High variability and non-neutral evolution of the mammalian avpr1a gene

<p>Abstract</p> <p>Background</p> <p>The arginine-vasopressin 1a receptor has been identified as a key determinant for social behaviour in <it>Microtus </it>voles, humans and other mammals. Nevertheless, the genetic bases of complex phenotypic traits like differences in social and mating behaviour among species and individuals remain largely unknown. Contrary to previous studies focusing on differences in the promotor region of the gene, we investigate here the level of functional variation in the coding region (exon 1) of this locus.</p> <p>Results</p> <p>We detected high sequence diversity between higher mammalian taxa as well as between species of the genus <it>Microtus</it>. This includes length variation and radical amino acid changes, as well as the presence of distinct protein variants within individuals. Additionally, negative selection prevails on most parts of the first exon of the <it>arginine-vasopressin receptor 1a (avpr1a) </it>gene but it contains regions with higher rates of change that harbour positively selected sites. Synonymous and non-synonymous substitution rates in the <it>avpr1a </it>gene are not exceptional compared to other genes, but they exceed those found in related hormone receptors with similar functions.</p> <p>Discussion</p> <p>These results stress the importance of considering variation in the coding sequence of <it>avpr1a </it>in regards to associations with life history traits (e.g. social behaviour, mating system, habitat requirements) of voles, other mammals and humans in particular.</p

A Drug-Sensitive Genetic Network Masks Fungi from the Immune System

Fungal pathogens can be recognized by the immune system via their β-glucan, a potent proinflammatory molecule that is present at high levels but is predominantly buried beneath a mannoprotein coat and invisible to the host. To investigate the nature and significance of “masking” this molecule, we characterized the mechanism of masking and consequences of unmasking for immune recognition. We found that the underlying β-glucan in the cell wall of Candida albicans is unmasked by subinhibitory doses of the antifungal drug caspofungin, causing the exposed fungi to elicit a stronger immune response. Using a library of bakers' yeast (Saccharomyces cerevisiae) mutants, we uncovered a conserved genetic network that is required for concealing β-glucan from the immune system and limiting the host response. Perturbation of parts of this network in the pathogen C. albicans caused unmasking of its β-glucan, leading to increased β-glucan receptor-dependent elicitation of key proinflammatory cytokines from primary mouse macrophages. By creating an anti-inflammatory barrier to mask β-glucan, opportunistic fungi may promote commensal colonization and have an increased propensity for causing disease. Targeting the widely conserved gene network required for creating and maintaining this barrier may lead to novel broad-spectrum antimycotics

A Candida albicans CRISPR system permits genetic engineering of essential genes and gene families

Candida albicans is a pathogenic yeast that causes mucosal and systematic infections with high mortality. The absence of facile molecular genetics has been a major impediment to analysis of pathogenesis. The lack of meiosis coupled with the absence of plasmids makes genetic engineering cumbersome, especially for essential functions and gene families. We describe a C. albicans CRISPR system that overcomes many of the obstacles to genetic engineering in this organism. The high frequency with which CRISPR-induced mutations can be directed to target genes enables easy isolation of homozygous gene knockouts, even without selection. Moreover, the system permits the creation of strains with mutations in multiple genes, gene families, and genes that encode essential functions. This CRISPR system is also effective in a fresh clinical isolate of undetermined ploidy. Our method transforms the ability to manipulate the genome of Candida and provides a new window into the biology of this pathogen.National Institutes of Health (U.S.) (GM035010

Genomic Scans Support Repetitive Continental Colonization Events during the Rapid Radiation of Voles (Rodentia: Microtus): the Utility of AFLPs versus Mitochondrial and Nuclear Sequence Markers

Single locus studies might not resolve phylogenetic relationships and the evolutionary history of taxa. The analysis of multiple markers promises higher resolution, and congruence among loci may indicate that the phylogenies represent the underlying species history. Here, we examine the utility of a genome-wide approach based on amplified fragment length polymorphisms (AFLP) and several DNA sequence markers in resolving phylogenetic signals in the rapidly radiating rodent genus Microtus which produced about 70 vole species within the last 1.2-2 myr. The current Holarctic distribution of Microtus is assumed to have resulted from three independent colonization events out of Asia to North America, Europe, and northern Asia without subsequent colonization, which would have led to deep splits between species from different continents. We investigated this hypothesis of three single colonization events by reconstructing the phylogenetic relationships among species from all three continents based on data from the first exon of the nuclear arginine vasopressin receptor 1a gene (EXON1), an adjacent noncoding region and the mitochondrial cytochrome b gene. The phylogenetic patterns obtained from these sequence markers are contrasted to genome-wide data on more than 1800 amplified fragment length polymorphisms (AFLP) analyzed for the same samples. Our results show that the single sequence markers partially resolve the phylogenetic relationships within Microtus, but with some incongruence mostly between EXON1 and the other loci. However, deeper nodes of the radiation are only weakly supported and neither the combination of the markers nor additional nuclear sequences improved the resolution significantly. AFLPs provided much stronger support for major continent-specific clades, and show also that reciprocal monophyly of American and European voles is incomplete. Our results demonstrate that Microtus voles colonized the American and European continents each repeatedly in several independent events on similar colonization routes during their radiation. More generally, this study supports the suitability of AFLPs as an alternative to sequence markers to resolve the evolutionary history of rapidly radiating tax

Antisense Transcription Controls Cell Fate in Saccharomyces cerevisiae

SummaryEntry into meiosis is a key developmental decision. We show here that meiotic entry in Saccharomyces cerevisiae is controlled by antisense-mediated regulation of IME4, a gene required for initiating meiosis. In MAT a/α diploids the antisense IME4 transcript is repressed by binding of the a1/α2 heterodimer at a conserved site located downstream of the IME4 coding sequence. MAT a/α diploids that produce IME4 antisense transcript have diminished sense transcription and fail to initiate meiosis. Haploids that produce the sense transcript have diminished antisense transcription and manifest several diploid phenotypes. Our data are consistent with transcription interference as a regulatory mechanism at the IME4 locus that determines cell fate

FASTER MT: Isolation of Pure Populations of a and α Ascospores from Saccharomyces cerevisiae

The budding yeast Saccharomyces cerevisiae has many traits that make it useful for studies of quantitative inheritance. Genome-wide association studies and bulk segregant analyses often serve as first steps toward the identification of quantitative trait loci. These approaches benefit from having large numbers of ascospores pooled by mating type without contamination by vegetative cells. To this end, we inserted a gene encoding red fluorescent protein into the MATa locus. Red fluorescent protein expression caused MATa and a/α diploid vegetative cells and MATa ascospores to fluoresce; MATα cells without the gene did not fluoresce. Heterozygous diploids segregated fluorescent and nonfluorescent ascospores 2:2 in tetrads and bulk populations. The two populations of spores were separable by fluorescence-activated cell sorting with little cross contamination or contamination with diploid vegetative cells. This approach, which we call Fluorescent Ascospore Technique for Efficient Recovery of Mating Type (FASTER MT), should be applicable to laboratory, industrial, and undomesticated, strains

Ruler Arrays Reveal Haploid Genomic Structural Variation

Despite the known relevance of genomic structural variants to pathogen behavior, cancer, development, and evolution, certain repeat based structural variants may evade detection by existing high-throughput techniques. Here, we present ruler arrays, a technique to detect genomic structural variants including insertions and deletions (indels), duplications, and translocations. A ruler array exploits DNA polymerase’s processivity to detect physical distances between defined genomic sequences regardless of the intervening sequence. The method combines a sample preparation protocol, tiling genomic microarrays, and a new computational analysis. The analysis of ruler array data from two genomic samples enables the identification of structural variation between the samples. In an empirical test between two closely related haploid strains of yeast ruler arrays detected 78% of the structural variants larger than 100 bp.United States. National Institutes of Health (Grant R01GM069676

Engineered yeast tolerance enables efficient production from toxified lignocellulosic feedstocks

Lignocellulosic biomass remains unharnessed for the production of renewable fuels and chemicals due to challenges in deconstruction and the toxicity its hydrolysates pose to fermentation microorganisms. Here, we show in Saccharomyces cerevisiae that engineered aldehyde reduction and elevated extracellular potassium and pH are sufficient to enable near-parity production between inhibitor-laden and inhibitor-free feedstocks. By specifically targeting the universal hydrolysate inhibitors, a single strain is enhanced to tolerate a broad diversity of highly toxified genuine feedstocks and consistently achieve industrial-scale titers (cellulosic ethanol of &gt;100 grams per liter when toxified). Furthermore, a functionally orthogonal, lightweight design enables seamless transferability to existing metabolically engineered chassis strains: We endow full, multifeedstock tolerance on a xylose-consuming strain and one producing the biodegradable plastics precursor lactic acid. The demonstration of "drop-in" hydrolysate competence enables the potential of cost-effective, at-scale biomass utilization for cellulosic fuel and nonfuel products alike

Genomic scans support repetitive continental colonization events during the rapid radiation of voles (Rodentia: Microtus): the utility of AFLPs versus mitochondrial and nuclear sequence markers

Background According to the World Health Organization position paper, immunodeficiency such as human immunodeficiency virus (HIV) infection is a relative contraindication for specific immunotherapy (SIT). Since the introduction of highly active antiretroviral therapy, a significant reconstitution of immune competence in individuals with HIV is possible.Case Report In a 52-year-old man, HIV infection was diagnosed in 1987. Antiretroviral therapy was started in 1998. He presented himself in July 2001 because of an increasingly severe seasonal rhinoconjunctivitis. Symptoms were not sufficiently alleviated by various antiallergic drugs.Results The investigations showed a relevant sensitization to tree pollens. Specific immunotherapy with a tree pollen mix (hazel, birch, ash, and alder, 25% each) was started in November 2001. Viral load at this time was less than 50 copies/mL, the CD4+ cell count was 307/μL. Therapy was given in monthly intervals until mid-April 2005 without any side effects. Viral load and CD4+ cell counts did not change during SIT. Clinically, rhinoconjunctivitis was experienced only intermittently and symptom relief was almost 90%.Conclusions This report indicates that in patients with well-controlled HIV infection on highly active antiretroviral therapy, SIT with pollen extracts is a potential and successful therapeutic option. Keywords: human immunodeficiency virus, pollen allergy, respiratory allergy, allergy treatment, specific immunotherap