Cis-regulatory variants affect gene expression dynamics in yeast.

Evolution of cis-regulatory sequences depends on how they affect gene expression and motivates both the identification and prediction of cis-regulatory variants responsible for expression differences within and between species. While much progress has been made in relating cis-regulatory variants to expression levels, the timing of gene activation and repression may also be important to the evolution of cis-regulatory sequences. We investigated allele-specific expression (ASE) dynamics within and between Saccharomyces species during the diauxic shift and found appreciable cis-acting variation in gene expression dynamics. Within-species ASE is associated with intergenic variants, and ASE dynamics are more strongly associated with insertions and deletions than ASE levels. To refine these associations, we used a high-throughput reporter assay to test promoter regions and individual variants. Within the subset of regions that recapitulated endogenous expression, we identified and characterized cis-regulatory variants that affect expression dynamics. Between species, chimeric promoter regions generate novel patterns and indicate constraints on the evolution of gene expression dynamics. We conclude that changes in cis-regulatory sequences can tune gene expression dynamics and that the interplay between expression dynamics and other aspects of expression is relevant to the evolution of cis-regulatory sequences.

Comparative Genomics Approaches Accurately Predict Deleterious Variants in Plants.

Recent advances in genome resequencing have led to increased interest in prediction of the functional consequences of genetic variants. Variants at phylogenetically conserved sites are of particular interest, because they are more likely than variants at phylogenetically variable sites to have deleterious effects on fitness and contribute to phenotypic variation. Numerous comparative genomic approaches have been developed to predict deleterious variants, but the approaches are nearly always assessed based on their ability to identify known disease-causing mutations in humans. Determining the accuracy of deleterious variant predictions in nonhuman species is important to understanding evolution, domestication, and potentially to improving crop quality and yield. To examine our ability to predict deleterious variants in plants we generated a curated database of 2,910 Arabidopsis thaliana mutants with known phenotypes. We evaluated seven approaches and found that while all performed well, their relative ranking differed from prior benchmarks in humans. We conclude that deleterious mutations can be reliably predicted in A. thaliana and likely other plant species, but that the relative performance of various approaches does not necessarily translate from one species to another.

Adaptive introgression of anticoagulant rodent poison resistance by hybridization between old world mice.

Polymorphisms in the vitamin K 2,3-epoxide reductase subcomponent 1 (vkorc1) of house mice (Mus musculus domesticus) can cause resistance to anticoagulant rodenticides such as warfarin [1-3]. Here we show that resistant house mice can also originate from selection on vkorc1 polymorphisms acquired from the Algerian mouse (M. spretus) through introgressive hybridization. We report on a polymorphic introgressed genomic region in European M. m. domesticus that stems from M. spretus, spans >10 Mb on chromosome 7, and includes the molecular target of anticoagulants vkorc1 [1-4]. We show that in the laboratory, the homozygous complete vkorc1 allele of M. spretus confers resistance when introgressed into M. m. domesticus. Consistent with selection on the introgressed allele after the introduction of rodenticides in the 1950s, we found signatures of selection in patterns of variation in M. m. domesticus. Furthermore, we detected adaptive protein evolution of vkorc1 in M. spretus (Ka/Ks = 1.54-1.93) resulting in radical amino acid substitutions that apparently cause anticoagulant tolerance in M. spretus as a pleiotropic effect. Thus, positive selection produced an adaptive, divergent, and pleiotropic vkorc1 allele in the donor species, M. spretus, which crossed a species barrier and produced an adaptive polymorphic trait in the recipient species, M. m. domesticus.

Genome-wide analysis of the cis-regulatory modules of divergent gene pairs in yeast.

In budding yeast, approximately a quarter of adjacent genes are divergently transcribed (divergent gene pairs). Whether genes in a divergent pair share the same regulatory system is still unknown. By examining transcription factor (TF) knockout experiments, we found that most TF knockout only altered the expression of one gene in a divergent pair. This prompted us to conduct a comprehensive analysis in silico to estimate how many divergent pairs are regulated by common sets of TFs (cis-regulatory modules, CRMs) using TF binding sites and expression data. Analyses of ten expression datasets show that only a limited number of divergent gene pairs share CRMs in any single dataset. However, around half of divergent pairs do share a regulatory system in at least one dataset. Our analysis suggests that genes in a divergent pair tend to be co-regulated in at least one condition; however, in most conditions, they may not be co-regulated.

Functional and evolutionary correlates of gene constellations in the Drosophila melanogaster genome that deviate from the stereotypical gene architecture.

BACKGROUND: The biological dimensions of genes are manifold. These include genomic properties, (e.g., X/autosomal linkage, recombination) and functional properties (e.g., expression level, tissue specificity). Multiple properties, each generally of subtle influence individually, may affect the evolution of genes or merely be (auto-)correlates. Results of multidimensional analyses may reveal the relative importance of these properties on the evolution of genes, and therefore help evaluate whether these properties should be considered during analyses. While numerous properties are now considered during studies, most work still assumes the stereotypical solitary gene as commonly depicted in textbooks. Here, we investigate the Drosophila melanogaster genome to determine whether deviations from the stereotypical gene architecture correlate with other properties of genes. RESULTS: Deviations from the stereotypical gene architecture were classified as the following gene constellations: Overlapping genes were defined as those that overlap in the 5-prime, exonic, or intronic regions. Chromatin co-clustering genes were defined as genes that co-clustered within 20 kb of transcriptional territories. If this scheme is applied the stereotypical gene emerges as a rare occurrence (7.5%), slightly varied schemes yielded between approximately 1%-50%. Moreover, when following our scheme, paired-overlapping genes and chromatin co-clustering genes accounted for 50.1 and 42.4% of the genes analyzed, respectively. Gene constellation was a correlate of a number of functional and evolutionary properties of genes, but its statistical effect was approximately 1-2 orders of magnitude lower than the effects of recombination, chromosome linkage and protein function. Analysis of datasets on male reproductive proteins showed these were biased in their representation of gene constellations and evolutionary rate Ka/Ks estimates, but these biases did not overwhelm the biologically meaningful observation of high evolutionary rates of male reproductive genes. CONCLUSION: Given the rarity of the solitary stereotypical gene, and the abundance of gene constellations that deviate from it, the presence of gene constellations, while once thought to be exceptional in large Eukaryote genomes, might have broader relevance to the understanding and study of the genome. However, according to our definition, while gene constellations can be significant correlates of functional properties of genes, they generally are weak correlates of the evolution of genes. Thus, the need for their consideration would depend on the context of studies.

Co-expression of adjacent genes in yeast cannot be simply attributed to shared regulatory system.

BACKGROUND: Adjacent gene pairs in the yeast genome have a tendency to express concurrently. Sharing of regulatory elements within the intergenic region of those adjacent gene pairs was often considered the major mechanism responsible for such co-expression. However, it is still in debate to what extent that common transcription factors (TFs) contribute to the co-expression of adjacent genes. In order to resolve the evolutionary aspect of this issue, we investigated the conservation of adjacent pairs in five yeast species. By using the information for TF binding sites in promoter regions available from the MYBS database http://cg1.iis.sinica.edu.tw/~mybs/, the ratios of TF-sharing pairs among all the adjacent pairs in yeast genomes were analyzed. The levels of co-expression in different adjacent patterns were also compared. RESULTS: Our analyses showed that the proportion of adjacent pairs conserved in five yeast species is relatively low compared to that in the mammalian lineage. The proportion was also low for adjacent gene pairs with shared TFs. Particularly, the statistical analysis suggested that co-expression of adjacent gene pairs was not noticeably associated with the sharing of TFs in these pairs. We further proposed a case of the PAC (polymerase A and C) and RRPE (rRNA processing element) motifs which co-regulate divergent/bidirectional pairs, and found that the shared TFs were not significantly relevant to co-expression of divergent promoters among adjacent genes. CONCLUSION: Our findings suggested that the commonly shared cis-regulatory system does not solely contribute to the co-expression of adjacent gene pairs in yeast genome. Therefore we believe that during evolution yeasts have developed a sophisticated regulatory system that integrates both TF-based and non-TF based mechanisms(s) for concurrent regulation of neighboring genes in response to various environmental changes.

MYBS: a comprehensive web server for mining transcription factor binding sites in yeast.

Correct interactions between transcription factors (TFs) and their binding sites (TFBSs) are of central importance to gene regulation. Recently developed chromatin-immunoprecipitation DNA chip (ChIP-chip) techniques and the phylogenetic footprinting method provide ways to identify TFBSs with high precision. In this study, we constructed a user-friendly interactive platform for dynamic binding site mapping using ChIP-chip data and phylogenetic footprinting as two filters. MYBS (Mining Yeast Binding Sites) is a comprehensive web server that integrates an array of both experimentally verified and predicted position weight matrixes (PWMs) from eleven databases, including 481 binding motif consensus sequences and 71 PWMs that correspond to 183 TFs. MYBS users can search within this platform for motif occurrences (possible binding sites) in the promoters of genes of interest via simple motif or gene queries in conjunction with the above two filters. In addition, MYBS enables users to visualize in parallel the potential regulators for a given set of genes, a feature useful for finding potential regulatory associations between TFs. MYBS also allows users to identify target gene sets of each TF pair, which could be used as a starting point for further explorations of TF combinatorial regulation. MYBS is available at http://cg1.iis.sinica.edu.tw/~mybs/.

Effects of Tamm-Horsfall protein on the protection of MCDK cells from oxalate induced free radical injury.

Renal cell injury and fixed particle formation is one of the theories of urinary stone formation. The exposure of renal epithelial cells to oxalate ions and calcium oxalate monohydrate crystals can cause free radical generation and increase lipid peroxidation. Tamm-Horsfall protein (THP) has a protective effect on the production of free radicals in vitro. We aimed to show that THP (and its deglycosylated products, D-THP) could protect culture cells from free radical injury in vivo as well as the possible mechanism by which this is done. Exposure of Madin-Darby canine kidney (MDCK) cells to Ox resulted in a significant increase in the release LDH, NBT and MDA, as well as an increase in caspase 3 activity, all of which were further elevated when COM crystals were added. With the addition of THP at 500 nM, there was a significant decrease in the release of LDH and the production of MDA and NBT. A decrease in capase 3 activity was observed when 500 nM THP was added to the culture medium that reached 32.7% and 40.4% of inhibition in CaOx+THP and CaOx+COM+THP, respectively. THP decreased the adhesion of COM crystals to the MDCK cells but lost its effect when THP was deglycosylated. The results indicate that both Ox and COM crystals cause the release of LDH, MDA, NBT and increase the activity of capase 3 in MDCK cells. As a free radical scavenger, THP reduces the amount of free radicals and provides significant protection at a critical concentration of 500 nM. The deglycosylated THP decreased the effect of the protection of the MDCK cells from oxalate-induced injury and an increase of adhesion of the COM crystals to the MDCK cells. Therefore, the effects of THP on the protection of oxalate induced radical injury may be partly due to its intact glycosylation and its adhesion to the cell membrane.