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Previews the intramolecular NBS/CBS interaction, which potentiates the pathogenic scaffold/hub function of CagA. Finally, the authors investigated CagAphosphatidylserine (PS) interactions, as this influences the transport of CagA and its localization to the inner plasma membrane (Murata-Kamiya et al., 2010). Two arginines (R624 and R626), crucial for the CagA-PS interaction, were located in the a18 helix of Domain-II. Structural and mutational analyses identified that R624 and R626 form a basic amino acid cluster with several lysine residues (position 613, 614, 617, 621, 625, 631, and 635), providing a positive electrostatic surface potential important for CagA-PS docking. The molecular orientation of delivered CagA relative to the plasma membrane was also deduced from these studies. Taken together, the above findings provide a comprehensive structural picture of the N terminus of CagA. The tertiary structure of CagA1-876 is unique, with no homologous other structure yet reported. Structure-function data reported here clearly indicate that the N terminus may

serve as a regulatory element for the C terminus. However, the current CagA1-876 structure still has gaps due to resolution problems. Thus, more efforts are necessary to obtain high-resolution crystal structures of the N and C terminus and CagAFL. It will be also highly interesting to investigate in future studies how other known interaction partners including RUNX3, integrin-b1, apoptosisstimulating protein of p53 (ASPP2), CagF, and others interact with the individual segments of CagA1-876 and influence its function. Efforts to cocrystallize CagA with these or other interaction partners may be helpful here. The present studies of Hayashi et al. (2012) provide important knowledge for the understanding of H. pylori-triggered pathology and developing inhibitors for more effective new therapy against this important pathogen.

Bagnoli, F., Buti, L., Tompkins, L., Covacci, A., and Amieva, M.R. (2005). Proc. Natl. Acad. Sci. USA 102, 16339–16344. Hatakeyama, M. (2008). Curr. Opin. Microbiol. 11, 30–37. Hayashi, T., Senda, M., Morohashi, H., Higashi, H., Horio, M., Kashiba, Y., Nagase, L., Sasaya, D., Shimizu, T., Venugopalan, N., et al. (2012). Cell Host Microbe 12, this issue, 20–33. Jime´nez-Soto, L.F., Kutter, S., Sewald, X., Ertl, C., Weiss, E., Kapp, U., Rohde, M., Pirch, T., Jung, K., Retta, S.F., et al. (2009). PLoS Pathog. 5, e1000684. Kwok, T., Zabler, D., Urman, S., Rohde, M., Hartig, R., Wessler, S., Misselwitz, R., Berger, J., Sewald, N., Ko¨nig, W., and Backert, S. (2007). Nature 449, 862–866. Moese, S., Selbach, M., Zimny-Arndt, U., Jungblut, P.R., Meyer, T.F., and Backert, S. (2001). Proteomics 1, 618–629. Mueller, D., Tegtmeyer, N., Brandt, S., Yamaoka, Y., De Poire, E., Sgouras, D., Wessler, S., Torres, J., Smolka, A., and Backert, S. (2012). J. Clin. Invest. 122, 1553–1566.

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NETs Tangle with HIV Craig N. Jenne1,2 and Paul Kubes1,3,* 1Calvin,

Phoebe & Joan Snyder Institute for Chronic Diseases of Critical Care Medicine 3Department of Physiology and Pharmacology University of Calgary, Calgary, Alberta, Canada T2N 4N1 *Correspondence: [email protected] http://dx.doi.org/10.1016/j.chom.2012.07.002 2Department

Neutrophil extracellular traps (NETs) are comprised of extracellular DNA coated in cytotoxic proteins capable of ensnaring and killing bacteria. Saitoh et al. (2012) expand our understanding of NETs into antiviral immunity by demonstrating that HIV induces the formation of NETs, which can bind and neutralize viral particles. Neutrophils are the foot soldiers of the innate immune response. These cells are often the first to respond to pathogens, arriving rapidly and deploying a vast array of antimicrobial agents in an effort to control the early phases of infection. This arsenal of effector mechanisms include the ability to phagocytose pathogens and the release of large numbers of anti-

microbial peptides, cytokines, and reactive oxygen species (ROS). More recently, a novel effector mechanism has been described whereby in response to a pathogen, the nuclear DNA of the neutrophil decondenses and is released from the cell to form a large sticky web. These Neutrophil extracellular traps (NETs) are decorated with histones—and antimicro-

bial proteins such as neutrophil elastase, myeloperoxidase (MPO), and a-defensin (Brinkmann et al., 2004). Furthermore, NETs have been demonstrated to both ensnare and directly kill a number of different bacteria. Although viral infection has classically been studied in the context of the adaptive immune response, a significant

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amount of evidence exists to escape NETs. The best charsuggest a critical role for the acterized of these strategies innate immune system in is the production of nucleases the host antiviral immune by some bacterial strains to response. The innate immune digest NET structures (Busystem detects viral pathchanan et al., 2006). The ogen-associated molecular expression of these nuclepatterns (PAMPs) through a ases contributes to bacterial diverse array of pattern revirulence and facilitates the cognition receptors (PRR), evasion of the host innate including Toll-like receptor immune response. (TLR)-3, 7, and 8, as well Importantly, in the current as RIG-I and MDA5, sugstudy the authors also identigesting the evolution of fied a means by which HIV is an antiviral response within able to regulate NET producthis arm of the immune tion by neutrophils; however, system. Furthermore, neutrorather than targeting the phils themselves have been neutrophil or NETs directly, demonstrated to be essential HIV modulates the local Figure 1. Interactions between HIV, Neutrophils, DC, and NETs for the early control of highly immune microenvironment. Upon detection of HIV through either TLR7 or TLR8, neutrophils initiate ROS pathogenic influenza infecSaitoh et al. (2012) demongeneration. The production of ROS results in the decondensation of the neutrophil nucleus and the eventual release of NETs. NETs are capable of entions; however, the specific strate that HIV is able to bind snaring HIV virions and, through the action of granular proteins bound to the mechanisms involved in this to the surface of dendritic DNA strands, neutralizing the viral particles. HIV is not simply the target of antiviral response remained cells (DC) via CD209 (DCthis innate immune response but rather actively modulates the process. Viral binding to CD209 on DC results in the production of IL-10 by the DC. IL-10 unclear (Tate et al., 2009). SIGN), a C-type lectin remodulates neutrophil function, reducing ROS production and attenuating In this issue, Saitoh et al. ceptor, and induce the proNET release. (2012) link the production of duction of IL-10, a potent NETs by human neutrophils, anti-inflammatory cytokine. previously thought of as an antibacterial induce NET production by neutrophils is Treatment of activated neutrophils with effector mechanism, to the host antiviral dependent on TLR-7, TLR-8, and the supernatants from HIV-treated DC or response. In a series of elegant experi- generation of ROS (Figure 1). These find- with purified IL-10 attenuated NET proments, Saitoh et al. (2012) are able to ings are particularly interesting, as the duction and significantly reduced the demonstrate that neutrophils release initiation of NET release by bacteria or virucidal capacity of human neutrophils. NETs in response to exposure to HIV bacterial ligands is also dependent on It is important to note that these studies virions alone, and that these extracellular ROS production (Fuchs et al., 2007). were carried out using an in vitro cellDNA structures are able to bind and, These findings suggest that regardless culture system. Although the work by more importantly, directly inactivate the of the specific nature of the pathogen Saitoh et al. (2012) beautifully characHIV virus. This viral neutralization was (bacterial versus viral), stimulation of terize the antiviral potential of NETs and dependent on the presence of MPO and neutrophils through one of a number of the complex dynamic that exists between a-defensin within the NET structure, as different PRR can result in the activation HIV, DC, and neutrophils, it will be impordestruction of NETs with DNase, inhibition of a common innate effector response, tant to understand how these mechaof MPO, or blocking antibodies against culminating in the dissolution of the nisms function within the context of the a-defensin abrogated the ability of the neutrophil’s nuclei and the release of host immune response to viruses in vivo. neutrophils to neutralize HIV. Using NETs in an effort to ensnare and neu- The authors propose that this interaction superresolution structured illumination tralize the invading pathogen. It remains could occur in the mucosa, but one could microscopy (SR-SIM), Saitoh et al. (2012) unclear from this study whether this also imagine NETs being formed in blood were able to capture striking 3D images NET production actually led to neutrophil as was reported by Clark et al. (2007). In of NETs at a much higher resolution death or whether neutrophils maintained fact, trying to catch viruses under flow than is possible with conventional fluo- viability and helped to eliminate NET conditions could be quite difficult, but rescence microscopy. The images reveal caught pathogens. the release of NETs could facilitate the sheets of DNA rather than the previously This interaction between the neutrophil capture of many viral particles by a single reported strands or strings. Moreover, and the virus is not a one-way street cell. Moreover, the efficacy of this novel these studies clearly illustrate the intri- whereby the neutrophil simply reacts antiviral response is dependent on the cacy of the extracellular DNA structures to the pathogen but rather a complex ability of neutrophils to deploy their and directly visualize the capture of intact and dynamic relationship in which the NETs in such a manner that they are virus particles by NETs. pathogen attempts to modulate the neu- able to ensnare free virus. This mechaThrough the use of a series of chemical trophil’s response. As with most host- nism has great potential in limiting viral and protein inhibitors, Saitoh et al. (2012) effector processes, pathogens have dissemination but may be of limited value further demonstrate the ability of HIV to evolved strategies and mechanisms to in dealing with virally infected host cells. 6 Cell Host & Microbe 12, July 19, 2012 ª2012 Elsevier Inc.

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Previews Furthermore, as indicated by the authors, some viral infections induce neutropenia and, as such, these viruses may be able to limit the antiviral potential of the neutrophil response. Regardless of the potential complexities of the in vivo response, the study by Saitoh et al. (2012) represents a de facto antiviral response and, as suggested by the authors, also represents an exciting opportunity for the development of new therapeutic approaches to viral infections. Strategies targeting neutrophils, and specifically the generation of NETs, may prove to be important tools in limiting viral replication and spread. However, one must be cautious in developing NETs as an antiviral therapy. NETs, like most immune effector mechanisms, are only effective if they are a component of a balanced response. Recent studies have indicated excess NET formation can result in host tissue damage and may be associated with a number of human disease states (Clark et al., 2007; Narasaraju et al., 2011; Thomas et al., 2012; Leffler et al., 2012). Future immunother-

apies directed at controlling viral infections through the generation of NETs will have to strike a balance between viral neutralization and host tissue damage. These findings by Saitoh et al. (2012) open an entirely new chapter in our understanding of the innate immune response to viruses and blur the lines between antiviral and antimicrobial immunity. Furthermore, this study represents a unique opportunity for the development of antiviral therapies, particularly when viral infections are found in the context of neutrophilic inflammation.

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ACKNOWLEDGMENTS We thank H. Neto, B. Petri, and C.H. Wong for their critical review of this manuscript. C.N.J. is supported by Alberta Innovates Health Solutions.

Saitoh, T., Komano, J., Saitoh, Y., Misawa, T., Takahama, M., Kozaki, T., Uehata, T., Iwasaki, H., Omori, H., Yamaoka, S., Yamamoto, N., and Akira, S. (2012). Cell Host & Microbe 12, this issue, 109–116. Tate, M.D., Deng, Y.M., Jones, J.E., Anderson, G.P., Brooks, A.G., and Reading, P.C. (2009). J. Immunol. 183, 7441–7450.

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