Diverse Immune Systems Research Articles

Novel adaptive immune systems in pristine Antarctic soils

Antarctic environments are dominated by microorganisms, which are vulnerable to viral infection. Although several studies have investigated the phylogenetic repertoire of bacteria and viruses in these poly-extreme environments with freezing temperatures, high ultra violet irradiation levels, low moisture availability and hyper-oligotrophy, the evolutionary mechanisms governing microbial immunity remain poorly understood. Using genome-resolved metagenomics, we test the hypothesis that Antarctic poly-extreme high-latitude microbiomes harbour diverse adaptive immune systems. Our analysis reveals the prevalence of prophages in bacterial genomes (Bacteroidota and Verrucomicrobiota), suggesting the significance of lysogenic infection strategies in Antarctic soils. Furthermore, we demonstrate the presence of diverse CRISPR-Cas arrays, including Class 1 arrays (Types I-B, I-C, and I-E), alongside systems exhibiting novel gene architecture among their effector cas genes. Notably, a Class 2 system featuring type V variants lacks CRISPR arrays, encodes Cas1 and Cas2 adaptation module genes. Phylogenetic analysis of Cas12 effector proteins hints at divergent evolutionary histories compared to classified type V effectors and indicates that TnpB is likely the ancestor of Cas12 nucleases. Our findings suggest substantial novelty in Antarctic cas sequences, likely driven by strong selective pressures. These results underscore the role of viral infection as a key evolutionary driver shaping polar microbiomes.

Distribution of specific prokaryotic immune systems correlates with host optimal growth temperature.

Prokaryotes encode an arsenal of highly diverse immune systems to protect themselves against invading nucleic acids such as viruses, plasmids and transposons. This includes invader-interfering systems that neutralize invaders to protect their host, and abortive-infection systems, which trigger dormancy or cell death in their host to offer population-level immunity. Most prokaryotic immune systems are found across different environments and prokaryotic phyla, but their distribution appears biased and the factors that influence their distribution are largely unknown. Here, we compared and combined the prokaryotic immune system identification tools DefenseFinder and PADLOC to obtain an expanded view of the immune system arsenal. Our results show that the number of immune systems encoded is positively correlated with genome size and that the distribution of specific immune systems is linked to phylogeny. Furthermore, we reveal that certain invader-interfering systems are more frequently encoded by hosts with a relatively high optimum growth temperature, while abortive-infection systems are generally more frequently encoded by hosts with a relatively low optimum growth temperature. Combined, our study reveals several factors that correlate with differences in the distribution of prokaryotic immune systems and extends our understanding of how prokaryotes protect themselves from invaders in different environments.

Bacteriophages avoid autoimmunity from cognate immune systems as an intrinsic part of their life cycles.

Dormant prophages protect lysogenic cells by expressing diverse immune systems, which must avoid targeting their cognate prophages upon activation. Here we report that multiple Staphylococcus aureus prophages encode Tha (tail-activated, HEPN (higher eukaryotes and prokaryotes nucleotide-binding) domain-containing anti-phage system), a defence system activated by structural tail proteins of incoming phages. We demonstrate the function of two Tha systems, Tha-1 and Tha-2, activated by distinct tail proteins. Interestingly, Tha systems can also block reproduction of the induced tha-positive prophages. To prevent autoimmunity after prophage induction, these systems are inhibited by the product of a small overlapping antisense gene previously believed to encode an excisionase. This genetic organization, conserved in S. aureus prophages, allows Tha systems to protect prophages and their bacterial hosts against phage predation and to be turned off during prophage induction, balancing immunity and autoimmunity. Our results show that the fine regulation of these processes is essential for the correct development of prophages' life cycle.

Functionally comparable but evolutionarily distinct nucleotide-targetingeffectors help identify conserved paradigms across diverse immune systems.

While nucleic acid-targeting effectors are known to be central to biological conflicts and anti-selfish element immunity, recent findings have revealed immune effectors that target their building blocks and the cellular energy currency-free nucleotides. Through comparative genomics and sequence-structure analysis, we identified several distinct effector domains, which we named Calcineurin-CE, HD-CE, and PRTase-CE. These domains, along with specific versions of the ParB and MazG domains, are widely present in diverse prokaryotic immune systems and are predicted to degrade nucleotides by targeting phosphate or glycosidic linkages. Our findings unveil multiple potential immune systems associated with at least 17 different functional themes featuring these effectors. Some of these systems sense modified DNA/nucleotides from phages or operate downstream of novel enzymes generating signaling nucleotides. We also uncovered a class of systems utilizing HSP90- and HSP70-related modules as analogs of STAND and GTPase domains that are coupled to these nucleotide-targeting- or proteolysis-induced complex-forming effectors. While widespread in bacteria, only a limited subset of nucleotide-targeting effectors was integrated into eukaryotic immune systems, suggesting barriers to interoperability across subcellular contexts. This work establishes nucleotide-degrading effectors as an emerging immune paradigm and traces their origins back to homologous domains in housekeeping systems.

Restriction endonuclease cleavage of phage DNA enables resuscitation from Cas13-induced bacterial dormancy.

Type VI CRISPR systems protect against phage infection using the RNA-guided nuclease Cas13 to recognize viral messenger RNA. Upon target recognition, Cas13 cleaves phage and host transcripts non-specifically, leading to cell dormancy that is incompatible with phage propagation. However, whether and how infected cells recover from dormancy is unclear. Here we show that type VI CRISPR and DNA-cleaving restriction-modification (RM) systems frequently co-occur and synergize to clear phage infections and resuscitate cells. In the natural type VI CRISPR host Listeria seeligeri, we show that RM cleaves the phage genome, thus removing the source of phage transcripts and enabling cells to recover from Cas13-induced cellular dormancy. We find that phage infections are neutralized more effectively when Cas13 and RM systems operate together. Our work reveals that type VI CRISPR immunity is cell-autonomous and non-abortive when paired with RM, and hints at other synergistic roles for the diverse host-directed immune systems in bacteria.

Control of bacterial immune signaling by a WYL domain transcription factor.

Bacteria use diverse immune systems to defend themselves from ubiquitous viruses termed bacteriophages (phages). While the molecular mechanisms of many such systems have been detailed in recent years, how these systems are controlled and the signals that activate them remain mostly unknown. Here, we identify a transcription factor associated with a fraction of CBASS immune systems that we term CapW. CapW is similar to the recently-discovered BrxR transcription factor found in some BREX immune systems, and homologs of CapW/BrxR are associated with a variety of bacterial immune systems. Combining X-ray crystallography and biochemical analysis, we show that CapW forms a homodimer and binds a palindromic DNA sequence in the CBASS promoter region. Two crystal structures of CapW suggest that the protein switches from an unliganded, DNA binding-competent state to a ligand-bound state unable to bind DNA. Using a reporter assay, we show that CapW strongly represses CBASS gene expression in uninfected cells and that CBASS expression is increased either upon phage infection or the addition of DNA damaging drugs. These results suggest that CapW enables its associated CBASS system to respond not only to phage infection but also to DNA damage, a universal signal of cell stress. We propose two hypotheses for the role of CapW and the functionally-related CapH+CapP system: first, these regulators may enable multiple bacterial immune systems to coordinate their functions in an infected cell, to mount a multi-pronged defensive response. Alternatively, CapW and CapH+CapP may enable bacteria to combat the conversion of phages from lysogenic to lytic lifestyles, by responding to the same stress signal(s) that induce prophage mobilization.

A conserved signaling pathway activates bacterial CBASS immune signaling in response to DNA damage.

To protect themselves from the constant threat of bacteriophage (phage) infection, bacteria have evolved diverse immune systems including restriction-modification, CRISPR-Cas, and many others. Here, we describe the discovery of a two-protein transcriptional regulator module associated with hundreds of CBASS immune systems and demonstrate that this module drives the expression of its associated CBASS system in response to DNA damage. We show that the helix-turn-helix transcriptional repressor CapH binds the promoter region of its associated CBASS system to repress transcription until it is cleaved by the metallopeptidase CapP. CapP is activated in vitro by single-stranded DNA, and in cells by DNA-damaging drugs. Together, CapH and CapP drive increased expression of their associated CBASS system in response to DNA damage. We identify CapH- and CapP-related proteins associated with diverse known and putative bacterial immune systems including DISARM and Pycsar antiphage operons. Overall, our data highlight a mechanism by which bacterial immune systems can sense and respond to a universal signal of cell stress, potentially enabling multiple immune systems to mount a coordinated defensive response against an invading pathogen.

Shared TIR enzymatic functions regulate cell death and immunity across the tree of life.

In the 20th century, researchers studying animal and plant signaling pathways discovered a protein domain that is shared across diverse innate immune systems: the Toll/interleukin-1/resistance gene (TIR) domain. The TIR domain is found in several protein architectures and was defined as an adaptor that mediates protein-protein interactions in animal innate immunity and developmental signaling pathways. However, studies of nerve degeneration in animals-and subsequent breakthroughs in plant, bacterial, and archaeal systems-revealed that TIR domains possess enzymatic activities. We provide a synthesis of TIR functions and the role of various related TIR enzymatic products in evolutionarily diverse immune systems. These studies may ultimately guide interventions that would span the tree of life, from treating human neurodegenerative disorders and bacterial infections to preventing plant diseases.

Control of bacterial immune signaling by a WYL domain transcription factor.

Bacteria use diverse immune systems to defend themselves from ubiquitous viruses termed bacteriophages (phages). Many anti-phage systems function by abortive infection to kill a phage-infected cell, raising the question of how they are regulated to avoid cell killing outside the context of infection. Here, we identify a transcription factor associated with the widespread CBASS bacterial immune system, that we term CapW. CapW forms a homodimer and binds a palindromic DNA sequence in the CBASS promoter region. Two crystal structures of CapW suggest that the protein switches from an unliganded, DNA binding-competent state to a ligand-bound state unable to bind DNA. We show that CapW strongly represses CBASS gene expression in uninfected cells, and that phage infection causes increased CBASS expression in a CapW-dependent manner. Unexpectedly, this CapW-dependent increase in CBASS expression is not required for robust anti-phage activity, suggesting that CapW may mediate CBASS activation and cell death in response to a signal other than phage infection. Our results parallel concurrent reports on the structure and activity of BrxR, a transcription factor associated with the BREX anti-phage system, suggesting that CapW and BrxR are members of a family of universal defense signaling proteins.

A unique mode of nucleic acid immunity performed by a multifunctional bacterial enzyme

The perpetual arms race between bacteria and their viruses (phages) has given rise to diverse immune systems, including restriction-modification and CRISPR-Cas, which sense and degrade phage-derived nucleic acids. These complex systems rely upon production and maintenance of multiple components to achieve antiphage defense. However, the prevalence and effectiveness of minimal, single-component systems that cleave DNA remain unknown. Here, we describe a unique mode of nucleic acid immunity mediated by a single enzyme with nuclease and helicase activities, herein referred to as Nhi (nuclease-helicase immunity). This enzyme provides robust protection against diverse staphylococcal phages and prevents phage DNA accumulation in cells stripped of all other known defenses. Our observations support a model in which Nhi targets and degrades phage-specific replication intermediates. Importantly, Nhi homologs are distributed in diverse bacteria and exhibit functional conservation, highlighting the versatility of such compact weapons as major players in antiphage defense.

SARS-CoV-2 Virus is Evolving to Adapt : A Fast Check Note

Coronavirus Disease 19 (COVID-19) caused by Severe Acute Respiratory Syndrome Coronavirus-2 (SARS-CoV-2), a highly transmissible and pathogenic coronavirus, has spread at an alarming rate throughout the world since Dec. 2019, claiming 2.196 million deaths globally, as per 30th Jan. 2021, and the count is on. The reason behind this ongoing rapid transmission and global spread of SARS-CoV-2 is that the virus is evolving to become more transmissible as it spreads across the world due to its fast host migration. The virus is rapidly evolving to adapt to the different geo‑climate environments, diverse host immune systems, and other protective counter-measures (such as prolong host survival) by accumulating adaptive mutations, deletions, and recombination. This note focuses on those mutations and the prominent viral strains that are noteworthy for epidemiological and biological reasons.

Identification of a Type IV-A CRISPR-Cas System Located Exclusively on IncHI1B/IncFIB Plasmids in Enterobacteriaceae.

Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR) are diverse immune systems found in many prokaryotic genomes that target invading foreign DNA such as bacteriophages and plasmids. There are multiple types of CRISPR with arguably the most enigmatic being Type IV. During an investigation of CRISPR carriage in clinical, multi-drug resistant, Klebsiella pneumoniae, a Type IV-A3 CRISPR-Cas system was detected on plasmids from two K. pneumoniae isolates from Egypt (isolated in 2002–2003) and a single K. pneumoniae isolate from the United Kingdom (isolated in 2017). Sequence analysis of all other genomes available in GenBank revealed that this CRISPR-Cas system was present on 28 other plasmids from various Enterobacteriaceae hosts and was never found on a bacterial chromosome. This system is exclusively located on IncHI1B/IncFIB plasmids and is associated with multiple putative transposable elements. Expression of the cas loci was confirmed in the available clinical isolates by RT-PCR. In all cases, the CRISPR-Cas system has a single CRISPR array (CRISPR1) upstream of the cas loci which has several, conserved, spacers which, amongst things, match regions within conjugal transfer genes of IncFIIK/IncFIB(K) plasmids. Our results reveal a Type IV-A3 CRISPR-Cas system exclusively located on IncHI1B/IncFIB plasmids in Enterobacteriaceae that is likely to be able to target IncFIIK/IncFIB(K) plasmids presumably facilitating intracellular, inter-plasmid competition.

The regulatory genome constrains protein sequence evolution: implications for the search for disease-associated genes.

The development of explanatory models of protein sequence evolution has broad implications for our understanding of cellular biology, population history, and disease etiology. Here we analyze the GTEx transcriptome resource to quantify the effect of the transcriptome on protein sequence evolution in a multi-tissue framework. We find substantial variation among the central nervous system tissues in the effect of expression variance on evolutionary rate, with highly variable genes in the cortex showing significantly greater purifying selection than highly variable genes in subcortical regions (Mann–Whitney U p = 1.4 × 10−4). The remaining tissues cluster in observed expression correlation with evolutionary rate, enabling evolutionary analysis of genes in diverse physiological systems, including digestive, reproductive, and immune systems. Importantly, the tissue in which a gene attains its maximum expression variance significantly varies (p = 5.55 × 10−284) with evolutionary rate, suggesting a tissue-anchored model of protein sequence evolution. Using a large-scale reference resource, we show that the tissue-anchored model provides a transcriptome-based approach to predicting the primary affected tissue of developmental disorders. Using gradient boosted regression trees to model evolutionary rate under a range of model parameters, selected features explain up to 62% of the variation in evolutionary rate and provide additional support for the tissue model. Finally, we investigate several methodological implications, including the importance of evolutionary-rate-aware gene expression imputation models using genetic data for improved search for disease-associated genes in transcriptome-wide association studies. Collectively, this study presents a comprehensive transcriptome-based analysis of a range of factors that may constrain molecular evolution and proposes a novel framework for the study of gene function and disease mechanism.

An adverse outcome pathway for immune-mediated and allergic hepatitis: a case study with the NSAID diclofenac

Many drugs have the potential to cause drug-induced liver injury (DILI); however, underlying mechanisms are diverse. The concept of adverse outcome pathways (AOPs) has become instrumental for risk assessment of drug class effects. We report AOPs specific for immune-mediated and drug hypersensitivity/allergic hepatitis by considering genomic, histo- and clinical pathology data of mice and dogs treated with diclofenac. The findings are relevant for other NSAIDs and drugs undergoing iminoquinone and quinone reactive metabolite formation. We define reactive metabolites catalyzed by CYP monooxygenase and myeloperoxidases of neutrophils and Kupffer cells as well as acyl glucuronides produced by uridine diphosphoglucuronosyl transferase as molecular initiating events (MIE). The reactive metabolites bind to proteins and act as neo-antigen and involve antigen-presenting cells to elicit B- and T-cell responses. Given the diverse immune systems between mice and dogs, six different key events (KEs) at the cellular and up to four KEs at the organ level are defined with mechanistic plausibility for the onset and progression of liver inflammation. With mice, cellular stress response, interferon gamma-, adipocytokine- and chemokine signaling provided a rationale for the AOP of immune-mediated hepatitis. With dogs, an erroneous programming of the innate and adaptive immune response resulted in mast cell activation; their infiltration into liver parenchyma and the shift to M2-polarized Kupffer cells signify allergic hepatitis and the occurrence of granulomas of the liver. Taken together, diclofenac induces divergent immune responses among two important preclinical animal species, and the injury pattern seen among clinical cases confirms the relevance of the developed AOP for immune-mediated hepatitis.

Characterization of Cas12a nucleases reveals diverse PAM profiles between closely-related orthologs.

CRISPR-Cas systems comprise diverse adaptive immune systems in prokaryotes whose RNA-directed nucleases have been co-opted for various technologies. Recent efforts have focused on expanding the number of known CRISPR-Cas subtypes to identify nucleases with novel properties. However, the functional diversity of nucleases within each subtype remains poorly explored. Here, we used cell-free transcription-translation systems and human cells to characterize six Cas12a single-effector nucleases from the V-A subtype, including nucleases sharing high sequence identity. While these nucleases readily utilized each other's guide RNAs, they exhibited distinct PAM profiles and apparent targeting activities that did not track based on phylogeny. In particular, two Cas12a nucleases encoded by Prevotella ihumii (PiCas12a) and Prevotella disiens (PdCas12a) shared over 95% amino-acid identity yet recognized distinct PAM profiles, with PiCas12a but not PdCas12a accommodating multiple G’s in PAM positions -2 through -4 and T in position -1. Mutational analyses transitioning PiCas12a to PdCas12a resulted in PAM profiles distinct from either nuclease, allowing more flexible editing in human cells. Cas12a nucleases therefore can exhibit widely varying properties between otherwise related orthologs, suggesting selective pressure to diversify PAM recognition and supporting expansion of the CRISPR toolbox through ortholog mining and PAM engineering.

Evolutionary perspective on the hematopoietic system through a colonial chordate: allogeneic immunity and hematopoiesis.

Evolution and selection have shaped diverse immune systems throughout phylogeny, the vast majority of which remain unexplored. Botryllus schlosseri is a colonial tunicate, a sister group to vertebrates, that develops as a chordate, then metamorphoses to an asexually reproductive invertebrate that every week makes the same body plan from budded stem cells. Genetically distinct B. schlosseri colonies can fuse to form a chimera, or reject each other based on allogeneic recognition. In chimeras, circulating germline and somatic stem cells participate in development; stem cells compete in all individuals in the fused colonies, with rejection preventing germline parasitism. Here we review the isolation and characterization of B. schlosseri hematopoietic stem cells (HSC) and their niches, and the role of the immune effector cells in allorecognition.

Antimicrobial peptide similarity and classification through rough set theory using physicochemical boundaries

BackgroundAntimicrobial peptides attract considerable interest as novel agents to combat infections. Their long-time potency across bacteria, viruses and fungi as part of diverse innate immune systems offers a solution to overcome the rising concerns from antibiotic resistance. With the rapid increase of antimicrobial peptides reported in the databases, peptide selection becomes a challenge. We propose similarity analyses to describe key properties that distinguish between active and non-active peptide sequences building upon the physicochemical properties of antimicrobial peptides. We used an iterative supervised machine learning approach to classify active peptides from inactive peptides with low false discovery rates in a relatively short computational search time.ResultsBy generating explicit boundaries, our method defines new categories of active and inactive peptides based on their physicochemical properties. Consequently, it describes physicochemical characteristics of similarity among active peptides and the physicochemical boundaries between active and inactive peptides in a single process. To build the similarity boundaries, we used the rough set theory approach; to our knowledge, this is the first time that this approach has been used to classify peptides. The modified rough set theory method limits the number of values describing a boundary to a user-defined limit. Our method is optimized for specificity over selectivity. Noting that false positives increase activity assays while false negatives only increase computational search time, our method provided a low false discovery rate. Published datasets were used to compare our rough set theory method to other published classification methods and based on this comparison, we achieved high selectivity and comparable sensitivity to currently available methods.ConclusionsWe developed rule sets that define physicochemical boundaries which allow us to directly classify the active sequences from inactive peptides. Existing classification methods are either sequence-order insensitive or length-dependent, whereas our method generates the rule sets that combine order-sensitive descriptors with length-independent descriptors. The method provides comparable or improved performance to currently available methods. Discovering the boundaries of physicochemical properties may lead to a new understanding of peptide similarity.

CRISPR RNA-guided DNA cleavage by reconstituted Type I-A immune effector complexes.

Diverse CRISPR-Cas immune systems protect archaea and bacteria from viruses and other mobile genetic elements. All CRISPR-Cas systems ultimately function by sequence-specific destruction of invading complementary nucleic acids. However, each CRISPR system uses compositionally distinct crRNP [CRISPR (cr) RNA/Cas protein] immune effector complexes to recognize and destroy invasive nucleic acids by unique molecular mechanisms. Previously, we found that Type I-A (Csa) effector crRNPs from Pyrococcus furiosus function in vivo to eliminate invader DNA. Here, we reconstituted functional Type I-A effector crRNPs in vitro with recombinant Csa proteins and synthetic crRNA and characterized properties of crRNP assembly, target DNA recognition and cleavage. Six proteins (Csa 4-1, Cas3″, Cas3', Cas5a, Csa2, Csa5) are essential for selective target DNA binding and cleavage. Native gel shift analysis and UV-induced RNA-protein crosslinking demonstrate that Cas5a and Csa2 directly interact with crRNA 5' tag and guide sequences, respectively. Mutational analysis revealed that Cas3″ is the effector nuclease of the complex. Together, our results indicate that DNA cleavage by Type I-A crRNPs requires crRNA-guided and protospacer adjacent motif-dependent target DNA binding to unwind double-stranded DNA and expose single strands for progressive ATP-dependent 3'-5' cleavage catalyzed by integral Cas3' helicase and Cas3″ nuclease crRNP components.

The Value of a Comparative Approach to Understand the Complex Interplay between Microbiota and Host Immunity.

The eukaryote immune system evolved and continues to evolve within a microbial world, and as such is critically shaped by—and in some cases even reliant upon—the presence of host-associated microbial species. There are clear examples of adaptations that allow the host to simultaneously tolerate and/or promote growth of symbiotic microbiota while protecting itself against pathogens, but the relationship between immunity and the microbiome reaches far beyond simple recognition and includes complex cross talk between host and microbe as well as direct microbiome-mediated protection against pathogens. Here, we present a broad but brief overview of how the microbiome is controlled by and interacts with diverse immune systems, with the goal of identifying questions that can be better addressed by taking a comparative approach across plants and animals and different types of immunity. As two key examples of such an approach, we focus on data examining the importance of early exposure on microbiome tolerance and immune system development and function, and the importance of transmission among hosts in shaping the potential coevolution between, and long-term stability of, host–microbiome associations. Then, by comparing existing evidence across short-lived plants, mouse model systems and humans, and insects, we highlight areas of microbiome research that are strong in some systems and absent in others with the hope of guiding future research that will allow for broad-scale comparisons moving forward. We argue that such an approach will not only help with identification of generalities in host–microbiome–immune interactions but also improve our understanding of the role of the microbiome in host health.

Pathogen-secreted proteases activate a novel plant immune pathway.

Mitogen-Activated Protein Kinase (MAPK) cascades play central roles in innate immune signaling networks in plants and animals1,2. In plants, however, the molecular mechanisms of how signal perception is transduced to MAPK activation remain elusive1. We report that pathogen-secreted proteases activate a previously unknown signaling pathway in Arabidopsis thaliana involving the Gα, Gβ and Gγ subunits of heterotrimeric G-protein complexes, which function upstream of a MAPK cascade. In this pathway, Receptor for Activated C Kinase 1 (RACK1) functions as a novel scaffold that binds to the Gβ subunit as well as to all three tiers of the MAPK cascade, thereby linking upstream G protein signaling to downstream activation of a MAPK cascade. The protease-G protein-RACK1-MAPK cascade modules identified in these studies are distinct from previously described plant immune signaling pathways such as the one elicited by bacterial flagellin, in which G proteins function downstream of or in parallel to a MAPK cascade without the involvement of the RACK1 scaffolding protein. The discovery of the novel protease-mediated immune signaling pathway described here was facilitated by the use of the broad host range, opportunistic bacterial pathogen Pseudomonas aeruginosa. The ability of P. aeruginosa to infect both plants and animals makes it an excellent model to identify novel types of immunoregulatory strategies that account for its niche adaptation to diverse host tissues and immune systems.