Negative selection in the thymus eliminates self-Ag–specific thymocytes, and inefficient selection has been linked to the development of T cell–mediated autoimmunity. To better understand the production of autoreactive T cells in the thymus, He et al. (p. 5858) transplanted thymic grafts from NOD mice of different ages into immunodeficient NOD.scid recipients. Mice receiving thymi from newborn mice developed diabetes, whereas those receiving grafts from 7- and 10-d-old NOD mice did not, despite developing a low level of insulitis. Interestingly, recipients of thymi from donors 10 d of age and older developed severe colitis characterized by T cells specific for colonic microbiota. Relative to those receiving older thymi, recipients of neonatal thymi demonstrated increased numbers of β cell–specific CD4+ and CD8+ T cells in their pancreatic lymph nodes (PLN). Changes in the frequency of autoreactive T cells did not result from alterations in regulatory T cell numbers or immunoregulation in the PLN, and the multiorgan inflammation observed following transfer of newborn thymi did not require the autoimmune regulator (AIRE). These data reveal a narrow temporal window for the thymic development of pancreas-specific T cells, which has implications for the understanding of both T cell development and autoimmunity.

Binding of collectin kidney 1 (CL-K1) to microbial molecular patterns, such as mannan or microbial DNA, initiates activation of MBL-associated serine proteases (MASPs) through the lectin binding pathway, which triggers C4 and downstream complement cascade events. However, many elements of this pathway remain undefined. In this issue, Henriksen et al. (p. 6117) identified a previously unknown binding partner for CL-K1, collectin liver 1 (CL-L1), present in plasma. They found that CL-L1 coprecipitated with CL-K1 proteins purified from plasma by EDTA-dependent affinity chromatography. Precipitates subjected to 2D-SDS-PAGE and deglycosylation analysis revealed that disulfide bridge linkages covalently bonded CL-L1 and CL-K1 in heteromeric complexes (CL-LK). Purified native high oligomeric CL-LK interacted with recombinant MASPs -1, -2, and -3 more efficiently than low oligomeric CL-LK in vitro, indicating that oligomerization played a crucial role in CL-LK activity. Sequential administration of CL-LK, MASP-2, and C4 to mannan- or DNA-coated plates resulted in increased C4b deposition on the plate surface compared with positive controls, demonstrating that CL-LK-MASP-2 interactions efficiently activated complement in vitro. These results provide important evidence of CL-LK heteromeric complexes circulating naturally in the plasma and suggest that these complexes may provide a potent means of triggering the lectin-binding pathway of complement activation.

The deoxycytidine deaminase APOBEC3G (A3G) catalyzes hypermutation in the HIV-1 genome, thereby restricting viral replication in T cells. A3G activity requires its encapsidation into HIV-1 virions lacking viral-encoded infectivity factor (Vif) through a process that remains unclear. The germinal center–associated nuclear protein (GANP) binds to and promotes the activity of activation-induced cytidine deaminase (AID), leading Maeda et al. (p. 6030) to hypothesize that GANP might also affect the activity of the related enzyme A3G. Indeed, GANP interacted with A3G in T cells, was encapsidated into HIV-1 virions along with A3G, and enhanced A3G localization into the core region of the virions. Within the virion core, GANP and A3G associated with viral genomic RNA, and GANP cooperated with A3G to modulate the packaging of cellular RNAs into virions. In vitro analysis of HIV-1 infectivity revealed that GANP augmented A3G-induced hypermutation of the viral genome and the resultant inhibition of viral infectivity. Conversely, knockdown or inhibition of endogenous GANP suppressed encapsidation of A3G and led to increased HIV-1 infectivity. The understanding of GANP-mediated control of A3G activity gleaned in this study may assist in the future development of HIV-1 vaccines.

Increased microbial drug resistance is a burgeoning threat to human health that requires new avenues of research to identify alternatives to antibiotic treatment. Increasingly, researchers have focused on exploiting evolutionarily ancient antimicrobial host defense proteins (HDPs) produced by immune cells and mucosal barriers to combat drug-resistant microbes. To that end, Campos-Salinas et al. (p. 6040) explored the antimicrobial properties of the neuropeptide urocortin II (UCN II), a molecule possessing neuroendocrine functions with structural and biochemical resemblance to many HDPs. Treating either gram-negative or -positive bacteria or the parasite Leishmania major with UCN II in vitro revealed that this peptide had potent bactericidal and leishmanicidal activity, exhibiting an EC50 below 5 μM. These antimicrobial properties depended upon mechanisms involving membrane depolarization in bacteria and membrane pore formation in L. major, but UCN II treatment did not affect the viability of mammalian cells in toxicity and hemolysis assays. UCN II also possessed bacterial killing abilities in vivo, as i.p. injection of this peptide into septic mice increased survival of these animals and reduced bacterial loads, inflammatory mediators, and disease-associated histopathology. Lastly, the authors found that macrophages (Mϕ) infected with bacteria produced high levels of UCN II compared with naive Mϕ, indicating that UCN II is released as an immune countermeasure to bacterial infection. These results suggest that the UCN II peptide is an evolutionarily conserved host defense molecule that has promising therapeutic potential as an antimicrobial agent.

Dendritic cell (DC)–mediated cross-presentation of extracellular Ags to CD8+ T cells is often key to the development of CD8+ T cell immunity to vaccines, but the details of how Ags access the MHC I processing pathway remain poorly defined. Heat shock proteins such as glucose-regulated protein 170 (Grp170) have shown promise as adjuvants for DC-targeted tumor vaccines in preclinical studies. To elucidate how Grp170 may enhance cross-presentation and thereby augment vaccine efficacy, Wang et al. (p. 6010) treated DCs with a protein complex vaccine consisting of Grp170 linked to the melanoma Ag Gp100. The Grp170-gp100 complex localized to the endoplasmic reticulum (ER) following endocytosis, possibly through fusion of endosomes with the ER. Grp170 directly interacted with components of the ER-associated degradation (ERAD) machinery, which is involved in retrotranslocation of proteins from the ER to the cytosol, where they are degraded by the proteasome. Both ERAD and the proteasome were found to be required for Gp100 processing and subsequent activation of Gp100-specific CD8+ T cells. TAP activity was also required, suggesting a mechanism by which processed Gp100 peptides might access MHC I. A T cell adoptive transfer model confirmed that the ERAD pathway was required for activation of CD8+ T cells by the Grp170-gp100 vaccine in vivo. This study presents a mechanism by which the chaperone Grp170 can facilitate cross-presentation and thus augment tumor vaccine efficacy, with important implications for future vaccination strategies.

Gene mutations in three prime repair exonuclease 1 (TREX1), the most abundant DNA exonuclease in mammalian cells, are linked to several autoimmune diseases, including Aicardi–Goutières syndrome and systemic lupus erythematosus. In mice, Trex1 deficiency causes systemic inflammation and chronic production of type-1 IFN that leads to inflammatory myocarditis and premature death, but how Trex1 regulates inflammatory processes in individual tissues and cells has not been fully elucidated. To shed light on these mechanisms, Pereira-Lopes et al. (p. 6128) characterized the inflammatory phenotype in tissues and immune cells of Trex1−/− mice. They found upregulation of inflammatory genes in multiple organs, including the brain, in Trex1−/− mice compared with wild-type (WT) controls. Bone marrow–derived macrophages (BMDMs) or dendritic cells stimulated with proinflammatory TLR4, 7, and 9 ligands increased expression of Trex1 compared with a control fibroblast cell line. Trex1−/− BMDMs treated with these same TLR ligands produced increased quantities of proinflammatory cytokines compared with WT BMDMs, indicating that Trex1 may negatively regulate inflammatory pathways in activated macrophages. Trex1−/− BMDMs primed overnight with TLR ligands and pulsed with the model Ag, OVA, had increased CD86 surface expression and caused increased proliferation of OVA-specific CD4+ T cells. TLR ligand treatment also caused impaired phagocytosis in Trex1−/− BMDMs compared with controls. Together, these data suggest that Trex1 may negatively regulate pathways in macrophages that promote inflammation and predispose individuals toward the development of autoimmunity.