The gastrointestinal tract is constantly challenged by foreign antigens and commensal bacteria but nonetheless is able to maintain a state of immunological quiescence. Recent advances have highlighted the importance of active suppression by regulatory lymphocytes and immunosuppressive cytokines in controlling mucosal immunity. Failures of these mechanisms contribute to the development of inflammatory bowel disease, but how these regulatory networks are established remains unclear. Here, we demonstrate key roles for ␣v integrins in regulation of mucosal immunity. We report that deletion of ␣v in the immune system causes severe colitis, autoimmunity, and cancer. Mice lacking immune cell ␣v have fewer regulatory T (Treg) cells in the colon and corresponding increases in activated T cells and T cell cytokine production, leading to colitis. Using conditional gene targeting, we demonstrate that this is specifically attributable to loss of ␣v from myeloid cells. Furthermore, we show that gut-associated macrophages and dendritic cells fail both to remove apoptotic cells efficiently and to induce Treg cells. Our results identify a vital role for myeloid ␣v integrins in generating mucosal Treg cells and emphasize the importance of antigen-presenting cells in establishing immune tolerance.immunoregulation ͉ phagocytosis ͉ apoptotic ͉ dendritic cell ͉ regulatory T cell I ntegrins are dimeric cell-surface receptors composed of ␣ and ␤ subunits (1). ␣v is the most promiscuous ␣ subunit, associating with five different ␤ subunits (␤1, ␤3, ␤5, ␤6, and ␤8) and participates in many important cellular processes, including cell adhesion, migration, survival, and growth factor signaling. ␣v integrins are expressed in many tissues throughout development, and ␣v knockout mice die at midgestation from placental deficiencies or, immediately after birth, with cerebral and intestinal hemorrhage and cleft palate (2). The lethality of ␣v Ϫ/Ϫ mice is attributable mainly to loss of ␣v␤8, because ␤8 Ϫ/Ϫ mice show similar placental and neural vasculature abnormalities (3), and these defects are principally attributable to loss of ␣v␤8 from parenchymal tissues supporting blood vessels (4, 5). Knockouts of most other ␣v-associated ␤ integrins (␤3, ␤5, and ␤6) are viable and reveal roles for ␣v in repair of tissue injury, inflammation, and immune responses (1, 6-8). ␣v␤6 and ␣v␤8 are expressed in epithelium, where their functions include binding and activating TGF-␤ (9), and ␣v␤8 is also expressed by myeloid cells and T cells (10, 11). ␣v␤3 and ␣v␤5 are more widely expressed and are up-regulated by many cells after tissue injury. In the immune system, ␣v␤3 and ␣v␤5 are expressed by many different cell types and promote cell adhesion, migration, and survival. ␣v integrins also mediate the uptake of apoptotic cells (12-14), a process central to immune regulation and inflammation resolution (15).Here, we report that ␣v integrins are central to the normal regulation of immune responses in the intestine and that deletion of ␣v in the immune system lead...

SUMMARY Dengue is the most common vector-borne viral disease, causing nearly 400 million infections yearly. Currently there are no approved therapies. Antibody epitopes that elicit weak humoral responses may not be accessible by conventional B cell panning methods. To demonstrate an alternative strategy to generating a therapeutic antibody, we employed a non-immunodominant, but functionally relevant, epitope in domain III of the E protein, and engineered by structure-guided methods an antibody directed to it. The resulting antibody, Ab513, exhibits high-affinity binding to, and broadly neutralizes, multiple genotypes within all four serotypes. To assess therapeutic relevance of Ab513, activity against important human clinical features of dengue was investigated. Ab513 mitigates thrombocytopenia in a humanized mouse model, resolves vascular leakage, reduces viremia to nearly undetectable levels, and protects mice in a maternal transfer model of lethal antibody-mediated enhancement. The results demonstrate that Ab513 may reduce the public health burden from dengue.

To combat antimicrobial infections, new active molecules are needed. Antimicrobial peptides, ever abundant in nature, are a fertile starting point to develop new antimicrobial agents but suffer from low stability, low specificity, and off-target toxicity. These drawbacks have limited their development. To overcome some of these limitations, we developed antibody-bactericidal macrocyclic peptide conjugates (ABCs), in which the antibody directs the bioactive macrocyclic peptide to the targeted Gram-negative bacteria. We used cysteine S Ar chemistry to synthesize and systematically study a library of large (>30-mer) macrocyclic antimicrobial peptides (mAMPs) to discover variants with extended proteolytic stability in human serum and low hemolytic activity while maintaining bioactivity. We then conjugated, by using sortase A, these bioactive variants onto an Escherichia coli targeted monoclonal antibody. We found that these ABCs had minimized hemolytic activity and were able to kill E. coli at nanomolar concentrations. Our findings suggest macrocyclic peptides if fused to antibodies may facilitate the discovery of new agents to treat bacterial infections.

IgA nephropathy (IgAN) is the most prevalent cause of primary glomerular disease worldwide, and the cytokine A PRoliferation‐Inducing Ligand (APRIL) is emerging as a key player in IgAN pathogenesis and disease progression. For a panel of anti‐human APRIL antibodies with known antagonistic activity, we sought to define their structural mode of engagement to understand molecular mechanisms of action and aid rational antibody engineering. Reliable computational prediction of antibody‐antigen complexes remains challenging, and experimental methods such as X‐ray co‐crystallography and cryoEM have considerable technical, resource, and throughput barriers. To overcome these limitations, we implemented an integrated and accessible experimental‐computational workflow to more accurately predict structures of antibody‐APRIL complexes. Specifically, a yeast surface display library encoding site‐saturation mutagenized surface positions of APRIL was screened against a panel of anti‐APRIL antibodies to rapidly obtain a comprehensive biochemical profile of mutational impact on binding for each antibody. The experimentally derived mutational profile data were used as quantitative constraints in a computational docking workflow optimized for antibodies, resulting in robust structural models of antibody‐antigen complexes. The model results were confirmed by solving the cocrystal structure of one antibody‐APRIL complex, which revealed strong agreement with our model. The models were used to rationally select and engineer one antibody for cross‐species APRIL binding, which quite often aids further testing in relevant animal models. Collectively, we demonstrate a rapid, integrated computational‐experimental approach to robustly predict antibody‐antigen structures information, which can aid rational antibody engineering and provide insights into molecular mechanisms of action.