<p>The progression of an inflammatory response is largely dictated by soluble<br />factors termed cytokines reknown for their redundant and pleiotropic nature in<br />modulation of both immune and stromal cells. Individual members of the<br />interleukin-6 (IL-6)-type cytokine family possess both unique and shared biological<br />activities. These cytokines may participate in tissue remodelling by promoting<br />reconstruction ofextracellular matrix (ECM) following nonspecific tissue damage by<br />inflammatory cells. Consistent with this view, these cytokines upregulate expression<br />of an ECM protease inhibitor, tissue inhibitor of metalloproteinases-1 (TIMP-1) and<br />thus may alter net enzymatic degradation of ECM. The overall goal of this thesis<br />is to examine mechanisms by which TIMP-1 is regulated by IL-6-type cytokines,<br />especially by the cytokine oncostatin M (OSM). The promoter of TIMP-1 has<br />therefore been studied in detail to address the mechanisms by which OSM (and IL-6<br />type cytokines) regulate the transcription of the TIMP-1 gene. The approaches<br />undertaken have included deletion analysis of the TIMP-1 gene nucleotide<br />sequences proximal to the start of transcription to define DNA sequences<br />necessary/sufficient for cytokine-induced TIMP-1 promoter activity. In addition, the<br />binding of nuclear factors to these DNA elements and cytokine-response elements, their expression and involvement in the regulation of TIMP-1 transcription have been explored.</p> <p>We have identified sequences proximal to the start of TIMP-1 transcription<br />that are necessary for maximal responsiveness to OSM and IL-6. Deletion analysis<br />of the proximal TIMP-1 promoter (-95 to +47 TIMP-1 sequences) has identified a<br />nucleotide sequence within -59 to -53 ofthe murine TIMP-1 promoter that harbours<br />an AP-1 consensus DNA binding element. This element is necessary for maximal<br />OSM or IL-6 induced promoter activity of TIMP1-CAT reporter gene constructs<br />transfected into human hepatoma HepG2 cells. OSM is the most potent stimulus<br />(approx. 11-fold for OSM, and 4-fold for IL-6) of this response and additional<br />sequences 3-prime to +1 ofthe TIMP-1 gene are also necessary for maximal OSM<br />responsiveness.</p> <p>Electrophoretic mobility shift assays demonstrated two gel-shifted complexes<br />which bind the TIMP-1 AP-1 site. An AP-1 gel-shifted complex is present in the<br />absence of cytokine stimulation ("complex 1"), while OSM and not other IL-6-type<br />cytokines, stimulated the formation of a second AP-1 gel shifted complex.<br />Nuclear factors binding to TIMP-1 AP-1 complex 1 include junB, junD and fosrelated<br />antigens. However, unlike complex1, c-fos is present and necessary for the<br />formation of the OSM-induced TIMP-1 AP-1 complex2. Consistent with this, OSM<br />is a potent inducerofc-fos protein expression among IL-6-type cytokines. Both the<br />formation of complex2 and c-fos expression require new protein synthesis. JunB<br />and junO are constitutively expressed, while the expression of fos-related antigens are induced in response to OSM. In addition, although PMA was also a potent<br />inducer of c-fos expression, induction of TIMP-1 promoter activity by the<br />combination of PMA and IL-6 was comparable to IL-6 alone and did not equal the<br />significantly higher induction by OSM. Within the same cells, OSM and IL-6 equally<br />induced STATDNA-binding activity. An Ets-consensus site (nucleotides -45 to -40)<br />flanking the 3-prime end of the AP-1 site is a weak binding site for Ets-related<br />nuclear factors, and an SP-1 site near +1 (-11 to -6) is a strong binding siteforSP1<br />nuclear factors and related SP-1 site binding proteins. No STAT nuclear factor<br />binding to the proximal TIMP-1 promoterwas detected. Taken together, the TIMP1<br />AP-1 site and c-fos represent a unique target of OSM signalling and activation of<br />AP-1 complexes (possibly containing c-fos) by OSM as well as sequences<br />downstream of TIMP-1 +1 contribute to maximal responsiveness of the promoter<br />to this cytokine among IL-6 family members.</p> <p>The contribution of c-fos to OSM-induced TIMP-1 expression was further<br />explored in murine cells. OSM stimulates the expression of c-fos and activates<br />STATs 1, 3 and 5 DNA-binding activity in murine fibroblasts. As observed in<br />human cells, OSM but not other IL-6-type cytokines upregulated c-fos expression<br />which participated in complexes binding the TIMP-1 AP-1 site. OSM was also<br />unique among IL-6 family members in activating STAT5 DNA-binding activity in<br />murine fibroblasts. However, in contrast to observations in human HepG2 cells,<br />deletion analysis of the TIMP-1 promoter showed that the AP-1 site (-59/-53) was<br />v not necessary for OSM-mediated upregulation of the TIMP-1 proximal promoter<br />activity over basal levels in murine NIH3T3 fibroblasts and co-transfection of a<br />dominant-negative of AP-1 had no effect. However, transfection of dominant-negative<br />STATs1, 3 or5 (especially STAT3) could diminish cytokine-induced TIMP1<br />promoter activity. In addition, c-Fos was dispensable for OSM-mediated<br />upregulation ofTIMP-1 mRNA levels as TIMP-1 expression was detected in wildtype<br />and c-fos knockout murine lung fibroblasts. Consistent with deletion analysis<br />ofthe TIMP-1 promoter in human cells, dominant-negative AP-1 expression vectors<br />abrogated OSM-mediated TIMP-1 promoter activity, while Stat-dominant negative<br />expression vectors did not. Taken together, the examination of murine and human<br />systems suggests that AP-1 and STAT nuclear factors can contribute to the<br />regulation of the TIMP-1 promoter.</p> / Doctor of Philosophy (PhD)

Dengue fever is a rapidly growing concern to human health and is currently the most prevalent mosquito-borne viral disease worldwide. Although there are several vaccine candidates being tested in clinical trials, there are no vaccines publicly available to prevent this disease. Plant-based vaccines are rapidly becoming viable alternatives to traditional animal-based vaccines because they are safe, easy to manufacture, and more cost-efficient. The purpose of this project is to develop a vaccine against the dengue virus by producing a recombinant DENV2 protein, engineered by Dr. David Lo and his lab at University of California Riverside, in Nicotiana benthamiana plants through Tobacco Mosaic Virus (TMV) infection. Initial attempts to ligate the complete DENV2 epitope, a combination of hybrid flagellin sequences and the envelope protein from dengue viral serotype 2, into the pJL TRBO vector were incompatible with established protocols. However, a proof of concept test that replaced the DENV2 envelope protein with Green Fluorescent Protein (GFP) successfully inserted the new sequence into pJL TRBO. In the future, the DENV2 envelope protein sequence will be re-inserted into the construct and updated protocols will be repeated for DENV2 protein expression. The recombinant DENV2 proteins will be extracted from the plants after signs of infection become apparent and tested for their ability to induce an immunogenic response that produces pathogen-specific antibodies.

Periodontitis is a biofilm-initiated inflammatory disease, resulting in soft tissue damage and alveolar bone loss. A nucleic acid sensor, toll-like receptor 9 (TLR9), has been recently implicated in periodontal inflammation. This study utilized an in vivo periodontitis model using TLR9 knockout (TLR9-/-) mice to assess the role of TLR9 in periodontitis. Significant bone loss was observed in wild type, but not in TLR9-/- mice. Further experiments using ex vivo assays revealed significantly higher IL-6 production in splenocytes of wild type mice compared to knockout cells in response to Porphyromonas gingivalis (a keystone pathogen for periodontitis) challenge. In conclusion, TLR9 contributes to periodontal inflammation through promoting a heightened inflammatory response. Therapeutics targeted to TLR9 may be beneficial to control periodontal disease.

Leishmaniasis refers to the group of diseases caused by pathogenic protozoan parasites of the genus Leishmania. Nearly all human Leishmania spp. infections are initiated in mammalian skin through the bite of the phlebotomine sand fly vector. However, clinical manifestations vary greatly with infecting species. Leishmania major establish infection locally within the skin and cause chronic ulcerating skin lesions at the local cutaneous site of inoculation, in a syndrome known as Cutaneous Leishmaniasis (CL) Leishmania infantum parasites metastasize from the site of skin infection via unknown mechanisms, and establish infection within visceral organs usually without inducing skin pathology, resulting in the potentially fatal disseminated disease, Visceral Leishmaniasis (VL). Mouse studies suggest early responses at the skin infection site are critical determinants of subsequent adaptive immune responses in leishmaniasis, yet few studies address the role of keratinocytes, the most abundant immunoactive cell in the epidermis. We hypothesize that Leishmania infection causes keratinocytes to produce immunomodulatory factors that influence the outcome of infection. Incubation of primary or immortalized human keratinocytes with L. infantum or L. major elicited dramatically different responses. Keratinocytes incubated with L. infantum significantly increased expression of pro-inflammatory genes IL6, IL8, TNF, and IL1B by RT-qPCR; whereas keratinocytes exposed to L. major did not. Similar to live parasites, L. infantum-derived exosomes induced more IL8 mRNA compared to control or L. major-derived exosomes. Western blotting confirmed NFkBp65 phosphorylation in keratinocytes exposed to L. infantum but not L. major. However, no evidence of L. major inhibition of TNF-induced NFkBp65 phosphorylation was observed in simultaneously treated keratinocytes. To examine whether keratinocytes influence proximal host cells, L. infantum-infected human monocytes were co-cultured with keratinocytes across a transwell membrane. These studies suggested L. infantum-exposed keratinocytes release soluble factors that enhance monocyte control of intracellular L. infantum replication. L. major-exposed keratinocytes had no comparable effect. These data suggest L. infantum and L. major differentially activate keratinocytes to release factors that limit infection in monocytes. Microarray analyses performed on human keratinocytes exposed to either L. infantum or L. major promastigotes identified a limited number of transcripts increased by parasite exposure. Consistent with RT-qPCR observations, several inflammatory cytokine and chemokine genes were more strongly induced in L. infantum-exposed keratinocytes compared to L. major-exposed keratinocytes. Pathway analyses of genes induced by L. infantum-treated keratinocytes suggested that this interaction may induce neutrophil recruitment. Notably, AP1 transcription factor subunit genes were significantly down regulated in L. major-treated compared with L. infantum-treated or control keratinocytes. This suggests L. major may actively inhibits this keratinocyte activation, which might affect its ability to establish infection within host skin. In addition, ex vivo intradermal infection of human skin explants was explored as a method to compare keratinocyte responses to L. infantum or L. major in the context of whole skin tissue and the effects of vector salivary gland components are considered. The response of keratinocytes found in these studies using L. infantum and L. major may give insight into the local host pathologic responses to different Leishmania species leading to visceralizing versus cutaneous manifestations to infection. We propose that Leishmania spp. elicit or evade a pro-inflammatory response by keratinocytes at the site of infection, generating a microenvironment uniquely tailored to each Leishmania species.

Tim-3 (for T cell immunoglobulin and mucin domain 3) is a surface molecule expressed throughout the immune system that appears to mediate both stimulatory and inhibitory effects. Tim-3 is expressed by activated CD4 and CD8 T cells, which suggests a direct role in the regulation of T cell responses. Consistent with this possibility, previous studies have shown that blockade of interactions between Tim-3 and ligands in the context of mouse models for autoimmunity and chronic infection can augment T cell responses, which indicates that Tim-3 functions as an inhibitory receptor for T cells. However, other studies have provided evidence that Tim-3 can function to promote T cell responses, which suggests that Tim-3 acts as a stimulatory receptor. In addition, biochemical and cell culture studies have shown that Tim-3 can induce both inhibitory and stimulatory signaling pathways. These conflicting findings highlight that the role of Tim-3 in regulating T cell responses remains unclear and warrants further investigation. My studies sought to determine the role of Tim-3 in regulating T cell responses to microbial infections in vivo and to acute stimulation in vitro. Using Tim-3 KO mice and Tim-3 KO cells, I demonstrate that Tim-3 can directly enhance CD8 T cell responses to Listeria monocytogenes (LM). I also provide evidence that Tim-3 indirectly regulates CD4 T cell responses to LM. Alternatively, I show that Tim-3 may inhibit CD4 and CD8 T cell responses to the chronic viral infection LCMV-Clone 13, and that Tim-3 may be dispensable for T cell responses to the acute viral infection LCMV-Armstrong. Additionally, I demonstrate that Tim-3 expression in response to acute stimulation in vitro or in vivo marks populations of CD4 Th1 cells that are enriched for cells with effector function as measured by cytokine production and markers for degranulation. Collectively, these data suggest that Tim-3 may differentially regulate T cell effector function in a manner that is dependent on infectious environment encountered.

The immunosuppressive effects of chronic alcohol abuse are profound, wide-ranging and readily apparent at the body's barriers. In the skin, alcoholism is associated with an increased incidence and severity of infection; yet the precise immunologic alterations responsible remain poorly understood. Cutaneous homeostasis and immunity are afforded via coordinated efforts of tissue-specific immune cell networks. Here, the Meadows-Cook murine model of alcoholism was used to investigate the impact of chronic ethanol (EtOH) exposure upon the following: 1) the composition and function of skin-resident dendritic cells (DCs) and T cells in preimmune mice 2) infection outcome and host defense following Staphylococcus aureus skin infection 3) the induction of cutaneous oxidative stress Chronic EtOH feeding caused a baseline reduction in skin DCs and T cells with the most pronounced effects occurring in self-renewing compartments (i.e, Langerhans cells and γδ T cells). In addition, we found that EtOH-induced immune cell subset loss was often associated with dysfunction of the remaining population. For DCs, EtOH-induced hyporesponsiveness was observed in both in vitro and in vivo migration assays. Defects in the former system could be corrected via TNFα restoration. EtOH-induced dysfunction in skin T cells was evident by the decreased upregulation of JAML and diminished production of IL-17 by epidermal and dermal γδ T cells respectively. In a murine model of EtOH withdrawal, some but not all of the EtOH-induced defects occurring in skin DCs and T cells recovered after cessation of EtOH exposure. Prior to this work, the impact of chronic EtOH exposure upon the cutaneous immune system had not been investigated in a murine model of infection. Using a novel method of cutaneous S. aureus challenge, evidence of exacerbated staphylococcal disease in EtOH-fed mice included skin lesions that were larger and contained more organisms, greater weight loss and increased bacterial dissemination. Infected EtOH-fed mice demonstrated poor maintenance and induction of PMN responses in the skin and draining LNs respectively. Additionally, altered PMN dynamics in the skin of these mice corresponded with reduced production of IL-23 and IL-1β by CD11b+ myeloid cells and IL-17 production by γδ T cells, with the latter defect occurring in the draining LNs as well. In addition, IL-17 restoration via intradermal injection improved bacterial clearance defects in EtOH-fed mice. Taken together, these findings show that the EtOH-induced increase in S. aureus-related injury/illness (i.e., weight loss, bacterial burden, lesion size) corresponds with defects in the IL-23/IL-17 inflammatory axis and poor PMN accumulation at the site of infection and draining LNs. Finally, two complementary tools (mice deficient in molecules that promote or inhibit reactive oxygen species induction) were used to investigate the role of oxidative stress as a driver of cutaneous immune dysfunction. In these studies intriguing evidence was obtained indicating that some but not all of the mechanisms by which oxidative stress contributes to cutaneous immune dysfunction are initiated through the Thurman cascade. In conclusion, this report offers new information about the impact of EtOH on cutaneous host defense pathways and provides potential mechanisms of explaining why alcoholics are predisposed to severe skin infections.

The immune system has evolved to fight off numerous pathogens. The first line of defense against these pathogens are innate immune cells. Innate immune cells ingest pathogens and a family of cytosolic proteins, NOD-like receptors (NLR) and AIM2-like receptors (ALRs), recognize conserved sequences on pathogens. Recognition of pathogens by NLRs and ALRs alerts the immune system to the presence of an invader and subsequent control of infection. NLRs and ALRs also recognize endogenous cell danger signals that are released during cell stress. Pathogens and tumor cells are capable of growing in a host with limited detection by the immune system. This evasion leads to a suboptimal immune response to both the bacteria and cancer cells resulting in enhanced infection and tumor growth. We investigated how the bacteria Francisella tularensis escapes recognition by the AIM2 receptor. We identified a novel gene important in bacterial folate metabolism that helps the bacteria escape immune recognition. The identification of this gene will help in the development of treatments for F. tularensis infection. In addition, we investigated the role of the NLRC4 receptor in a mouse model of melanoma. We found that mice lacking NLRC4 developed significantly larger tumors and had a diminished immune response compared to mice that expressed NLRC4. We also observed decreased expression of NLRC4 in metastatic melanoma tissue in humans. Thus demonstrating that NLRC4 is important for initiating an immune response to melanoma and down regulating expression of NLRC4 is a way for the tumor to evade the immune response.

Viral infection of the central nervous system is complicated by the mostly irreplaceable nature of neurons, as the loss of neurons has the potential to result in permanent damage to brain function. However, whether neurons or other cells in the CNS sometimes survive infection and the effects of infection on neuronal function are largely unknown. To address this question, I used the rJHM strain (rJ) of mouse hepatitis virus, (MHV), a neurotropic coronavirus, which causes acute encephalitis in susceptible strains of mice. To determine whether neurons or other CNS cells survive acute infection with this virulent virus, I developed a recombinant JHMV that expresses Cre recombinase (rJ-Cre) and infected mice that universally expressed a silent (floxed) version of tdTomato. Infection of these mice with rJ-Cre resulted in expression of tdTomato in host cells. The results showed that some cells were able to survive the infection, as demonstrated by continued tdTomato expression after virus antigen could no longer be detected. Most notably, interneurons in the olfactory bulb, which are known to inhibit other neurons, represented a large fraction of the surviving cells. The results described in this thesis indicated that some neurons are resistant to virus-mediated cell death and provide a framework for studying the effects of prior coronavirus infection on neuron function. Another aspect of this thesis concerns the role of microglia in virus infections. Recent findings have highlighted roles for microglia in orchestrating normal development and refining neural network connectivity in the healthy central nervous system (CNS). Microglia are not only vital cells in maintaining CNS homeostasis, but also respond to injury, infection, and disease, by undergoing proliferation, and changes in transcription and morphology. Understanding the specific role of microglia in responding to viral infection is complicated by the presence of non-microglial myeloid cells with potentially overlapping function in the healthy brain, and by the rapid infiltration of hematopoietic myeloid cells into the brain in diseased states. Here, I used a CSF1R inhibitor that depletes microglia to examine the specific roles that microglia play in response to infection with a neurotropic coronavirus (MHV). My results show that microglia were required during the first days post infection to limit MHV replication, subsequent tissue damage and lethality. Additionally, microglia depletion resulted in ineffective T cell responses. These results reveal nonredundant, critical roles for microglia in the early innate and virus-specific T cell responses and for subsequent host protection from viral encephalitis. Overall these studies provide new insight into cells that survive MHV infection and into the role microglia play in the immune response to MHV. Specifically, many neurons, especially interneurons of the olfactory bulb survive MHV infection and microglia play a role in both the innate and adaptive immune response to viruses.

Sepsis occurs when infection enters the circulation resulting in harmful or lethal levels of pro- and anti-inflammatory cytokines that affects nearly 2 million people in the U.S. annually. Improved identification and treatment options have increased survival of patients shortly after sepsis. However, this management of sepsis subsequently revealed survivors have increased long-term mortality rates due to altered immune responses that increase susceptibility to secondary complications. My thesis further revealed how sepsis impacts CD8 T cell-mediated immunity that protects the host against sepsis unrelated complications that will be useful for improving the outcome of sepsis survivors. Using vaccinia virus (VacV) to generate circulating (TCIRCM) and skin resident (TRM) memory CD8 T cells, I showed sepsis preferentially diminishes the number and function of TCIRCM, while TRM in barrier tissues were unaffected in mice that received a low-severity model of sepsis. Despite optimal number and ‘sensing and alarming’ function of skin TRM, the capacity of these cells to promote a tissue-wide recall response was lost after sepsis that increased viral burden upon homologous skin infection. Decreased sensitivity of vascular endothelium to TRM-derived IFN-γ was the underlying mechanism that diminished localized recall responses after sepsis. Overall, my data stress the importance of understanding how sepsis-induced lesions in T cell-extrinsic factors contribute to diminished T cell-mediated immunity observed after sepsis. Sepsis influence on T cell-mediated immunity has previously been explored using model pathogens. However, sepsis survivors are susceptible to an array of secondary complications, including cancer, which often occurs in patients with no previous history of malignancy. This suggested the immunoparalysis phase of sepsis increases cancer incidence and/or progression that may contribute to the increased long-term mortality of this population. I showed that previously-septic mice had increased cancer-associated mortality weeks/months after sepsis recovery due to diminished frequency and function of tumor-infiltrating CD8 T cells (CD8 TILs) that mediate partial control of B16 melanoma. CD8 T cells with high expression of inhibitory receptors (denoted PD-1hi) that retain some effector functions in the tumor were preferentially eliminated in sepsis survivors. The rapid loss of PD-1hi CD8 TILs diminished the window of efficacy for checkpoint blockade therapy designed to improve the number and function of this T cell subset. Thus, the chronic immunoparalysis phase of sepsis is defined by diminished tumor-specific CD8 T cell responses that increases susceptibility to cancer mortality. Most sepsis patients have comorbidities at the time of sepsis onset but the capacity of sepsis to influence comorbidity burden remains unknown. Because cancer is the highest risk factor for sepsis, I examined the capacity of sepsis to influence the outcome of hosts bearing solid tumors. Sepsis was performed after B16 inoculation and recapitulated clinical data showing increased sepsis mortality in cancer patients. However, sepsis mortality only occurred in mice bearing large tumors, while hosts with less progressed tumors survived the septic event and had improved cancer prognosis. CD8 T cells were required for this phenomenon and the number of vasculature-excluded CD8 TILs was not altered after sepsis. Despite this, I showed CD8 TILs from mice that survived sepsis had increased activation and effector functions suggesting sepsis has the unexpected capacity to reinvigorate this subset. In total, sepsis impact on CD8 T cell responses alters host outcome to additional disease states that commonly affect sepsis patients, a notion that will be important for a growing number of sepsis survivors.

CD8 T cells have been demonstrated to be critical in the resolution of acute influenza A virus (IAV) infections. Previously our laboratory has demonstrated that the magnitude of the IAV-specific CD8 T cell response is inversely proportional to the initial IAV inoculum. The decrease in CD8 T cells observed during lethal dose IAV infections was shown tobe a result of apoptosis driven by FasL expressed on lymph node dendritic cells (LNDC) during lethal, but not sublethal, dose IAV infections. However the specific LNDC subset(s) responsible for FasL:Fas mediated apoptosis of IAV-specific CD8 T cells remains to be identified. The existence of multiple subsets of dendritic cells (DC) within the lymph node (LN) with distinct functions during viral infections suggest the possibility that a specific subset(s) may be responsible for this effect. Furthermore, the regulation of FasL expression on LNDC during lethal versus sublethal dose IAV infections was shown to be dependent on the levels of IL-12p40. However, the specific IL-12p40 containing cytokine, as well as which cells produce this cytokine within the LN remain unknown. Finally, whether or not the expression of FasL expression induced during infections is unique to IAV or if other pulmonary insults can mediate this effect is as of yet undetermined. Here we demonstrate that plasmacytoid DC (pDC), which accumulate in the lung draining LN, are the only LNDC subset eliminating IAV-specific CD8 T cells through FasL:Fas dependent mechanism both in vitro and in vivo during lethal dose IAV infections despite FasL expression by all LNDC subsets. Further we demonstrate that this disparity in LNDC FasL induced apoptosis is related to the individual DC subsets ability to present IAV antigen as pDC are the only LNDC subset incapable of viral antigen presentation via MHC class I during IAV infections. This dissertation further demonstrates that IL-12p40 homodimer (p402), which is produced by respiratory DC (rDC) and LNDC, and not IL-12p40 monomer controls FasL expression on LNDC during lethal dose IAV infections. Additionally I also demonstrate that IL-12Rβ1, which binds IL-12p40, is important for p402 mediated LNDC FasL expression. We further go on to demonstrate that rDC migration from the lungs to the LN is required for both LNDC p402 production and FasL expression. However, LNDC in isolation are unable maintain FasL expression suggesting their production of p402 may not me sufficient for FasL expression. Conversely, rDC are sufficient to induce FasL expression on LNDC from IL-12p40 deficient LN. Together these data suggest that rDC are a critical component of p402 mediated LNDC FasL expression. Finally, we demonstrate that the differential production of p402, and downstream LNDC FasL expression, observed during lethal versus sublethal dose IAV infections is not unique to IAV as intranasal stimulation with diverse TLR agonists also results in differential rDC p402 production and LNDC FasL expression. These data suggest that in addition to IAV, a multitude of pulmonary pathogens may regulate antigen-specific CD8 T cells through this pathway. Taken together the results present herein detail a mechanism of CD8 T cell regulation during IAV infections mediated through p402 induction of FasL expression on pDC within the LN. FasL expressing pDC then induce apoptosis of activated Fas+ IAV-specific CD8 T cells within the LN during lethal a IAV infections leading to a reduction in the number of IAV-specific CD8 T cells that reach the lung and as a result death of the host.