CRISPR/Cas systems in archaea: What array spacers can teach us about parasitism and gene exchange in the 3rd domain of life

CRISPR (Clustered, Regularly, Interspaced, Short, Palindromic Repeats) loci have been shown to provide prokaryotes with an adaptive immunity against viruses and plasmids. CRISPR arrays are transcribed and processed into small CRISPR RNA molecules, which base-pair with invading DNA or RNA and lead to its degradation by CRISPR-associated (Cas) protein complexes. New spacers can be acquired by active CRISPR/Cas systems, and thus the sequences of these spacers provide a record of the past “infection history” of the organism. Recently we used spacer sequences from archaeal genomes to infer gene exchange events among archaeal species and genera and to demonstrate that at least in this domain of life CRISPR indeed has an anti-viral role

Complexity, connectivity, and duplicability as barriers to lateral gene transfer

Laterally transferred genes are shown to be less involved in protein-protein interactions, and essential genes that exhibit low duplicability and high connectivity do exhibit mostly vertical descent

Deriving enzymatic and taxonomic signatures of metagenomes from short read data

<p>Abstract</p> <p>Background</p> <p>We propose a method for deriving enzymatic signatures from short read metagenomic data of unknown species. The short read data are converted to six pseudo-peptide candidates. We search for occurrences of Specific Peptides (SPs) on the latter. SPs are peptides that are indicative of enzymatic function as defined by the Enzyme Commission (EC) nomenclature. The number of SP hits on an ensemble of short reads is counted and then converted to estimates of numbers of enzymatic genes associated with different EC categories in the studied metagenome. Relative amounts of different EC categories define the enzymatic spectrum, without the need to perform genomic assemblies of short reads.</p> <p>Results</p> <p>The method is developed and tested on 22 bacteria for which there exist many EC annotations in Uniprot. Enzymatic signatures are derived for 3 metagenomes, and their functional profiles are explored.</p> <p>We extend the SP methodology to taxon-specific SPs (TSPs), allowing us to estimate taxonomic features of metagenomic data from short reads. Using recent Swiss-Prot data we obtain TSPs for different phyla of bacteria, and different classes of proteobacteria. These allow us to analyze the major taxonomic content of 4 different metagenomic data-sets.</p> <p>Conclusions</p> <p>The SP methodology can be successfully extended to applications on short read genomic and metagenomic data. This leads to direct derivation of enzymatic signatures from raw short reads. Furthermore, by employing TSPs, one obtains valuable taxonomic information.</p

Specific changes in the mammalian gut microbiome as a biomarker for oxytocin-induced behavioral changes

Humans are colonized by bacteria, fungi, archaea, and viruses, which are collectively referred to as the microbiome. Most of the microbiome resides in the gut and may easily be investigated via stool sampling and subsequent metagenomic DNA sequencing. Prolonged exposure to psychiatric pharmacological agents is often associated with marked gastrointestinal phenomena, including changes in food intake, bowel motility, gastric emptying, and transit time [1–3]. Unlike the relatively objective measurement of the microbiota composition, accurate assessment of patients’ therapy adherence and treatment outcomes represent a challenge in psychiatric medical care [4]. This is partly because, for most psychopharmacological agents, compliance and response to treatments are subjectively assessed based on self-reporting and physicians’ evaluations [5,6]. An interesting alternative is having changes in the psychiatric patients’ gut microbiota composition serve as a measurable proxy for monitoring patients’ compliance and the therapeutic effects of some drugs. It is yet unclear how behavioral changes and drug intake affect the microbiota; however, mounting evidence suggests that physical and mental disturbances may lead to changes in gastrointestinal (GI) motility [7,8] in both animals and humans [9–11]. Indeed, in humans, anger, fear, pain, and anxiety, as well as intensive exercise, results in changes in GI activity [8]. In rats, chronic stress results in initial delayed gastric emptying followed by acceleration later on [12]. Medication intake [13,14] and changes in stool consistency, gastric transit, and emptying time [15,16] also have a great impact on microbial composition

CEACAM1 recognition by bacterial pathogens is species-specific

<p>Abstract</p> <p>Background</p> <p>Carcinoembryonic antigen-related cell adhesion molecule 1 (CEACAM1), an immunoglobulin (Ig)-related glycoprotein, serves as cellular receptor for a variety of Gram-negative bacterial pathogens associated with the human mucosa. In particular, <it>Neisseria gonorrhoeae</it>, <it>N. meningitidis</it>, <it>Moraxella catarrhalis</it>, and <it>Haemophilus influenzae </it>possess well-characterized CEACAM1-binding adhesins. CEACAM1 is typically involved in cell-cell attachment, epithelial differentiation, neovascularisation and regulation of T-cell proliferation, and is one of the few CEACAM family members with homologues in different mammalian lineages. However, it is unknown whether bacterial adhesins of human pathogens can recognize CEACAM1 orthologues from other mammals.</p> <p>Results</p> <p>Sequence comparisons of the amino-terminal Ig-variable-like domain of CEACAM1 reveal that the highest sequence divergence between human, murine, canine and bovine orthologues is found in the β-strands comprising the bacteria-binding CC'FG-face of the Ig-fold. Using GFP-tagged, soluble amino-terminal domains of CEACAM1, we demonstrate that bacterial pathogens selectively associate with human, but not other mammalian CEACAM1 orthologues. Whereas full-length human CEACAM1 can mediate internalization of <it>Neisseria gonorrhoeae </it>in transfected cells, murine CEACAM1 fails to support bacterial internalization, demonstrating that the sequence divergence of CEACAM1 orthologues has functional consequences with regard to bacterial recognition and cellular invasion.</p> <p>Conclusions</p> <p>Our results establish the selective interaction of several human-restricted bacterial pathogens with human CEACAM1 and suggest that co-evolution of microbial adhesins with their corresponding receptors on mammalian cells contributes to the limited host range of these highly adapted infectious agents.</p

Metabolic-network-driven analysis of bacterial ecological strategies

Bacterial ecological strategies revealed by metabolic network analysis show that ecological diversity correlates with metabolic flexibility, faster growth rate and intense co-habitation

High tolerance to self-targeting of the genome by the endogenous CRISPR-Cas system in an archaeon

CRISPR-Cas systems allow bacteria and archaea to acquire sequence-specific immunity against selfish genetic elements such as viruses and plasmids, by specific degradation of invader DNA or RNA. However, this involves the risk of autoimmunity if immune memory against host DNA is mistakenly acquired. Such autoimmunity has been shown to be highly toxic in several bacteria and is believed to be one of the major costs of maintaining these defense systems. Here we generated an experimental system in which a non-essential gene, required for pigment production and the reddish colony color, is targeted by the CRISPR-Cas I-B system of the halophilic archaeon Haloferax volcanii. We show that under native conditions, where both the self-targeting and native crRNAs are expressed, self-targeting by CRISPR-Cas causes no reduction in transformation efficiency of the plasmid encoding the self-targeting crRNA. Furthermore, under such conditions, no effect on organismal growth rate or loss of the reddish colony phenotype due to mutations in the targeted region could be observed. In contrast, in cells deleted for the pre-crRNA processing gene cas6, where only the self-targeting crRNA exists as mature crRNA, self-targeting leads to moderate toxicity and the emergence of deletion mutants. Sequencing of the deletions caused by CRISPR-Cas self targeting indicated DNA repair via microhomology-mediated end joining

Native homing endonucleases can target conserved genes in humans and in animal models

In recent years, both homing endonucleases (HEases) and zinc-finger nucleases (ZFNs) have been engineered and selected for the targeting of desired human loci for gene therapy. However, enzyme engineering is lengthy and expensive and the off-target effect of the manufactured endonucleases is difficult to predict. Moreover, enzymes selected to cleave a human DNA locus may not cleave the homologous locus in the genome of animal models because of sequence divergence, thus hampering attempts to assess the in vivo efficacy and safety of any engineered enzyme prior to its application in human trials. Here, we show that naturally occurring HEases can be found, that cleave desirable human targets. Some of these enzymes are also shown to cleave the homologous sequence in the genome of animal models. In addition, the distribution of off-target effects may be more predictable for native HEases. Based on our experimental observations, we present the HomeBase algorithm, database and web server that allow a high-throughput computational search and assignment of HEases for the targeting of specific loci in the human and other genomes. We validate experimentally the predicted target specificity of candidate fungal, bacterial and archaeal HEases using cell free, yeast and archaeal assay

The Effect of Density-Dependent Phase on the Locust Gut Bacterial Composition

The desert locust demonstrates density-dependent phase polyphenism: For extended periods it appears in a non-aggregating, non-migrating phenotype, known as the solitary phase. When circumstances change, solitary individuals may aggregate and transform to the gregarious phenotype, which have a strong propensity for generating large swarms. Previous reports have suggested a role for gut-bacteria derived volatiles in the swarming phenomenon, and suggested that locusts are capable of manipulating their gut microbiome according to their density-dependent phases. Here, we directly tested this hypothesis for the first time. Using locusts of both phases from well-controlled laboratory cultures as well as gregarious field-collected individuals; and high-throughput sequencing. We characterized the hindgut bacterial community composition in the two phases of the desert locust. Our findings demonstrate that laboratory-reared gregarious and solitary locusts maintain a stable core of Enterobacter. However, while different generations of gregarious locust experience shifts in their Enterobacter’s relative abundance; the solitary locusts maintain a stable gut microbiome, highly similar to that of the field-collected locusts. Tentative phase differences in wild populations’ microbiome may thus be an indirect effect of environmental or other factors that push the swarming individuals to homogenous gut bacteria. We therefore conclude that there are phase-related differences in the population dynamics of the locust hindgut bacterial composition, but there is no intrinsic density-dependent mechanism directly affecting the gut microbiome