Dynamics of Cas10 Govern Discrimination between Self and Non-self in Type III CRISPR-Cas Immunity.

Adaptive immune systems must accurately distinguish between self and non-self in order to defend against invading pathogens while avoiding autoimmunity. Type III CRISPR-Cas systems employ guide RNA to recognize complementary RNA targets, which triggers the degradation of both the invader's transcripts and their template DNA. These systems can broadly eliminate foreign targets with multiple mutations but circumvent damage to the host genome. To explore the molecular basis for these features, we use single-molecule fluorescence microscopy to study the interaction between a type III-A ribonucleoprotein complex and various RNA substrates. We find that Cas10-the DNase effector of the complex-displays rapid conformational fluctuations on foreign RNA targets, but is locked in a static configuration on self RNA. Target mutations differentially modulate Cas10 dynamics and tune the CRISPR interference activity in vivo. These findings highlight the central role of the internal dynamics of CRISPR-Cas complexes in self versus non-self discrimination and target specificity.

Immunoglobulin genes, reproductive isolation and vertebrate speciation.

Reproductive isolation drives the formation of new species, and many genes contribute to this through Dobzhansky-Muller incompatibilities (DMIs). These incompatibilities occur when gene divergence affects loci encoding interacting products such as receptors and their ligands. We suggest here that the nature of vertebrate immunoglobulin (IG) genes must make them prone to DMIs. The genes of these complex loci form functional genes through the process of recombination, giving rise to a repertoire of heterodimeric receptors of incredible diversity. This repertoire, within individuals and within species, must defend against pathogens but must also avoid pathogenic self-reactivity. We suggest that this avoidance of autoimmunity is only achieved through a coordination of evolution between heavy- and light-chain genes, and between these genes and the rest of the genome. Without coordinated evolution, the hybrid offspring of two diverging populations will carry a heavy burden of DMIs, resulting in a loss of fitness. Critical incompatibilities could manifest as incompatibilities between a mother and her divergent offspring. During fetal development, biochemical differences between the parents of hybrid offspring could make their offspring a target of the maternal immune system. This hypothesis was conceived in the light of recent insights into the population genetics of IG genes. This has suggested that antibody genes are probably as susceptible to evolutionary forces as other parts of the genome. Further repertoire studies in human and nonhuman species should now help determine whether antibody genes have been part of the evolutionary forces that drive the development of species.

Innate immune recognition of double-stranded RNA triggers increased expression of NKG2D ligands after virus infection.

Self/non-self-discrimination by the innate immune system relies on germline-encoded, non-rearranging receptors expressed by innate immune cells recognizing conserved pathogen-associated molecular patterns. The natural killer group 2D (NKG2D) receptor is a potent immune-activating receptor that binds human genome-encoded ligands, whose expression is negligible in normal tissues, but increased in stress and disease conditions for reasons that are incompletely understood. Here it is not clear how the immune system reconciles receptor binding of self-proteins with self/non-self-discrimination to avoid autoreactivity. We now report that increased expression of NKG2D ligands after virus infection depends on interferon response factors activated by the detection of viral double-stranded RNA by pattern-recognition receptors (RIG-I/MDA-5) and that NKG2D ligand up-regulation can be blocked by the expression of viral dsRNA-binding proteins. Thus, innate immunity-mediated recognition of viral nucleic acids triggers the infected cell to release interferon for NK cell recruitment and to express NKG2D ligands to become more visible to the immune system. Finally, the observation that NKG2D-ligand induction is a consequence of signaling by pattern-recognition receptors that have been selected over evolutionary time to be highly pathogen-specific explains how the risks of autoreactivity in this system are minimized.

Ubiquitin-proteasome-mediated degradation of S-RNase in a solanaceous cross-compatibility reaction.

Many plants have a self-incompatibility (SI) system in which the rejection of self-pollen is determined by multiple haplotypes at a single locus, termed S. In the Solanaceae, each haplotype encodes a single ribonuclease (S-RNase) and multiple S-locus F-box proteins (SLFs), which function as the pistil and pollen SI determinants, respectively. S-RNase is cytotoxic to self-pollen, whereas SLFs are thought to collaboratively recognize non-self S-RNases in cross-pollen and detoxify them via the ubiquitination pathway. However, the actual mechanism of detoxification remains unknown. Here we isolate the components of a SCF(SLF) (SCF = SKP1-CUL1-F-box-RBX1) from Petunia pollen. The SCF(SLF) polyubiquitinates a subset of non-self S-RNases in vitro. The polyubiquitinated S-RNases are degraded in the pollen extract, which is attenuated by a proteasome inhibitor. Our findings suggest that multiple SCF(SLF) complexes in cross-pollen polyubiquitinate non-self S-RNases, resulting in their degradation by the proteasome.

Optimal T cell cross-reactivity and the role of regulatory T cells.

The T lymphocytes of the adaptive immune system constitute a highly diverse repertoire of clones expressing a unique T cell receptor (TCR). It has been argued that TCRs are cross-reactive, meaning that one receptor can recognize a multitude of epitopes. Cross-reactivity between self and foreign epitopes can potentially lead to autoimmune responses. Regulatory T cells (Tregs) down-regulate immune reactions, and play an important role in the avoidance of autoimmunity. We use a probabilistic modeling approach to investigate how suppression of antigen-presenting dendritic cells (DCs) by Tregs influences the probability of mounting a successful immune response against a pathogen while remaining self-tolerant. For T cell cross-reactivity values close to experimental estimates, we find that the presence of Tregs increases this success probability somewhat. However, the probability of a successful immune response remains relatively low for these cross-reactivity values, and the probability of success is optimized when T cells are more specific and no Tregs are formed. We conclude that DC suppression on its own is insufficient to provide an evolutionary benefit of regulatory T cells. Rejecting one intuitively likely hypothesis for the function of Tregs thus narrows down the search for the mechanisms by which they are suppressing inappropriate immune responses.

From structure to function - Ligand recognition by myeloid C-type lectin receptors.

The relevance of protein-glycan interactions in immunity has long been underestimated. Yet, the immune system possesses numerous classes of glycan-binding proteins, so-called lectins. Of specific interest is the group of myeloid C-type lectin receptors (CLRs) as they are mainly expressed by myeloid cells and play an important role in the initiation of an immune response. Myeloid CLRs represent a major group amongst pattern recognition receptors (PRRs), placing them at the center of the rapidly growing field of glycoimmunology. CLRs have evolved to encompass a wide range of structures and functions and to recognize a large number of glycans and many other ligands from different classes of biopolymers. This review aims at providing the reader with an overview of myeloid CLRs and selected ligands, while highlighting recent insights into CLR-ligand interactions. Subsequently, methodological approaches in CLR-ligand research will be presented. Finally, this review will discuss how CLR-ligand interactions culminate in immunological functions, how glycan mimicry favors immune escape by pathogens, and in which way immune responses can be affected by CLR-ligand interactions in the long term.

B-1 B cell progenitors transiently and partially express keratin 5 during differentiation in bone marrow.

BACKGROUND: Keratin 5 (K5) is a cytoskeletal tissue-specific protein expressed in the epithelial cells of skin and esophagus and ectopic K5 expression in lymphocytes has never been reported. OBJECTIVE: Here we demonstrate an ectopic epidermal self-protein expression in B-1 B cell by fate mapping of K5-expressing cells. METHODS: K5-Cre×CAG-CAT-loxP-EGFP double Tg (K5×GFP) mice that express enhanced GFP under the control of the K5 promoter were employed. RESULTS: Unexpectedly, B220(+)GFP(+) cells were found in LN, spleen, peripheral blood and peritoneal cavity. These cells were IgM(+)IgD(low)CD23(-)CD43(+)CD19(+)CD93(-), indicating that they were B-1 B cells. The number of B220(+)GFP(+) cells was significantly larger in spleen than in the other tissues tested. Although GFP(+) B-1 cells did not express K5 in the periphery, Lin(-)CD93(+)B220(low-neg)CD19(+) B-1 B cell progenitors expressed GFP and B220(+)CD93(+) progenitor cells expressed K5 and MHC-class II in BM, indicating that GFP(+) B-1 cells transiently expressed K5 and the progenitor cells were potential APC. GFP(+) B-1 cells in the periphery continued expressing MHC class II and had exogenous antigen-presenting capacity comparable to non-follicular B cells. GFP(+) B-1 cells spontaneously secreted more IgM than GFP(-) B-1 cells in vitro. CONCLUSION: These results indicate that B-1 B cells transiently and partially express K5 in BM and are potent for both natural antibody production and antigen presentation.

The natural defense system and the normative self model.

Infectious agents are not the only agressors, and the immune system is not the sole defender of the organism. In an enlarged perspective, the 'normative self model' postulates that a 'natural defense system' protects man and other complex organisms against the environmental and internal hazards of life, including infections and cancers. It involves multiple error detection and correction mechanisms that confer robustness to the body at all levels of its organization. According to the model, the self relies on a set of physiological norms, and NONself (meaning : Non Obedient to the Norms of the self) is anything 'off-norms'. The natural defense system comprises a set of 'civil defenses' (to which all cells in organs and tissues contribute), and a 'professional army ', made of a smaller set of mobile cells. Mobile and non mobile cells differ in their tuning abilities. Tuning extends the recognition capabilities of NONself by the mobile cells, which increase their defensive function. To prevent them to drift, which would compromise self/NONself discrimination, the more plastic mobile cells need to periodically refer to the more stable non mobile cells to keep within physiological standards.

In utero transplantation: Disparate ramifications.

In utero stem cell transplantation, which promises treatment for a host of genetic disorders early in gestation before disease effect stems from Ray Owen's seminal observation that self-tolerance, is acquired during gestation. To date, in utero transplantation (IUT) has proved useful in characterizing the hematopoietic stem cell. Recent observations support its use as an in vivo method to further understanding of self-tolerance. Preclinical development continues for its application as a treatment for childhood hematolymphoid diseases. In addition, IUT may offer therapeutic options in the treatment of diabetes among other diseases. Thus IUT serves as a technique or system important in both a basic and applied format. This review summarizes these findings.

Physiologically-Mediated Self/Non-Self Root Discrimination in Trifolium repens has Mixed Effects on Plant Performance.

Recent studies suggest that plant roots can avoid competition with other roots of the same plant, but the mechanism behind this behavior is yet largely unclear and their effects on plant performance hardly studied. We grew combinations of two ramets of Trifolium repens in a single pot that were either intact, disconnected for a shorter or longer time, or that belonged to different genotypes. Interconnected ramets developed lower root length and mass than any other combination of ramets, supporting the notion that self/non-self discrimination in T. repens was based entirely on physiological coordination between different roots that develop on the same plant, rather than biochemical allorecognition. These responses were consistent among eight field-collected genotypes, suggesting that self/non-self discrimination is a common feature in wild populations of white clover. There were no significant treatment x genotype interactions suggesting that genetic variation for self/non-self discrimination may be limited. Self-interactions resulted in lower to similar shoot biomass and number of ramets, but higher flowering probabilities, compared to non-self interactions. Thus, our results demonstrated that the performance consequences of self/non-self discrimination may be more complicated than previously thought.