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Abstract Background Figures of phylogenetic trees are widely used to illustrate the result of evolutionary analyses. However, one cannot easily extract a machine-readable representation from such images. Therefore, new software emerges that helps to preserve phylogenies digitally for future research. Results TreeSnatcher Plus is a GUI-driven JAVA application that semi-automatically generates a Newick format for multifurcating, arbitrarily shaped, phylogenetic trees contained in pixel images. It offers a range of image pre-processing methods and detects the topology of a depicted tree with adequate user assistance. The user supervises the recognition process, makes corrections to the image and to the topology and repeats steps if necessary. At the end TreeSnatcher Plus produces a Newick tree code optionally including branch lengths for rectangular and freeform trees. Conclusions Although illustrations of phylogenies exist in a vast number of styles, TreeSnatcher Plus imposes no limitations on the images it can process with adequate user assistance. Given that a fully automated digitization of all figures of phylogenetic trees is desirable but currently unrealistic, TreeSnatcher Plus is the only program that reliably facilitates at least a semi-automatic conversion from such figures into a machine-readable format.

Abstract Background The regulatory information encoded in the DNA of promoter regions usually enforces a minimal, non-zero distance between the coding regions of neighboring genes. However, the size of this minimal regulatory space is not generally known. In particular, it is unclear if minimal promoter size differs between species and between uni- and bi-directionally acting regulatory regions. Results Analyzing the genomes of 11 yeasts, we show that the lower size limit on promoter-containing regions is species-specific within a relatively narrow range (80-255 bp). This size limit applies equally to regions that initiate transcription on one or both strands, indicating that bi-directional promoters and uni-directional promoters are constrained similarly. We further find that young, species-specific regions are on average much longer than older regions, suggesting either a bias towards deletions or selection for genome compactness in yeasts. While the length evolution of promoter-less intergenic regions is well described by a simplistic, purely neutral model, regions containing promoters typically show an excess of unusually long regions. Regions flanked by divergently transcribed genes have a bi-modal length distribution, with short lengths found preferentially among older regions. These old, short regions likely harbor evolutionarily conserved bi-directionally active promoters. Surprisingly, some of the evolutionarily youngest regions in two of the eleven species (S. cerevisiae and K. waltii) are shorter than the lower limit observed in older regions. Conclusions The minimal chromosomal space required for transcriptional regulation appears to be relatively similar across yeast species, and is the same for uni-directional and bi-directional promoters. New intergenic regions created by genome rearrangements tend to evolve towards the more narrow size distribution found among older regions.

Abstract Background Many functional, structural and evolutionary features of human genes have been observed to correlate with expression breadth and/or gene age. Here, we systematically explore these correlations. Results Gene age and expression breadth are strongly correlated, but contribute independently to the variation of functional, structural and evolutionary features, even when we take account of variation in mRNA expression level. Human genes without orthologs in distant species ('young' genes) tend to be tissue-specific in their expression. As computational inference of gene function often relies on the existence of homologs in other species, and experimental characterization is facilitated by broad and high expression, young, tissue-specific human genes are often the least characterized. At the same time, young genes are most likely to be medically relevant. Conclusions Our results indicate that functional characterization of human genes is biased against young, tissue-specific genes that are mostly medically relevant. The biases should not be taken lightly because they may pose serious obstacles to our understanding of the molecular basis of human diseases. Future studies should thus be designed to specifically explore the properties of primate-specific genes.

Abstract Background Among bacteria and archaea, amino acid usage is correlated with habitat temperatures. In particular, protein surfaces in species thriving at higher temperatures appear to be enriched in amino acids that stabilize protein structure and depleted in amino acids that decrease thermostability. Does this observation reflect a causal relationship, or could the apparent trend be caused by phylogenetic relatedness among sampled organisms living at different temperatures? And do proteins from endothermic and exothermic vertebrates show similar differences? Results We find that the observed correlations between the frequencies of individual amino acids and prokaryotic habitat temperature are strongly influenced by evolutionary relatedness between the species analysed; however, a proteome-wide bias towards increased thermostability remains after controlling for phylogeny. Do eukaryotes show similar effects of thermal adaptation? A small shift of amino acid usage in the expected direction is observed in endothermic ('warm-blooded') mammals and chicken compared to ectothermic ('cold-blooded') vertebrates with lower body temperatures; this shift is not simply explained by nucleotide usage biases. Conclusion Protein homologs operating at different temperatures have different amino acid composition, both in prokaryotes and in vertebrates. Thus, during the transition from ectothermic to endothermic life styles, the ancestors of mammals and of birds may have experienced weak genome-wide positive selection to increase the thermostability of their proteins.

Abstract Background In all eukaryotic species examined, genes that are chromosomal neighbours are more similar in their expression than random gene pairs. Currently, it is still unclear how much of this local co-expression is caused by direct transcriptional interactions, and how much is due to shared chromatin environments. Results We analysed neighbouring genes in Arabidopsis thaliana. At large intergenic distances (>400 bp), divergently and convergently transcribed gene pairs show very similar levels of co-expression, mediated most likely by shared chromatin environments. At gene distances below 400 bp, co-expression is strongly enhanced only for divergently transcribed gene pairs, indicating bi-directional transcription from a single promoter. Conversely, co-expression is suppressed for short convergently or uni-directionally transcribed pairs. This suppression points to transcriptional interference concentrated at the 3' end, e.g., in the context of transcription termination. Conclusions Classifying linked gene pairs by their orientation, we are able to partially tease apart the different levels of regional expression modulation. (i) Regional chromatin characteristics modulate the accessibility for regulation and transcription, regardless of gene orientation; the strength of this chromatin effect can be assessed from divergently or convergently transcribed distant neighbours. (ii) Shared promoter regions up to 400 bp in length enhance the co-expression of close bi-directional neighbours. (iii) Transcriptional interference of close neighbours is concentrated at the 3' ends of genes, and reduces co-expression on average by 40%.

Abstract Background A prerequisite for a duplication to spread through and persist in a given population is retaining expression of both gene copies. Yet changing a gene's dosage is frequently detrimental to fitness. Consequently, dosage-sensitive genes are less likely to duplicate. However, in cases where the level of gene product is controlled, via negative feedback, by its own abundance, an increase in gene copy number can in principle be decoupled from an increase in protein while both copies remain expressed. Using data from the transcriptional networks of E. coli and S. cerevisiae, we test the hypothesis that genes under negative auto-regulation show enhanced duplicability. Results Controlling for several known correlates of duplicability, we find no statistically significant support in either E. coli or S. cerevisiae that transcription factors under negative auto-regulation hold a duplicability advantage over transcription factors with no auto-regulation. Conclusion Based on the analysis of transcriptional networks in E. coli and S. cerevisiae, there is no evidence that negative auto-regulation has contributed, on a genome-wide scale, to the variability in gene family sizes in these species.

Abstract Background Genome sequencing projects and comparative genomics studies typically aim to trace the evolutionary history of large gene sets, often requiring human inspection of hundreds of phylogenetic trees. If trees are checked for compatibility with an explicit null hypothesis (e.g., the monophyly of certain groups), this daunting task is greatly facilitated by an appropriate coloring scheme. Findings In this note, we introduce ColorTree, a simple yet powerful batch customization tool for phylogenic trees. Based on pattern matching rules, ColorTree applies a set of customizations to an input tree file, e.g., coloring labels or branches. The customized trees are saved to an output file, which can then be viewed and further edited by Dendroscope (a freely available tree viewer). ColorTree runs on any Perl installation as a stand-alone command line tool, and its application can thus be easily automated. This way, hundreds of phylogenic trees can be customized for easy visual inspection in a matter of minutes. Conclusion ColorTree allows efficient and flexible visual customization of large tree sets through the application of a user-supplied configuration file to multiple tree files.

Abstract Background The rate of molecular evolution varies widely between proteins, both within and among lineages. To what extent is this variation influenced by genome-wide, lineage-specific effects? To answer this question, we assess the rate variation between insect lineages for a large number of orthologous genes. Results When compared to the beetle Tribolium castaneum, we find that the stem lineage of flies and mosquitoes (Diptera) has experienced on average a 3-fold increase in the rate of evolution. Pairwise gene comparisons between Drosophila and Tribolium show a high correlation between evolutionary rates of orthologous proteins. Conclusion Gene specific divergence rates remain roughly constant over long evolutionary times, modulated by genome-wide, lineage-specific effects. Among the insects analysed so far, it appears that the Tribolium genes show the lowest rates of divergence. This has the practical consequence that homology searches for human genes yield significantly better matches in Tribolium than in Drosophila. We therefore suggest that Tribolium is better suited for comparisons between phyla than the widely employed dipterans.

Abstract Background Prior to the sequencing of the human genome it was typically assumed that, tandem duplication aside, gene order is for the most part random. Numerous observers, however, highlighted instances in which a ligand was linked to one of its cognate receptors, with some authors suggesting that this may be a general and/or functionally important pattern, possibly associated with recombination modification between epistatically interacting loci. Here we ask whether ligands are more closely linked to their receptors than expected by chance. Results We find no evidence that ligands are linked to their receptors more closely than expected by chance. However, in the human genome there are approximately twice as many co-occurrences of ligand and receptor on the same human chromosome as expected by chance. Although a weak effect, the latter might be consistent with a past history of block duplication. Successful duplication of some ligands, we hypothesise, is more likely if the cognate receptor is duplicated at the same time, so ensuring appropriate titres of the two products. Conclusion While there is an excess of ligands and their receptors on the same human chromosome, this cannot be accounted for by classical models of non-random gene order, as the linkage of ligands/receptors is no closer than expected by chance. Alternative hypotheses for non-random gene order are hence worth considering.

Plant roots are functionally heterogeneous in cellular architecture, transcriptome profile, metabolic state, and microbial immunity. We hypothesized that axial differentiation may also impact spatial colonization by root microbiota along the root axis. We developed two growth systems, ArtSoil and CD-Rhizotron, to grow and then dissect Arabidopsis thaliana roots into three segments. We demonstrate that distinct endospheric and rhizosphere bacterial communities colonize the segments, supporting the hypothesis of microbiota differentiation along the axis. Root metabolite profiling of each segment reveals differential metabolite enrichment and specificity. Bioinformatic analyses and GUS histochemistry indicate microbe-induced accumulation of SWEET2, 4, and 12 sugar uniporters. Profiling of root segments from sweet mutants shows altered spatial metabolic profiles and reorganization of endospheric root microbiota. This work reveals the interdependency between root metabolites and microbial colonization and the contribution of SWEETs to spatial diversity and stability of microbial ecosystem., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2024 The Author(s). Published by Elsevier Inc. All rights reserved.)

Sucrose, hexoses, and raffinose play key roles in the plant metabolism. Sucrose and raffinose, produced by photosynthesis, are translocated from leaves to flowers, developing seeds and roots. Translocation occurs in the sieve elements or sieve tubes of angiosperms. But how is sucrose loaded into and unloaded from the sieve elements? There seem to be two principal routes: one through plasmodesmata and one via the apoplasm. The best-studied transporters are the H+/SUCROSE TRANSPORTERs (SUTs) in the sieve element-companion cell complex. Sucrose is delivered to SUTs by SWEET sugar uniporters that release these key metabolites into the apoplasmic space. The H+/amino acid permeases and the UmamiT amino acid transporters are hypothesized to play analogous roles as the SUT-SWEET pair to transport amino acids. SWEETs and UmamiTs also act in many other important processes-for example, seed filling, nectar secretion, and pollen nutrition. We present information on cell type-specific enrichment of SWEET and UmamiT family members and propose several members to play redundant roles in the efflux of sucrose and amino acids across different cell types in the leaf. Pathogens hijack SWEETs and thus represent a major susceptibility of the plant. Here, we provide an update on the status of research on intercellular and long-distance translocation of key metabolites such as sucrose and amino acids, communication of the plants with the root microbiota via root exudates, discuss the existence of transporters for other important metabolites and provide potential perspectives that may direct future research activities., (© The Author(s) 2021. Published by Oxford University Press on behalf of American Society of Plant Biologists.)

The leaf vasculature plays a key role in solute translocation. Veins consist of at least seven distinct cell types, with specific roles in transport, metabolism, and signaling. Little is known about leaf vascular cells, in particular the phloem parenchyma (PP). PP effluxes sucrose into the apoplasm as a basis for phloem loading, yet PP has been characterized only microscopically. Here, we enriched vascular cells from Arabidopsis leaves to generate a single-cell transcriptome atlas of leaf vasculature. We identified at least 19 cell clusters, encompassing epidermis, guard cells, hydathodes, mesophyll, and all vascular cell types, and used metabolic pathway analysis to define their roles. Clusters comprising PP cells were enriched for transporters, including SWEET11 and SWEET12 sucrose and UmamiT amino acid efflux carriers. We provide evidence that PP development occurs independently from ALTERED PHLOEM DEVELOPMENT, a transcription factor required for phloem differentiation. PP cells have a unique pattern of amino acid metabolism activity distinct from companion cells (CCs), explaining differential distribution/metabolism of amino acids in veins. The kinship relation of the vascular clusters is strikingly similar to the vein morphology, except for a clear separation of CC from the other vascular cells including PP. In summary, our single-cell RNA-sequencing analysis provides a wide range of information into the leaf vasculature and the role and relationship of the leaf cell types., (© The Author(s) 2021. Published by Oxford University Press on behalf of American Society of Plant Biologists.)

Previously, we have identified single nucleotide polymorphisms (SNPs) rs7958904 and rs4759314 in long non-coding RNA HOX transcript antisense RNA (HOTAIR) were significantly associated with risk of colorectal and gastric cancer, respectively. Here, we aimed to investigate the association between HOTAIR SNPs and cervical cancer (CC) susceptibility. A total of 1209 cases and 1348 controls were enrolled for association study and genotyped with TaqMan allelic discrimination method. The Cancer Genome Atlas (TCGA) database was utilized for in vivo analysis of allele-specific HOTAIR expression. MTT assay was employed for evaluation of allele-specific cell proliferation. The rs7958904 CC genotype was related to an increased risk of cervical cancer compared with the GG/GC genotypes (OR = 1.57, 95% CI = 1.10-2.25). TCGA database showed the CC tissues with rs7958904 CC genotype had higher HOTAIR expression than those with GG genotype (P = 0.046). MTT assay demonstrated a growth-promoting role of rs7958904 C allele on CC cells. Further functional studies on the effect of rs7958904 on biological behavior of CC cells are needed to confirm and extend our findings. In conclusion, HOTAIR rs7958904 might influence CC susceptibility through modulation of CC cell proliferation, and could serve as a diagnostic biomarker.

Objective: Glyoxalase 1 (GLO1) is ubiquitously expressed in the cytosol of the cell and is the major opponent against the reactive metabolite methylglyoxal, which is involved in the development of atherosclerosis. Nondiabetic individuals with an increased hemoglobin A1c (HbA1c) level are at higher risk for development of cardiovascular diseases. As such, this study investigated whether there was an association between reduced GLO1 activity in atherosclerotic lesions of nondiabetic patients with an increased HbA1c level., Methods: HbA1c level was determined in venous blood of patients with carotid artery disease. Protein level of GLO1 was measured in endarterectomy-derived carotid artery plaques by Western blotting. Activity was measured by spectrophotometric assay in the plaques as well as in the erythrocytes; GLO1 activity in erythrocytes was compared with that in a cohort of healthy individuals (n = 15; 33% men; average age, 60 years)., Results: There were 36 patients with carotid artery disease (69% men; average age, 69 years) included in this study and divided into two equal groups: group I, HbA1c < 5.7% (<39 mmol/mol); and group II, 5.7% ≤ HbA1c < 6.5% (39 mmol/mol ≤ HbA1c < 48 mmol/mol). GLO1 activity in carotid plaques was reduced by 29% in group II compared with group I (P = .048), whereas protein expression was unchanged (P = .25). Analysis of GLO1 activity in erythrocytes revealed no difference between the groups (P = .36) or in comparison to healthy controls (P = .15). Examination of clinical parameters showed an increased amount of patients with concomitant peripheral arterial disease in group II (44% vs 10%; P = .020)., Conclusions: Reduction of GLO1 activity in atherosclerotic lesions of nondiabetic patients with increased HbA1c is associated with a functional involvement of this protective enzyme in atherogenesis. Systemic GLO1 activity seems to be independent of both HbA1c and localized atherosclerosis as it was unchanged between group I and group II as well as compared with healthy controls, respectively., (Copyright © 2016 Society for Vascular Surgery. Published by Elsevier Inc. All rights reserved.)

The sex chromosomes and autosomes spend different times in the germ line of the two sexes. If cell division is mutagenic and if the sexes differ in number of cell divisions, then we expect that sequences on the X and Y chromosomes and autosomes should mutate at different rates. Tests of this hypothesis for several mammalian species have led to conflicting results. At the same time, recent evidence suggests that the chromosomal location of genes on autosomes affects their rate of evolution at synonymous sites. This suggests a mutagenic source different from germ cell replication. To correctly interpret the previous estimates of male mutation bias, it is crucial to understand the degree and range of this local similarity. With a carefully chosen randomization protocol, local similarity in synonymous rates of evolution can be detected in human-rodent and mouse-rat comparisons. However, the synonymous-site similarity in the mouse-rat comparison remains weak. Simulations suggest that this difference between the mouse-human and the mouse-rat comparisons is not artifactual and that there is therefore a difference between humans and rodents in the local patterns of mutation or selection on synonymous sites (conversely, we show that the previously reported absence of a local similarity in nonsynonymous rates of evolution in the human-rodent comparison was a methodological artifact). We show that linkage effects have a long-range component: not one in a million random genomes shows such levels of autosomal heterogeneity. The heterogeneity is so great that more autosomes than expected by chance have rates of synonymous evolution comparable with that of the X chromosome. As autosomal heterogeneity cannot be owing to different times spent in the germ line, this demonstrates that the dominant determiner of synonymous rates of evolution is not, as has been conjectured, the time spent in the male germ line.

Unlabelled: For the assessment of the efficacy of clinical gene therapy trials, different imaging modalities have been developed that enable a noninvasive assessment of location, magnitude, and duration of transduced gene expression in vivo. These imaging methods rely on a combination of an appropriate marker gene and a radiolabeled or paramagnetic marker substrate that can be detected by PET or MRI. Here, we assess whether the nucleoside analog 2'-fluoro-2'-deoxy-1beta-D-arabinofuranosyl-5-iodouracil (FIAU), a specific marker substrate for herpes simplex virus type 1 thymidine kinase (HSV-1-tk) gene expression, penetrates the blood-brain barrier (BBB) as an essential prerequisite for a noninvasive assessment of HSV-1-tk gene expression in gliomas., Methods: No-carrier-added [(124)I]FIAU was synthesized by reacting the precursor 2'-fluoro-2'-deoxy-1beta-D-arabinofuranosyluracil (FAU) with carrier-free [(124)I]NaI. The course of biodistribution of [(124)I]FIAU was investigated in anesthetized cats (n = 3; organs) and in one patient with a recurrent glioblastoma (plasma and brain) by PET imaging over several hours (cats, 1-22 h) to several days (patient, 1-68 h). FIAU PET was performed in conjunction with multitracer PET imaging (cerebral blood flow and cerebral metabolic rate of O(2) in cats only; cerebral metabolic rate of glucose and [(11)C]methionine in all subjects). A region-of-interest analysis was performed on the basis of coregistered high-resolution MR images. The average radioactivity concentration was determined, decay corrected, and recalculated as percentage injected dose per gram of tissue (%ID/g) or as standardized uptake values (SUVs)., Results: The average chemical yield of [(124)I]FIAU synthesis was 54.6% +/- 6.8%. The chemical and radiochemical purities of [(124)I]FIAU were found to be >98% and >95%, respectively. In cats, the kinetic analysis of [(124)I]FIAU-derived radioactivity showed an early peak (1-2 min after injection) in heart and kidneys (0.20 %ID/g; SUV, 4.0) followed by a second peak (10-20 min after injection) in liver and spleen (0.16 %ID/g; SUV, 3.2) with subsequent clearance from tissues and a late peak in the bladder (10-15 h after injection). In the unlesioned cat brain, no substantial [(124)I]FIAU uptake occurred throughout the measurement (<0.02 %ID/g; SUV, <0.4). In the patient, [(124)I]FIAU uptake in normal brain was also very low (<0.0002 %ID/g; SUV, <0.16). In contrast, the recurrent glioblastoma revealed relatively high levels of [(124)I]FIAU-derived radioactivity (5-10 min after injection; 0.001 %ID/g; SUV, 0.8), which cleared slowly over the 68-h imaging period., Conclusion: The PET marker substrate FIAU does not penetrate the intact BBB significantly and, hence, is not the marker substrate of choice for the noninvasive localization of HSV-1-tk gene expression in the central nervous system under conditions in which the BBB is likely to be intact. However, substantial levels of [(124)I]FIAU-derived radioactivity may occur within areas of BBB disruption (e.g., glioblastoma), which is an essential prerequisite for imaging clinically relevant levels of HSV-1-tk gene expression in brain tumors after gene therapy by FIAU PET. For this purpose, washout of nonspecific radioactivity should be allowed for several days.

Modern multiring positron emission tomographs allow the acquisition of 3-D data sets to increase their sensitivity. A substantial part of this data is due to scattered radiation. The authors describe the experimental dependence of point source scatter distributions on energy window setting, source location, and scatter volume in geometries relevant for brain studies. The point source scatter distribution was parametrized accurately by a broad, 2-D Gaussian, which included a shift parameter to account for asymmetry of the scatter medium relative to the source. This parametrization was used to formulate two fast scatter correction algorithms suitable for brain scans. In both algorithms, a 2-D subset of the measured projections was transformed into a scatter projection. An image of the 3-D scatter distribution was reconstructed using 2-D algorithms. It was then subtracted from the total (true+scattered) 3-D image. Both algorithms were implemented in different combinations with the additional attenuation correction and were tested on point source and phantom measurements. It was shown that, for the situation typical for brain scans, reconstructed scatter fractions could be reduced to 5% or less.