Altered IFN- −γ Mediated Immunity and Transcriptional Expression Patterns in N -Ethyl-N-Nitrosourea−Induced STAT4 Mutants Confer Susceptibility to Acute This information is current as Typhoid-like Disease of September 25, 2021. Megan M. Eva, Kyoko E. Yuki, Shauna M. Dauphinee, Jeremy A. Schwartzentruber, Michal Pyzik, Marilène Paquet, Mark Lathrop, Jacek Majewski, Silvia M. Vidal and Danielle Malo Downloaded from J Immunol 2014; 192:259-270; Prepublished online 27 November 2013; doi: 10.4049/jimmunol.1301370 http://www.jimmunol.org/content/192/1/259 http://www.jimmunol.org/ Supplementary http://www.jimmunol.org/content/suppl/2013/11/27/jimmunol.130137 Material 0.DC1 References This article cites 76 articles, 24 of which you can access for free at: http://www.jimmunol.org/content/192/1/259.full#ref-list-1 by guest on September 25, 2021 Why The JI? Submit online. • Rapid Reviews! 30 days* from submission to initial decision • No Triage! Every submission reviewed by practicing scientists • Fast Publication! 4 weeks from acceptance to publication *average Subscription Information about subscribing to The Journal of Immunology is online at: http://jimmunol.org/subscription Permissions Submit copyright permission requests at: http://www.aai.org/About/Publications/JI/copyright.html Email Alerts Receive free email-alerts when new articles cite this article. Sign up at: http://jimmunol.org/alerts The Journal of Immunology is published twice each month by The American Association of Immunologists, Inc., 1451 Rockville Pike, Suite 650, Rockville, MD 20852 Copyright © 2013 by The American Association of Immunologists, Inc. All rights reserved. Print ISSN: 0022-1767 Online ISSN: 1550-6606. The Journal of Immunology Altered IFN-g–Mediated Immunity and Transcriptional Expression Patterns in N-Ethyl-N-Nitrosourea–Induced STAT4 Mutants Confer Susceptibility to Acute Typhoid-like Disease Megan M. Eva,*,† Kyoko E. Yuki,*,† Shauna M. Dauphinee,*,† Jeremy A. Schwartzentruber,*,‡ Michal Pyzik,*,† Marile`ne Paquet,x Mark Lathrop,*,‡ Jacek Majewski,*,‡ Silvia M. Vidal,*,†,{ and Danielle Malo*,†,{ Salmonella enterica is a ubiquitous Gram-negative intracellular bacterium that continues to pose a global challenge to human health. The etiology of Salmonella pathogenesis is complex and controlled by pathogen, environmental, and host genetic factors. In Downloaded from fact, patients immunodeficient in genes in the IL-12, IL-23/IFN-g pathway are predisposed to invasive nontyphoidal Salmonella infection. Using a forward genomics approach by N-ethyl-N-nitrosourea (ENU) germline mutagenesis in mice, we identified the Ity14 (Immunity to Typhimurium locus 14) pedigree exhibiting increased susceptibility following in vivo Salmonella challenge. A DNA-binding domain mutation (p.G418_E445) in Stat4 (Signal Transducer and Activator of Transcription Factor 4) was the causative mutation. STAT4 signals downstream of IL-12 to mediate transcriptional regulation of inflammatory immune responses. In mutant Ity14 mice, the increased splenic and hepatic bacterial load resulted from an intrinsic defect in innate cell function, IFN-g–mediated http://www.jimmunol.org/ immunity, and disorganized granuloma formation. We further show that NK and NKT cells play an important role in mediating control of Salmonella in Stat4Ity14/Ity14 mice. Stat4Ity14/Ity14 mice had increased expression of genes involved in cell–cell interactions and communication, as well as increased CD11b expression on a subset of splenic myeloid dendritic cells, resulting in compromised recruitment of inflammatory cells to the spleen during Salmonella infection. Stat4Ity14/Ity14 presented upregulated compensatory mechanisms, although inefficient and ultimately Stat4Ity14/Ity14 mice develop fatal bacteremia. The following study further elucidates the pathophysiological impact of STAT4 during Salmonella infection. The Journal of Immunology, 2014, 192: 259–270. cute foodborne bacterial infections remain a major public waterborne illnesses, from a self-limiting, localized gastroenteritis by guest on September 25, 2021 health problem associated with high morbidity and mor- to the more severe, potentially fatal systemic disease of typhoid A tality worldwide. The intracellular bacterium Salmonella fever (2). Certain infected individuals (1–4%) may further develop enterica continues to pose a global challenge to human health (1). recurrent infections or become asymptomatic chronic carriers acting In humans, Salmonella infections cause a range of foodborne and as a reservoir for pathogen persistence and dissemination in the population (3, 4). Typhoid fever is strictly caused by an infection *Department of Human Genetics, McGill University, Montreal, Quebec H3G 0B1, with the human-restricted S. enterica serovars Typhi and Paratyphi. Canada; †Complex Traits Group, McGill University, Montreal, Quebec H3G 0B1, It is primarily endemic in developing areas of the world where poor Canada; ‡McGill University and Genome Quebec Innovation Centre, Montreal, Que- x sanitation and lack of access to clean drinking water are common bec H3B 1S6, Canada; De´partement de Pathologie et de Microbiologie, Faculte´ de Me´decine Ve´te´rinaire, Universite´ de Montre´al, Saint-Hyacinthe, Quebec J2S 5B5, (5). In contrast, nontyphoidal Salmonella (NTS) serovars, such as { Canada; and Department of Medicine, McGill University, Montreal, Quebec H3G Salmonella Typhimurium and Salmonella Enteritidis, are capable of 0B1, Canada infecting a broad spectrum of hosts, resulting in intestinal and di- Received for publication May 23, 2013. Accepted for publication October 25, 2013. arrheal disease (salmonellosis). NTS serovars are the second leading This work was supported by funds from the Team Program of the Canadian Institutes cause of foodborne illnesses in North America. Approximately 5% of Health Research (to S.M.V. and D.M.). M.M.E. was recipient of a studentship from the Research Institute of the McGill University Health Centre. K.E.Y. received a Fac- of patients with salmonellosis are at increased risk of further de- ulty of Medicine Internal Studentship award. S.M.D. received a Faculty of Medicine veloping invasive transient bacteremia and sepsis (6). Indeed, dis- David Lin fellowship, a Research Institute of the McGill University Health Centre Salmonella fellowship, and a Fonds de la Recherche du Que´bec fellowship. S.M.V. holds a Can- ease manifestation of -related infections in humans is ada Research chair and D.M. is a McGill Dawson scholar. dependent on the complex interaction between environmental fac- The sequences presented in this article have been submitted to the ArrayExpress tors, bacterial serotype, and host genetic factors. database (http://www.ebi.ac.uk/arrayexpress) under accession number E-MTAB-1931. The outcome of S. enterica infection relies on the activation of Address correspondence and reprint requests to Dr. Danielle Malo, Department of both early innate functions and adaptive humoral and cell-mediated Human Genetics and Department of Medicine, McGill University Life Sciences immune responses of the host (7, 8). During systemic Salmonella Complex, Bellini Building, 3649 Promenade Sir-William-Osler, Montreal, QC H3G 0B1, Canada. E-mail address: [email protected] infection, rapid neutrophilic infiltration and phagocytosis by tissue The online version of this article contains supplemental material. macrophages are crucial in limiting hepatosplenic infection. Acti- Abbreviations used in this article: DC, dendritic cell; ENU, N-ethyl-N-nitrosourea; vated macrophages limit Salmonella replication and dissemination IPA, Ingenuity pathway analysis; KO, knockout; MFI, mean fluorescence intensity; to new foci by phagolysosomal bacterial killing and secretion of in- NTS, nontyphoidal Salmonella; PMN, polymorphonuclear cell; SNP, single nucleo- flammatory chemokines and cytokines, including TNF-a, IL-12, tide polymorphism. IL-18, and IFN-g (7). The inflammatory environment results in Copyright Ó 2013 by The American Association of Immunologists, Inc. 0022-1767/13/$16.00 the rapid recruitment of leukocytes that increase the ability for www.jimmunol.org/cgi/doi/10.4049/jimmunol.1301370 260 ROLE OF STAT4 IN SALMONELLA INFECTION intracellular pathogen killing and formation of granulomas. Im- following sublethal invasive S. Typhimurium challenge. Overall, portantly, IL-12 induces IFN-g secretion from NK cells, which is Stat4Ity14/Ity14 mice have increased mortality following infection, critical in activating macrophages in the early response against with a concomitant progressive increase in splenic and hepatic systemic Salmonella infections (9, 10). Protective immunity is bacterial load. Using genome-wide expression microarrays in spleen further mediated by IFN-g production from activated CD4+ Th1 tissue from wild-type and Ity14 mutants, we studied the impact of cells as well as the cytotoxic response of CD8+ T lymphocytes Stat4 on the inflammatory immune response to Salmonella infec- (11). Indeed, immunodeficient mice lacking IFN-g or IFN-gR fail tion. We validated the importance of IFN-g–mediated immunity to resolve primary infection with an attenuated Salmonella strain during systemic Salmonella infection. We further show that NK and (12). Moreover, neutralization of IL-12 in a mouse model of ty- NKT cells play an important early role in controlling Salmonella in phoid fever abrogates host immunity to primary Salmonella in- Stat4Ity14/Ity14 mice. During oral infection using a model of intestinal fection (13). In addition, clinical reports