Kinetics of the CRISPR-Cas9 effector complex assembly and the role of 3'-terminal segment of guide RNA.

CRISPR-Cas9 is widely applied for genome engineering in various organisms. The assembly of single guide RNA (sgRNA) with the Cas9 protein may limit the Cas9/sgRNA effector complex function. We developed a FRET-based assay for detection of CRISPR-Cas9 complex binding to its targets and used this assay to investigate the kinetics of Cas9 assembly with a set of structurally distinct sgRNAs.

Function of the CRISPR-Cas System of the Human Pathogen Clostridium difficile.

Unlabelled: Clostridium difficile is the cause of most frequently occurring nosocomial diarrhea worldwide. As an enteropathogen, C. difficile must be exposed to multiple exogenous genetic elements in bacteriophage-rich gut communities.

Development of giant bacteriophage ϕKZ is independent of the host transcription apparatus.

Unlabelled: Pseudomonas aeruginosa bacteriophage ϕKZ is the type representative of the giant phage genus, which is characterized by unusually large virions and genomes. By unraveling the transcriptional map of the ∼ 280-kb ϕKZ genome to single-nucleotide resolution, we combine 369 ϕKZ genes into 134 operons. Early transcription is initiated from highly conserved AT-rich promoters distributed across the ϕKZ genome and located on the same strand of the genome.

Activity and bacterial diversity of snow around Russian Antarctic stations.

The diversity and temporal dynamics of bacterial communities in pristine snow around two Russian Antarctic stations was investigated. Taxonomic analysis of rDNA libraries revealed that snow communities were dominated by bacteria from a small number of operational taxonomic units (OTUs) that underwent dramatic swings in abundance between the 54th (2008-2009) and 55th (2009-2010) Russian Antarctic expeditions. Moreover, analysis of the 55th expedition samples indicated that there was very little, if any, correspondence in abundance of clones belonging to the same OTU present in rDNA and rRNA libraries.

Cooperativity and interaction energy threshold effects in recognition of the -10 promoter element by bacterial RNA polymerase.

RNA polymerase (RNAP) melts promoter DNA to form transcription-competent open promoter complex (RPo). Interaction of the RNAP σ subunit with non-template strand bases of a conserved -10 element (consensus sequence T-12A-11T-10A-9A-8T-7) is an important source of energy-driving localized promoter melting. Here, we used an RNAP molecular beacon assay to investigate interdependencies of RNAP interactions with -10 element nucleotides.

Host RNA polymerase inhibitors encoded by ϕKMV-like phages of Pseudomonas.

Escherichia coli bacteriophage T7 is a founding member of a large clade of podoviruses encoding a single-subunit RNA polymerase (RNAP). Phages of the family rely on host RNAP for transcription of early viral genes; viral RNAP transcribes non-early viral genes. T7 and its close relatives encode an inhibitor of host RNAP, the gp2 protein.

The mechanism of microcin C resistance provided by the MccF peptidase.

The heptapeptide-nucleotide microcin C (McC) is a potent inhibitor of enteric bacteria growth. Inside a sensitive cell, McC is processed by aminopeptidases, which release a nonhydrolyzable aspartyl-adenylate, a strong inhibitor of aspartyl-tRNA synthetase. The mccABCDE operon is sufficient for McC production and resistance of the producing cell to McC.

Transcription, processing and function of CRISPR cassettes in Escherichia coli.

CRISPR/Cas, bacterial and archaeal systems of interference with foreign genetic elements such as viruses or plasmids, consist of DNA loci called CRISPR cassettes (a set of variable spacers regularly separated by palindromic repeats) and associated cas genes. When a CRISPR spacer sequence exactly matches a sequence in a viral genome, the cell can become resistant to the virus. The CRISPR/Cas systems function through small RNAs originating from longer CRISPR cassette transcripts.