Multiplex PCR method for detection of variability in genes encoding the alpha proteins of Streptococcus agalactiae

The majority of Streptococcus agalactiae strains express one or more surface-anchored proteins that vary by strain. These proteins, which are characteristic for S. agalactiae, and are able to induce protective antibodies, include the alpha C, Rib proteins, alpha-like protein 2, and alpha-like protein 3. In this study was developed multiplex PCR method for detection of genes encoding these proteins, and its occurrence within a various S. agalactiae isolates of bovine origin. Also are reported two new genes from bovine isolates of S. agalactiae amplified by PCR, encoding other putative members of the family, alphalike protein 6, and alpha-like protein 7. They contain an overall genetic organization highly similar to that of the alpha C and Rib proteins.

Design, construction and optimization of a synthetic RNA polymerase operon in Escherichia coli.

Prokaryotic genes encoding functionally related proteins are often clustered in operons. The compact structure of operons allows for co-transcription of the genes, and for co-translation of the polycistronic messenger RNA to the corresponding proteins. This leads to reduced regulatory complexity and enhanced gene expression efficiency, and as such to an overall metabolic benefit for the protein production process in bacteria and archaea. Interestingly, the genes encoding the subunits of one of the most conserved and ubiquitous protein complexes, the RNA polymerase, are not clustered in a single operon. Rather, its genes are scattered in all known prokaryotic genomes, generally integrated in different ribosomal operons. To analyze the impact of this genetic organization on the fitness of Escherichia coli, we constructed a bacterial artificial chromosome harboring the genes encoding the RNA polymerase complex in a single operon. Subsequent deletion of the native chromosomal genes led to a reduced growth on minimal medium. However, by using adaptive laboratory evolution the growth rate was restored to wild-type level. Hence, we show that a highly conserved genetic organization of core genes in a bacterium can be reorganized by a combination of design, construction and optimization, yielding a well-functioning synthetic genetic architecture.

The genetic organization of longitudinal subcortical volumetric change is stable throughout the lifespan

Development and aging of the cerebral cortex show similar topographic organization and are governed by the same genes. It is unclear whether the same is true for subcortical regions, which follow fundamentally different ontogenetic and phylogenetic principles. We tested the hypothesis that genetically governed neurodevelopmental processes can be traced throughout life by assessing to which degree brain regions that develop together continue to change together through life. Analyzing over 6000 longitudinal MRIs of the brain, we used graph theory to identify five clusters of coordinated development, indexed as patterns of correlated volumetric change in brain structures. The clusters tended to follow placement along the cranial axis in embryonic brain development, suggesting continuity from prenatal stages, and correlated with cognition. Across independent longitudinal datasets, we demonstrated that developmental clusters were conserved through life. Twin-based genetic correlations revealed distinct sets of genes governing change in each cluster. Single nucleotide polymorphisms-based analyses of 38127 cross-sectional MRIs showed a similar pattern of genetic volume-volume correlations. In conclusion, coordination of subcortical change adheres to fundamental principles of lifespan continuity and genetic organization.

Genome sequencing of the neotype strain CBS 554.65 reveals the MAT1-2 locus of Aspergillus niger

Background Aspergillus niger is a ubiquitous filamentous fungus widely employed as a cell factory thanks to its abilities to produce a wide range of organic acids and enzymes. Due to its economic importance and its role as model organism to study fungal fermentation, its genome was one of the first Aspergillus genomes to be sequenced in 2007. Nowadays, the genome sequences of at least five other A. niger strains are available. These, however, do not include the neotype strain CBS 554.65. Results In this study, the genome of CBS 554.65 was sequenced with PacBio. A high-quality nuclear genome sequence consisting of 17 contigs with a N50 value of 4.07 Mbp was obtained. The sequencing covered all the 8 centromeric regions of the chromosomes. In addition, a complete circular mitochondrial DNA assembly was obtained. In silico analyses revealed the presence of a MAT1-2-1 gene in this genome, contrary to the so far sequenced A. niger strains, which all contain a MAT1-1-1 gene. An alignment at the MAT locus showed a different position of the MAT1-1-1 gene of ATCC 1015 compared to the MAT1-2-1 gene of CBS 554.65, relative to the surrounding genes. In addition, 24 other sequenced isolates of A. niger showed a 1:1 ratio of MAT1-1 and MAT1-2 loci. While the genetic organization of the MAT1-2 locus of CBS 554.65 is similar to what is found in other aspergilli, the genetic organization of the MAT1-1 locus is flipped in all sequenced strains. Conclusions This study, besides providing a high-quality genome sequence of an important A. niger strain, suggests the occurrence of genetic flipping or switching events at the MAT1-1 locus of A. niger. These results provide new insights in the mating system of A. niger and could contribute to the investigation and potential discovery of sexuality of this so far asexual fungal species.

Bacteria have numerous distinctive groups of phage–plasmids with conserved phage and variable plasmid gene repertoires

Plasmids and temperate phages are key contributors to bacterial evolution. They are usually regarded as very distinct. However, some elements, termed phage–plasmids, are known to be both plasmids and phages, e.g. P1, N15 or SSU5. The number, distribution, relatedness and characteristics of these phage–plasmids are poorly known. Here, we screened for these elements among ca. 2500 phages and 12000 plasmids and identified 780 phage–plasmids across very diverse bacterial phyla. We grouped 92% of them by similarity of gene repertoires to eight defined groups and 18 other broader communities of elements. The existence of these large groups suggests that phage–plasmids are ancient. Their gene repertoires are large, the average element is larger than an average phage or plasmid, and they include slightly more homologs to phages than to plasmids. We analyzed the pangenomes and the genetic organization of each group of phage–plasmids and found the key phage genes to be conserved and co-localized within distinct groups, whereas genes with homologs in plasmids are much more variable and include most accessory genes. Phage–plasmids are a sizeable fraction of the sequenced plasmids (∼7%) and phages (∼5%), and could have key roles in bridging the genetic divide between phages and other mobile genetic elements.

Genetic organization and cellular specificity of S. aureus leukocidins

Currently, infections produced by the gram-positive bacteria S. aureus represent a significant healthcare burden throughout the world. This is attributed to the ability of this bacterium to develop antibiotic resistance and efficiently evade human immune response. Therefore, research effort of many scientific laboratories worldwide is directed toward characterization of the genetic organization and molecular mechanisms responsible for S. aureus pathogenesis. This report is aimed to describe the growing body of evidence related to our understanding of the genetic organization and molecular interactions of the S. aureus leukocidins with the human cells that play an important role in bacterial pathogenesis, and represent a significant healthcare burden. Understanding of the genetic organization linked with additional mechanisms responsible for the realization of toxic potential, can help us to develop a better personalized approach for therapy against S. aureus infections. Thus, improved understanding of the molecular interactions between S. aureus leucocidins, and cell surface receptors may lead to the development of the alternative anti-microbial agents, that either independently or in combination with the current antibiotic treatment regimens will be used as an effective treatment strategy in clinic.

Study of the genetic organization of the strain Rhizobium leguminosarum bv. trifolii forming a symbiosis with clover Trifolium ambiguum

R. leguminosarum bv. trifolii strains are characterized by narrow host specificity. We have identified a strain that forms nodules on several types of clover and studied the genetic organization of its symbiotic region.

Bacteria have numerous phage-plasmid families with conserved phage and variable plasmid gene repertoires

ABSTRACTPlasmids and temperate phages are mobile genetic elements driving bacterial evolution. They are usually regarded as very distinct. However, some elements, termed phage-plasmids, are known to be both plasmids and phages, e.g. P1, N15 or SSU5. The number, distribution, relatedness and characteristics of these phage-plasmids are poorly known. Here, we screened for these elements among ca. 14000 phages and plasmids and identified 780 phage-plasmids across very diverse bacterial phyla. We grouped 92% of them by similarity of gene repertoires to define 8 families and 18 other broader communities of elements. The existence of these large groups suggests that phage-plasmids are ancient. Their gene repertoires are large, the average element is larger than an average phage or plasmid, and they include slightly more homologs to phages than to plasmids. We analyzed the pangenomes and the genetic organization of each group of phage-plasmids and found the key phage genes to be conserved and co-localized within families, whereas genes with homologs in plasmids are much more variable and include most accessory genes. Phage-plasmids are a sizeable fraction of all phages and plasmids and could have key roles in bridging the genetic divide between phages and other mobile genetic elements.

High-throughput discovery and characterization of human transcriptional effectors

SummaryThousands of proteins localize to the nucleus; however, it remains unclear which contain transcriptional effectors. Here, we develop HT-recruit - a pooled assay where protein libraries are recruited to a reporter, and their transcriptional effects are measured by sequencing. Using this approach, we measure gene silencing and activation for thousands of domains. We find a relationship between repressor function and evolutionary age for the KRAB domains, discover Homeodomain repressor strength is collinear with Hox genetic organization, and identify activities for several Domains of Unknown Function. Deep mutational scanning of the CRISPRi KRAB maps the co-repressor binding surface and identifies substitutions that improve stability/silencing. By tiling 238 proteins, we find repressors as short as 10 amino acids. Finally, we report new activator domains, including a divergent KRAB. Together, these results provide a resource of 600 human proteins containing effectors and demonstrate a scalable strategy for assigning functions to protein domains.