyV agents are available. Several compounds have been reported to possess anti-PyV activity in vitro, but none have shown efficacy in clinical trials. Productive PyV infection involves usurping the cellular retrograde vesicular transport pathway to enable endocytosed virions to navigate to the endoplasmic reticulum where virion uncoating begins. Compounds inhibiting this pathway have been shown to reduce infection by simian virus 40 (SV40), JCPyV, and BKPyV in tissue culture. In this study, we investigated the potential of Retro-2.1, a retrograde transport inhibitor, to limit infection by mouse polyomavirus (MuPyV) in vivo. We found that Retro-2.1 significantly reduced MuPyV levels in the kidney during acute infection without affecting renal function or the MuPyV-specific CD8 T cell response. To approximate the clinical setting of PyV resurgence in immunocompromised hosts, we showed that antibody-mediated depletion of T cells in persistently infected mice elevated MuPyV levels in the kidney and that Retro-2.1 blunted this increase in virus levels. In summary, these data indicate that inhibition of retrograde vesicular transport in vivo controls infection in a natural PyV mouse model and supports development of these compounds as potential therapeutic agents for individuals at risk for human PyV-associated diseases. IMPORTANCE PyVs can cause significant morbidity and mortality in immunocompromised individuals. No clinically efficacious anti-PyV therapeutic agents are available. A recently identified inhibitor of retrograde transport, Retro-2cycl, blocks movement of PyV virion-containing vesicles from early endosomes to the endoplasmic reticulum, an early step in the PyV life cycle. Retro-2cycl and its derivatives have been shown to inhibit infection by human PyVs in tissue culture. Here, we demonstrate that a derivative of Retro-2cycl, Retro-2.1, reduces infection by MuPyV in the kidneys of acutely infected mice. Mimicking the common clinical scenario of PyV resurgence, we further show that MuPyV levels increase in the kidneys of immunocompromised, persistently infected mice and that this increase is inhibited by Retro-2.1. These data provide the first evidence for control of a natural PyV infection in vivo by administration of an inhibitor of retrograde transport.

Maintenance of PD-1 on brain-resident memory CD8 T cells is antigen independent.

Infection of the central nervous system (CNS) by murine polyomavirus (MuPyV), a persistent natural mouse pathogen, establishes brain-resident memory CD8 T cells (bTRM) that uniformly and chronically express programmed cell death protein 1 (PD-1) irrespective of the expression of αE integrin CD103, a TRM cell marker. In contrast, memory antiviral CD8 T cells in the spleen are PD-1-, despite viral loads being similar in both the brain and spleen during persistent infection. Repetitive antigen engagement is central to sustained PD-1 expression by T cells in chronic viral infections; however, recent evidence indicates that expression of inhibitory receptors, including PD-1, is part of the TRM differentiation program. Here we asked whether PD-1 expression by CD8 bTRM cells during persistent MuPyV encephalitis is antigen dependent. By transferring MuPyV-specific CD8 bTRM cells into the brains of naive mice and mice infected with cognate epitope-sufficient and -deficient MuPyVs, we demonstrate that antigen and inflammation are dispensable for PD-1 maintenance. In vitro and direct ex vivo analyses indicate that CD103- MuPyV-specific CD8 bTRM retain functional competence. We further show that the Pdcd-1 promoter of anti-MuPyV bTRM cells is epigenetically fixed in a demethylated state in the brain. In contrast, the PD-1 promoter of splenic antiviral memory CD8 T cells undergoes remethylation after being demethylated during acute infection. These data show that PD-1 expression is an intrinsic property of brain TRM cells in a persistent CNS viral infection.

Alloimmunity But Not Viral Immunity Promotes Allograft Loss in a Mouse Model of Polyomavirus-Associated Allograft Injury.

BACKGROUND: The interplay between viral infection and alloimmunity is known to influence the fate of transplanted organs. Clarifying how local virus-associated inflammation/injury and antiviral immunity can alter host alloimmune responses in transplantation remains a critical question. METHODS: We used a mouse model of polyomavirus (PyV) infection and kidney transplantation to investigate the roles of direct viral pathology, the antiviral immune response, and alloimmunity in the pathogenesis of PyV-associated allograft injury. We have previously shown that an effective primary T cell response is required in PyV-associated graft injury. RESULTS: Here we show that the transfer of primed antidonor, but not antiviral, T cells results in PyV-associated allograft injury. In further studies, we use a surrogate minor antigen model (ovalbumin) and show that only antidonor specific T cells and not antiviral specific T cells are sufficient to mediate injury. Lastly, we demonstrate that local but not systemic virus-mediated inflammation and injury within the graft itself are required. CONCLUSIONS: These data suggest that in this mouse model, the predominant mechanism of allograft injury in PyV-associated injury is due to an augmented alloimmune T cell response driven by virus-induced inflammation/injury within the graft. These studies highlight the important interplay between viral infection and alloimmunity in a model system.

TCR stimulation strength is inversely associated with establishment of functional brain-resident memory CD8 T cells during persistent viral infection.

Establishing functional tissue-resident memory (TRM) cells at sites of infection is a newfound objective of T cell vaccine design. To directly assess the impact of antigen stimulation strength on memory CD8 T cell formation and function during a persistent viral infection, we created a library of mouse polyomavirus (MuPyV) variants with substitutions in a subdominant CD8 T cell epitope that exhibit a broad range of efficiency in stimulating TCR transgenic CD8 T cells. By altering a subdominant epitope in a nonstructural viral protein and monitoring memory differentiation of donor monoclonal CD8 T cells in immunocompetent mice, we circumvented potentially confounding changes in viral infection levels, virus-associated inflammation, size of the immunodominant virus-specific CD8 T cell response, and shifts in TCR affinity that may accompany temporal recruitment of endogenous polyclonal cells. Using this strategy, we found that antigen stimulation strength was inversely associated with the function of memory CD8 T cells during a persistent viral infection. We further show that CD8 TRM cells recruited to the brain following systemic infection with viruses expressing epitopes with suboptimal stimulation strength respond more efficiently to challenge CNS infection with virus expressing cognate antigen. These data demonstrate that the strength of antigenic stimulation during recruitment of CD8 T cells influences the functional integrity of TRM cells in a persistent viral infection.