Risk, severity, and outcome of infection depend on the interplay of pathogen virulence and host susceptibility. Systematic identification of genetic susceptibility to infection is being undertaken through genome-wide association studies, but how to expeditiously move from genetic differences to functional mechanisms is unclear. Here, we use genetic association of molecular, cellular, and human disease traits and experimental validation to demonstrate that genetic variation affects expression of VAC14, a phosphoinositide-regulating protein, to influence susceptibility to serovar Typhi ( Typhi) infection. Decreased VAC14 expression increased plasma membrane cholesterol, facilitating docking and invasion. This increased susceptibility at the cellular level manifests as increased susceptibility to typhoid fever in a Vietnamese population. Furthermore, treating zebrafish with a cholesterol-lowering agent, ezetimibe, reduced susceptibility to Typhi. Thus, coupling multiple genetic association studies with mechanistic dissection revealed how VAC14 regulates invasion and typhoid fever susceptibility and may open doors to new prophylactic/therapeutic approaches.

Salmonella hijack host machinery in order to invade cells and establish infection. While considerable work has described the role of host proteins in invasion, much less is known regarding how natural variation in these invasion-associated host proteins affects Salmonella pathogenesis. Here we leveraged a candidate cellular GWAS screen to identify natural genetic variation in the ARHGEF26 (Rho Guanine Nucleotide Exchange Factor 26) gene that renders lymphoblastoid cells susceptible to Salmonella Typhi and Typhimurium invasion. Experimental follow-up redefined ARHGEF26’s role in Salmonella epithelial cell infection. Specifically, we identified complex serovar-by-host interactions whereby ARHGEF26 stimulation of S. Typhi and S. Typhimurium invasion into host cells varied in magnitude and effector-dependence based on host cell type. While ARHGEF26 regulated SopB- and SopE-mediated S. Typhi (but not S. Typhimurium) infection of HeLa cells, the largest effect of ARHGEF26 was observed with S. Typhimurium in polarized MDCK cells through a SopB- and SopE2-independent mechanism. In both cell types, knockdown of the ARHGEF26-associated protein DLG1 resulted in a similar phenotype and serovar specificity. Importantly, we show that ARHGEF26 plays a critical role in S. Typhimurium pathogenesis by contributing to bacterial burden in the enteric fever murine model, as well as inflammation in the colitis infection model. In the enteric fever model, SopB and SopE2 are required for the effects of Arhgef26 deletion on bacterial burden, and the impact of sopB and sopE2 deletion in turn required ARHGEF26. In contrast, SopB and SopE2 were not required for the impacts of Arhgef26 deletion on colitis. A role for ARHGEF26 on inflammation was also seen in cells, as knockdown reduced IL-8 production in HeLa cells. Together, these data reveal pleiotropic roles for ARHGEF26 during infection and highlight that many of the interactions that occur during infection that are thought to be well understood likely have underappreciated complexity.

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Salmonella hijack host machinery in order to invade cells and establish infection. While considerable work has described the role of host proteins in invasion, much less is known regarding how natural variation in these invasion-associated host proteins affects Salmonella pathogenesis. Here we leveraged a candidate cellular GWAS screen to identify natural genetic variation in the ARHGEF26 (Rho Guanine Nucleotide Exchange Factor 26) gene that renders lymphoblastoid cells susceptible to Salmonella Typhi and Typhimurium invasion. Experimental follow-up redefined ARHGEF26’s role in Salmonella epithelial cell invasion, identified serovar specific interactions, implicated ARHGEF26 in SopE-mediated invasion, and revealed that the ARHGEF26-associated proteins DLG1 and SCRIB facilitate S. Typhi uptake. Importantly, we show that ARHGEF26 plays a critical role in S. Typhimurium pathogenesis by contributing to bacterial burden in the enteric fever murine model, as well as inflammation in the gastroenteritis infection model. The impact of ARHGEF26 on inflammation was also seen in cells, as knockdown reduced IL-8 production in HeLa cells. Together, these data reveal pleiotropic roles for ARHGEF26 function during infection and highlight that many of the interactions that occur during infection that are thought to be well understood likely have underappreciated complexity.Author SummaryDuring infection, Salmonella manipulates host cells into engulfing the bacteria and establishing an intracellular niche. While many studies have identified genes involved in different stages of this Salmonella invasion process, few studies have examined how differences between human hosts contribute to infection susceptibility. Here we leveraged a candidate genetic screen to identify natural genetic variation in the human ARHGEF26 gene that correlates with Salmonella invasion. Springboarding from this result, we experimentally tested and revised existing models of ARHGEF26’s role in Salmonella invasion, discovered an additional new role for ARHGEF26 during Salmonella disease, and confirmed our findings in mouse models. Building on how ARHGEF26 functions in other contexts, we implicated two ARHGEF26-interacting host proteins as contributors to Salmonella pathobiology. Collectively, these results identify a potential source of inter-person diversity in susceptibility to Salmonella disease, expand our molecular understanding of Salmonella infection to include a multifaceted role for ARHGEF26, and identify several important future directions that will be important to understand how Salmonella recruit and manipulate ARHGEF26 as well as how ARHGEF26 is able to drive Salmonella-beneficial processes.

25Clearance of intracellular pathogens, such as Leishmania (L.) major, depends on 26 a well-regulated adaptive T cell response. Here we describe a pathogen-encoded 27 mechanism to alter T cell recruitment by suppressing CXCL10, a chemokine that recruits 28 CD4+ and CD8+ T cells by signaling through the CXCR3 receptor. L. major suppresses 29 CXCL10 through the virulence factor and protease, glycoprotein-63 (gp63). GP63 cleaves 30 CXCL10 after amino acid A81, impairing T-cell chemotaxis in vitro. GP63 from either 31 extracellular promastigotes or intracellular amastigotes is capable of cleaving CXCL10. 32Consistent with CXCL10 cleavage during infection, we observed GP63-mediated 33 impairment of activated CD8+ T-cells in the draining lymph nodes of C57BL/6J WT mice. 34Correspondingly, in C57BL/6J WT mice, gp63 deletion resulted in slower lesion 35 development and a smaller maximum lesion size. However, infection in C57BL/6J cxcr3-/-36 mice revealed the delay to lesion development required CXCR3 signaling. Finally, 37Salmonella enterica and Chlamydia trachomatis infection also suppress CXCL10, 38 demonstrating convergent evolution of this immune evasion strategy in diverse 39 intracellular pathogens. Understanding mechanisms of CXCL10 evasion may facilitate 40 the development of novel therapeutic strategies to treat intracellular pathogens. 41 42 Author Summary 45Leishmaniasis is an infectious disease that affects over one million people 46 annually. It is caused by intracellular parasites that have evolved to evade the host's 47 attempts to eliminate the parasite. The most common form of disease, cutaneous 48 leishmaniasis, causes disfiguring skin lesions if the host immune system does not 49 appropriately respond to the infection. A family of molecules called chemokines are critical 50 for the recruitment of specific subsets of immune cells required to eliminate infection. 51Here, we demonstrate a novel mechanism that the parasite Leishmania (L.) major 52 employs to disrupt the immune response by interfering with chemokine signaling. By this 53 strategy, L. major uses a protease to cleave chemokines known to recruit T-cells required 54 for fighting off infection. We show that L. major cleavage of these immune signals changes 55 the recruitment of CD8+ T-cells and alters disease progression in mice. Finally, we 56 observe that multiple common human intracellular pathogens, including Chlamydia 57 trachomatis and Salmonella enterica, interfere with the same chemokines, suggesting a 58 strong selective pressure to avoid this component of the immune response. Our study 59 provides new insights into how intracellular pathogens interact with the host immune 60 response to enhance pathogen survival and exacerbate disease outcomes. 61Leishmania, the etiologic agent for leishmaniasis, serve as a paradigm for studying T cell 65 polarization in response to intracellular pathogens [1]. However, persistent gaps in the 66 understanding of host and pathogen factors that influence T cell polarization and immune 67 cell recruitment...