Insertive Condom-Protected and Condomless Vaginal Sex Both Have a Profound Impact on the Penile Immune Correlates of HIV Susceptibility
by Avid Mohammadi, Sareh Bagherichimeh, Yoojin Choi, Azadeh Fazel, Elizabeth Tevlin, Sanja Huibner, Zhongtian Shao, David Zuanazzi, Jessica L. Prodger, Sara V. Good, Wangari Tharao & Rupert Kaul
Summary: In heterosexual men, the penis is the primary site of Human Immunodeficiency Virus (HIV) acquisition. Levels of inflammatory cytokines in the coronal sulcus are associated with an increased HIV risk, and we hypothesized that these may be altered after insertive penile sex. Therefore, we designed the Sex, Couples and Science Study (SECS study) to define the impact of penile–vaginal sex on the penile immune correlates of HIV susceptibility. We found that multiple coronal sulcus cytokines increased dramatically and rapidly after sex, regardless of condom use, with a return to baseline levels by 72 hours. The changes observed after condomless sex were strongly predicted by cytokine concentrations in the vaginal secretions of the female partner, and were similar in circumcised and uncircumcised men. We believe that these findings have important implications for understanding the immunopathogenesis of penile HIV acquisition; in addition, they have important implications for the design of clinical studies of penile HIV acquisition and prevention.
by Quang Vinh Phan, Boris Bogdanow, Emanuel Wyler, Markus Landthaler, Fan Liu, Christian Hagemeier & Lüder Wiebusch
Summary: Human cytomegalovirus (HCMV) infection is associated with systemic disease in immunocompromised individuals and congenitally infected neonates. Animal CMVs and their bacterial artificial chromosome (BAC) clones have been utilized as models for CMV infection and thereby contributed immensely to the understanding of pathogenesis, host immune response and underlying molecular mechanism of CMV infections. As the closest relative to HCMV, the chimpanzee CMV (CCMV) holds a great potential as a model system for HCMV infection but its application was limited due to the lack of tools and data for functional genomic analyses. Here, the cloning of the CCMV as a BAC vector made its viral genome available to gene targeting techniques that allow the efficient application of reverse genetic strategies. Furthermore, the multi-omic datasets created in this study provide an in-depth view of the viral gene repertoire and the host cell responses to infection, confirming the close phylogenetic relationship between HCMV and CCMV on a system level. Taken together, the newly established CCMV–BAC system presents a framework for HCMV modeling and comparative studies to address key questions in evolutionary processes and infection mechanisms.
RplI Interacts with 5′ UTR of exsA to Repress Its Translation and Type III Secretion System in Pseudomonas aeruginosa
by Dan Wang, Xinxin Zhang, Liwen Yin, Qi Liu, Zhaoli Yu, Congjuan Xu, Zhenzhen Ma, Yushan Xia, Jing Shi, Yuehua Gong, Fang Bai, Zhihui Cheng, Weihui Wu, Jinzhong Lin & Yongxin Jin
Summary: Ribosomes provide all living organisms the capacity to synthesize proteins. The production of many ribosomal proteins is often controlled by an autoregulatory feedback mechanism. P. aeruginosa is an opportunistic human pathogen and its type III secretion system (T3SS) is a critical virulence determinant in host infections. In this study, by screening a Tn5 mutant library, we identified rplI, encoding ribosomal large subunit protein L9, as a novel repressor for the T3SS. Further exploring the regulatory mechanism, we found that the RplI protein interacts with the 5’ UTR (5’ untranslated region) of exsA, a gene coding for transcriptional activator of the T3SS. Such an interaction likely blocks ribosome loading on the exsA 5’ UTR, inhibiting the initiation of exsA translation. The significance of this work is in the identification of a novel repressor for the T3SS and elucidation of its molecular mechanism. Furthermore, this work provides evidence for individual ribosomal protein regulating mRNA translation beyond its autogenous feedback control.
by Patrick Günther, Dennis Quentin, Shehryar Ahmad, Kartik Sachar, Christos Gatsogiannis, John C. Whitney & Stefan Raunser
Summary: Bacteria have developed a variety of strategies to compete for nutrients and limited resources. One system widely used by Gram-negative bacteria is the T6 secretion system which delivers a plethora of effectors into competing bacterial cells. Known functions of effectors are degradation of the cell wall, the depletion of essential metabolites such as NAD+ or the cleavage of DNA. RhsA is an effector from the widespread plant-protecting bacteria Pseudomonas protegens. We found that RhsA forms a closed cocoon similar to that found in bacterial Tc toxins and metazoan teneurin proteins. The effector cleaves its polypeptide chain by itself in three pieces, namely the N-terminal domain including a seal, the cocoon and the actual toxic component which potentially cleaves DNA. The toxic component is encapsulated in the large cocoon, so that the effector producing bacterium is protected from the toxin. In order for the toxin to exit the cocoon, we propose that the seal, which closes the cocoon at one end, is removed by mechanical forces during injection of the effector by the T6 secretion system. We further hypothesize about different scenarios for the delivery of the toxin into the cytoplasm of the host cell. Together, our findings expand the knowledge of the mechanism of action of the T6 secretion system and its essential role in interbacterial competition.
Non-Neutralizing Antibodies Targeting the Immunogenic Regions of HIV-1 Envelope Reduce Mucosal Infection and Virus Burden in Humanized Mice
by Catarina E. Hioe, Guangming Li, Xiaomei Liu, Ourania Tsahouridis, Xiuting He, Masaya Funaki, Jéromine Klingler, Alex F. Tang, Roya Feyznezhad, Daniel W. Heindel, Xiao-Hong Wang, David A. Spencer, Guangnan Hu, Namita Satija, Jérémie Prévost, Andrés Finzi, Ann J. Hessell, Shixia Wang, Shan Lu, Benjamin K. Chen, Susan Zolla-Pazner, Chitra Upadhyay, Raymond Alvarez & Lishan Su
Summary: In the past decade, HIV-1 has infected an estimated 1.5 to 2 million people every year, but vaccines needed to control this pandemic are unavailable. Among vaccines tested in the human efficacy trials, the RV144 vaccine regimen showed a modest efficacy and revealed non-neutralizing antibodies against the virus envelope glycoproteins as a correlate of reduced virus acquisition. To design more efficacious HIV-1 vaccines, a better understanding about antiviral mechanisms of these antibodies is needed. Here non-neutralizing monoclonal antibodies against two immunogenic sites on the virus envelope were evaluated for passive administration to humanized mice that were subsequently challenged with HIV-1. The antibodies did not block mucosal HIV-1 infection but reduced virus burden. The level of virus reduction correlated with the antibody binding potency and the effector functions mediated through their Fc fragments, which included antibody-dependent phagocytosis and complement activation, but not the commonly studied antibody-dependent cellular cytotoxicity. The importance of the Fc functions was further demonstrated by reduced virus control when mutations were introduced to decrease Fc activities. This study provides new evidence for the important contribution of multiple Fc-dependent antibody functions in immune control against HIV-1.
Variability in an Effector Gene Promoter of a Necrotrophic Fungal Pathogen Dictates Epistasis and Effector-Triggered Susceptibility in Wheat
by Evan John, Silke Jacques, Huyen T. T. Phan, Lifang Liu, Danilo Pereira, Daniel Croll, Karam B. Singh, Richard P. Oliver & Kar-Chun Tan
Summary: Breeding for durable resistance to fungal diseases in crops is a continual challenge for crop breeders. Fungal pathogens evolve ways to overcome host resistance by masking themselves through effector evolution and evasion of broad-spectrum defense responses. Association studies on mapping populations infected by isolate mixtures are often used by researchers to seek out novel sources of genetic resistance. Disease resistance quantitative trait loci (QTL) are often minor or inconsistent across environments. This is a particular problem with septoria diseases of cereals such as septoria nodorum blotch (SNB) of wheat caused by Parastagonospora nodorum. The fungus uses a suite of necrotrophic effectors (NEs) to cause SNB. We characterized a genetic element, called PE401, in the promoter of the major NE gene Tox1, which is present in some P. nodorum isolates. PE401 functions as a transcriptional repressor of Tox1 and exerts epistatic control on another major SNB resistance QTL in the host. In the context of crop protection, constant surveillance of the pathogen population for the frequency of PE401 in conjunction with NE diversity will enable agronomists to provide the best advice to growers on which wheat varieties can be tailored to provide optimal SNB resistance to regional pathogen population genotypes.
Mutational Analysis of Aedes aegypti Dicer 2 Provides Insights into the Biogenesis of Antiviral Exogenous Small Interfering RNAs
by Rommel J. Gestuveo, Rhys Parry, Laura B. Dickson, Sebastian Lequime, Vattipally B. Sreenu, Matthew J. Arnold, Alexander A. Khromykh, Esther Schnettler, Louis Lambrechts, Margus Varjak & Alain Kohl
Summary: Aedes aegypti mosquitoes that transmit human-pathogenic viruses rely on the exogenous small interfering RNA (exo-siRNA) pathway as part of antiviral responses. This pathway is triggered by virus-derived double-stranded RNA (dsRNA) produced during viral replication that is then cleaved by Dicer 2 (Dcr2) into virus-derived small interfering RNAs (vsiRNAs). These vsiRNAs target viral RNA, leading to suppression of viral replication. The importance of Dcr2 in this pathway has been intensely studied in the Drosophila melanogaster model but is largely lacking in mosquitoes. Here, we have identified conserved and functionally relevant amino acids in the helicase and RNase III domains of Ae. aegypti Dcr2 that are important in its silencing activity and antiviral responses against Semliki Forest virus (SFV). Small RNA sequencing of SFV-infected mosquito cells with functional or mutated Dcr2 gave new insights into the nature and origin of vsiRNAs. The findings of this study, together with the different molecular tools we have previously developed to investigate the exo-siRNA pathway of mosquito cells, have started to uncover important properties of Dcr2 that could be valuable in understanding mosquito-arbovirus interactions and potentially in developing or assisting vector control strategies.
Probing the Structure and Function of the Protease Domain of Botulinum Neurotoxins Using Single-Domain Antibodies
by Kwok-ho Lam, Jacqueline M. Tremblay, Kay Perry, Konstantin Ichtchenko, Charles B. Shoemaker & Rongsheng Jin
Summary: Botulinum neurotoxins (BoNTs) are extremely toxic to humans by causing flaccid paralysis of botulism. The catalytic light chain (LC) of BoNTs is the warhead of the toxin, which is mainly responsible for BoNT’s neurotoxic effects. As an endopeptidase, LC is delivered by the toxin to inside neurons where it specifically cleaves neuronal SNARE proteins and causes muscle paralysis. While the currently available equine and human antitoxin sera can prevent further intoxication, they do not promote recovery from paralysis that has already occurred. We strike to develop single-domain variable heavy-chain (VHH) antibodies targeting the LC of BoNT/A (LC/A) and BoNT/B (LC/B) as antidotes to inhibit or eliminate the intraneuronal LC protease. Here, we report the identification and characterization of large panels of new and unique VHHs that bind to LC/A or LC/B. Using a combination of X-ray crystallography and biochemical assays, we reveal that VHHs exploit diverse mechanisms to interact with LC/A and LC/B and inhibit their protease activity, and such knowledge can be harnessed to predict their specificity towards different toxin subtypes within each serotype. We anticipate that the new VHHs and their characterization reported here will contribute to the development of improved botulism therapeutics having high potencies and broad specificities.
by Clinton O. Ogega, Nicole E. Skinner, Andrew I. Flyak, Kaitlyn E. Clark, Nathan L. Board, Pamela J. Bjorkman, James E. Crowe Jr., Andrea L. Cox, Stuart C. Ray & Justin R. Bailey
Summary: Antiviral immunity relies on production of protective immunoglobulin G (IgG) by B cells, but many hepatitis C virus (HCV)-infected individuals have very low levels of HCV-specific IgG in their serum. Elucidating mechanisms underlying this suboptimal IgG expression remains paramount in guiding therapeutic and vaccine strategies. In this study, we developed a highly specific method to capture HCV-specific B cells and characterized their surface protein expression. Two proteins analyzed were Fc receptor-like protein 5 (FCRL5), a cell surface receptor for IgG, and programmed cell death protein-1 (PD-1), a marker of lymphocyte activation and exhaustion. We measured serum levels of anti-HCV IgG in these subjects and demonstrated that overexpression of FCRL5 and PD-1 on memory B cells was associated with reduced anti-E2 IgG levels. This study uses HCV as a viral model, but the findings may be applicable to many viral infections, and they offer new potential targets to enhance antiviral IgG production.