Phylogenomic analysis of gene co‐expression networks reveals the evolution of functional modules

Molecular evolutionary studies correlate genomic and phylogenetic information with the emergence of new traits of organisms. These traits are, however, the consequence of dynamic gene networks composed of functional modules, which might not be captured by genomic analyses. Here, we established a method that combines large‐scale genomic and phylogenetic data with gene co‐expression networks to extensively study the evolutionary make‐up of modules in the moss Physcomitrella patens, and in the angiosperms Arabidopsis thaliana and Oryza sativa (rice). We first show that younger genes are less annotated than older genes. By mapping genomic data onto the co‐expression networks, we found that genes from the same evolutionary period tend to be connected, whereas old and young genes tend to be disconnected. Consequently, the analysis revealed modules that emerged at a specific time in plant evolution. To uncover the evolutionary relationships of the modules that are conserved across the plant kingdom, we added phylogenetic information that revealed duplication and speciation events on the module level. This combined analysis revealed an independent duplication of cell wall modules in bryophytes and angiosperms, suggesting a parallel evolution of cell wall pathways in land plants. We provide an online tool allowing plant researchers to perform these analyses at http://www.gene2function.de .     

The evolutionary analysis reveals domain fusion of proteins with Frizzled-like CRD domain

Frizzleds (FZDs) are transmembrane receptors in the Wnt signaling pathway and they play pivotal roles in developments. The Frizzled-like extracellular Cysteine-rich domain (Fz-CRD) has been identified in FZDs and other proteins. The origin and evolution of these proteins with Fz-CRD is the main interest of this study. We found that the Fz-CRD exists in FZD, SFRP, RTK, MFRP, CPZ, CORIN, COL18A1 and other proteins. Our systematic analysis revealed that the Fz-CRD domain might have originated in protists and then fused with the Frizzled-like seven-transmembrane domain (7TM) to form the FZD receptors, which duplicated and diversified into about 11 members in Vertebrates. The SFRPs and RTKs with the Fz-CRD were found in sponge and expanded in Vertebrates. Other proteins with Fz-CRD may have emerged during Vertebrate evolution through domain fusion. Moreover, we found a glycosylation site and several conserved motifs in FZDs, which may be related to Wnt interaction. Based on these results, we proposed a model showing that the domain fusion and expansion of Fz-CRD genes occurred in Metazoa and Vertebrates. Our study may help to pave the way for further research on the conservation and diversification of Wnt signaling functions during evolution.     

基于全基因组系统发生的草菇基因家族显著扩增

【目的】从基因组层次研究草菇低温自溶异常代谢的分子特征。【方法】对21个真菌物种进行全基因组系统发生分析,进而选取其中具有代表性的物种进行比较基因组学分析,系统研究草菇异常代谢分子特征。【结果】全基因组系统发生分析结果显示草菇位于草腐菌所形成簇的底端。基于全基因组系统发生树,由于担子菌和子囊菌属于完全不同的演化路径,所以选取担子菌中具有代表性的9个物种进行比较基因组学分析,结果显示相比于其它草腐菌,草菇基因家族具有一定的收缩趋势。进一步对不同范畴的基因家族数目进行比较,结果显示3个大于200的草菇基因家族(fam1、fam4和fam6)分别发生了显著扩增,且在总数上也显著高于其它物种,表明草菇的这一分子特征与其异常代谢相关。【结论】3个草菇基因家族(200)显著扩增提示其特定基因家族功能加强,很可能与草菇低温自溶密切相关。     

Robertsonian fusion and centric fission in karyotype evolution of higher plants

Robertsonian fusion and centric fission are uniquely detectable in comparative studies of karyotype patterns. They are the most important types of karyotype change in animals but seem to be relatively uncommon in higher plants. Both modify intra- and interchromosomal recombination and linkage relationships and consequently patterns of genetic variation. When differentiating populations or species they can produce postulating barriers to gene flow. The number of reported cases of fusion or fission in higher plants has increased over the years but remains low, and most of these are casual comparisons of karyotypes without any follow-up investigation. This review focuses on more adequate studies made in a few groups. Studies in the Tradescantieae produce the strongest evidence for fusion as a type of orthoselection in the subfamily. Some species ofLycoris are also considered to have evolved their karyotypes in that way. Some genera of slipper orchids and the cycad genusZamia have populations where atypical chromosome number increase can be attributed to fission probably as a result of stressful influences. It is suggested that fusion may have been involved in the evolution of many stable karyotypes and that fission is generally a secondary destabilizing mechanism which may lead to refusion in the long term. Their proven incidence remains making it unwise to suggest that they have been major influences in karyotype evolution in higher plants.     

Rapid tagging and integration of genes in Giardia intestinalis

We developed a series of plasmids that allow C-terminal tagging of any gene in its endogenous locus in Giardia intestinalis, with different epitope tags (triple hemagglutinin [3HA] and triple Myc [3Myc]) and selection markers (puromycin, neomycin, and a newly developed marker, blasticidin). Using these vectors, cyclin B and aurora kinase were tagged, expressed, and localized.     

Phylogenomic analysis resolves the relationships among net‐winged beetles (Coleoptera: Lycidae) and reveals the parallel evolution of morphological traits

Net‐winged beetles (Coleoptera: Lycidae) are a diverse group of elateroids known for aposematism and neoteny. Phylogenetic analyses of morphological and molecular data have revealed different results with respect to within‐group relationships. In this study, we recovered a highly supported phylogenomic phylogeny and identified seven subfamilies: Dexorinae stat.n. , Calochrominae stat.n. , Erotinae, Ateliinae, Lycinae, Lyropaeinae stat.n. and Metriorrhynchinae stat.n. Our results suggest that female neoteny evolved multiple times. Therefore, the development of similar morphological modifications in neotenics may be linked and may have produced characteristics such as body miniaturization, structural simplification, i.e. reduction of mouthparts, fewer antennomeres and palpomeres, uniquely shaped terminal palpomeres, shortened elytra, the loss of coadaptation between the elytra and pronotum, and others. Additional traits evolved in parallel due to similarities in biology, function and sexual selection. These characteristics include mimetic similarities, the presence of the rostrum, pronotal carinae and elytral costae, and the structure of male genitalia. By comparing the phylogenomic topology with the evolution of morphological characters, we were able to identify evolutionary trends in lycids and compare them with analogues for other neotenic elateroids. These traits have not been accepted as homoplasies due to the ambiguous phylogenetic signal from Sanger sequencing markers.     

Molecular analysis of some camel cytochrome P450 enzymes reveals lower evolution and drug-binding properties

Camels bear unique genotypes and phenotypes for adaptation of their harsh environment. They have unique visual systems, sniffing, water metabolism, and heat-control mechanisms that are different from other creatures. The recent announcement for the complete sequence of camel genome will allow for the discovery of many secrets of camel life. In this context, the genetic bases of camel drug-metabolizing enzymes are still unknown. Furthermore, the genomic content of camel that rendered it highly susceptible to some drugs (as monensin and salinomycin) and became easily intoxicated needs to be investigated. The objectives of this work are the annotation of camel genome and retrieval of camel for cytochrome P450 (CYP) 1A1, 2C, and 3A enzymes. This is followed by comprehensive phylogenetic, evolution, molecular modeling, and docking studies. In comparison with the human enzymes, camel CYPs showed lower evolution rate, especially CYP1A1. Furthermore, the binding of monensin, salinomycin, alfa-naphthoflavone, felodepine, and ritonavir was weaker in camel enzymes. Interestingly, rerank score indicated instable binding of monensin and salinomycin with camel CYP1A1 as well as salinomycin with camel CYP2C. The results of this work suggest that camels are more susceptible to toxicity with compounds undergoing metabolic oxidation. This conclusion was based on lower evolution rate and lower binding potency of camels compared with the human enzymes.     

Phylogenomic analysis of pseudocryptic diversity reveals the new genus Deltalsia (Rhodomelaceae,Rhodophyta)

Molecular analyses, in combination with morphological studies, provide invaluable tools for delineating red algal taxa. However, molecular datasets are incomplete and taxonomic revisions are often required once additional species or populations are sequenced. The small red alga Conferva parasitica was described from the British Isles in 1762 and then reported from other parts of Europe. Conferva parasitica was traditionally included in the genus Pterosiphonia (type species P. cloiophylla in Schmitz and Falkenberg 1897), based on its morphological characters, and later transferred to Symphyocladia and finally to Symphyocladiella using molecular data from an Iberian specimen. However, although morphological differences have been observed between specimens of Symphyocladiella parasitica from northern and southern Europe they have yet to be investigated in a phylogenetic context. In this study, we collected specimens from both regions, studied their morphology and analyzed rbcL and cox1 DNA sequences. We determined the phylogenetic position of a British specimen using a phylogenomic approach based on mitochondrial and plastid genomes. Northern and southern European populations attributed to S. parasitica represent different species. Symphyocladiella arecina sp. nov. is proposed for specimens from southern Europe, but British specimens were resolved as a distant sister lineage to the morphologically distinctive Amplisiphonia, so we propose the new genus Deltalsia for this species. Our study highlights the relevance of using materials collected close to the type localities for taxonomic reassessments, and showcases the utility of genome-based phylogenies for resolving classification issues in the red algae.     

Changes in allozyme pattern of the protozoan parasite Giardia intestinalis.

The present study compares the allelic profiles of Giardia intestinalis grown in vivo and in vitro. Three clinical isolates of G. intestinalis were established in suckling mice and subsequently adapted to in vitro culture to test the null hypothesis that samples of the same clinical isolate grown in different culture conditions have identical allelic profiles. For each isolate, a mouse-derived and an axenically cultured sample were analysed electrophoretically at 11 enzyme loci. In each case, the axenically cultured sample of each isolate showed marked allelic differences from its corresponding in vivo sample. These data suggest that there may be either regulated expression of alternative genes encoding distinct isozymes (i.e. gene switching) or selection by different growth conditions of specific genotypes from a mixture present within the original clinical isolate. Although these hypotheses are not tested in this study, the data highlight the importance of confirming that allozymes (or isozymes) are stable genetic characters for the identification and characterization of protozoan taxa.     

Photocatalytic disinfection of Giardia intestinalis and Acanthamoeba castellani cysts in water

In this study, disinfection of water containing Giardia intestinalis and Acanthamoeba castellani cysts with TiO2 and modified catalyst silver loaded TiO2 (Ag-TiO2) was investigated. Destruction of the parasites was evaluated after UV illumination of the suspension consisting 5 x 10(8)-13.5 x 10(8)cysts/mL in the presence of 2g/L neat or modified TiO2 at neutral pH. In the initial stage, the solid photocatalyst particles penetrated the cyst wall and then oxidant species produced by TiO2/UV destroyed both cell wall and intracellular structure. In the case of G. intestinalis inactivation (disinfection) performance of TiO2/UV system reached 52.5% only after 25 min illumination and total parasite disinfection was achieved after 30 min illumination. However, silver loaded TiO2 seemed to be more effective as this loading provided better catalytic action as well as additional antimicrobial properties. Cell viability tests showed that parasite cysts, their walls in particular, were irreversibly damaged and cysts did not re-grow. Nevertheless the studied system seemed to be ineffective for the inactivation of A. castellani. Inactivation percentages of TiO2/UV and Ag-TiO2/UV systems were far lower than that of UV alone, being 50.1% and 46.1%, respectively.     

Novel structural components of the ventral disc and lateral crest in Giardia intestinalis

Giardia intestinalis is a ubiquitous parasitic protist that is the causative agent of giardiasis, one of the most common protozoan diarrheal diseases in the world. Giardia trophozoites attach to the intestinal epithelium using a specialized and elaborate microtubule structure, the ventral disc. Surrounding the ventral disc is a less characterized putatively contractile structure, the lateral crest, which forms a continuous perimeter seal with the substrate. A better understanding of ventral disc and lateral crest structure, conformational dynamics, and biogenesis is critical for understanding the mechanism of giardial attachment to the host. To determine the components comprising the ventral disc and lateral crest, we used shotgun proteomics to identify proteins in a preparation of isolated ventral discs. Candidate disc-associated proteins, or DAPs, were GFP-tagged using a ligation-independent high-throughput cloning method. Based on disc localization, we identified eighteen novel DAPs, which more than doubles the number of known disc-associated proteins. Ten of the novel DAPs are associated with the lateral crest or outer edge of the disc, and are the first confirmed components of this structure. Using Fluorescence Recovery After Photobleaching (FRAP) with representative novel DAP::GFP strains we found that the newly identified DAPs tested did not recover after photobleaching and are therefore structural components of the ventral disc or lateral crest. Functional analyses of the novel DAPs will be central toward understanding the mechanism of ventral disc-mediated attachment and the mechanism of disc biogenesis during cell division. Since attachment of Giardia to the intestine via the ventral disc is essential for pathogenesis, it is possible that some proteins comprising the disc could be potential drug targets if their loss or disruption interfered with disc biogenesis or function, preventing attachment.     

Common Coinfections of Giardia intestinalis and Helicobacter pylori in Non-Symptomatic Ugandan Children

BackgroundThe protozoan parasite Giardia intestinalis and the pathogenic bacterium Helicobacter pylori are well known for their high prevalences in human hosts worldwide. The prevalence of both organisms is known to peak in densely populated, low resource settings and children are infected early in life. Different Giardia genotypes/assemblages have been associated with different symptoms and H. pylori with induction of cancer. Despite this, not much data are available from sub-Saharan Africa with regards to the prevalence of different G. intestinalis assemblages and their potential association with H. pylori infections.Conclusions/SignificanceThis study shows that Giardia assemblage B dominates in children in Kampala, Uganda and that the presence of H. pylori is an associated risk factor for G. intestinalis infection.     

Kinetic and physical characterization of the inducible UDP-N-acetylglucosamine pyrophosphorylase from Giardia intestinalis

The UDP-N-acetylglucosamine pyrophosphorylase in Giardia intestinalis (GiUAP) is one of the five inducible enzymes to synthesize UDP-GalNAc, which is an important precursor for cyst wall synthesis. The recombinant UDP-N-acetylglucosamine pyrophosphorylase (rGiUAP) and its mutants G108A and G210A were expressed and identified by SDS-PAGE, size-exclusion chromatography, Western hybridization, and MALDI mass spectrometry. Sequence comparison with other eukaryotic UAPs has identified three specific motifs. Within these motifs alanine substitution for Gly(108) or Gly(210) dramatically reduced the pyrophosphate synthesis, suggesting these amino acids are catalytic residues. Besides, the rGiUAP was found to have relaxed binding to other uridine-based nucleotides, suggesting the substrate binding pocket is specific to uridine rather than phosphate group(s). Moreover, thermal denaturation analysis showed a significant increase in T(m) for the rGiUAP and G108A upon binding of the substrate Mg-UTP. In contrast, G210A showed a decreased T(m) upon binding of Mg-UTP. These results showed that binding of Mg-UTP increases protein stability of the rGiUAP, and the catalytic residue Gly(210) plays a significant role in stabilizing the protein structure. Such stabilization effect induced by substrate binding might be physiologically important as it favors the production of UDP-GlcNAc and hence the downstream GalNAc, which is crucial to survival of Giardia. These results help to define the essential amino acids for catalysis in the GiUAP and reveal the role of Mg-UTP binding in regulation of protein stability.     

Comparative analysis of SET domain proteins in maize and Arabidopsis reveals multiple duplications preceding the divergence of monocots and dicots

Histone proteins play a central role in chromatin packaging, and modification of histones is associated with chromatin accessibility. SET domain [Su(var)3-9, Enhancer-of-zeste, Trithorax] proteins are one class of proteins that have been implicated in regulating gene expression through histone methylation. The relationships of 22 SET domain proteins from maize (Zea mays) and 32 SET domain proteins from Arabidopsis were evaluated by phylogenetic analysis and domain organization. Our analysis reveals five classes of SET domain proteins in plants that can be further divided into 19 orthology groups. In some cases, such as the Enhancer of zeste-like and trithorax-like proteins, plants and animals contain homologous proteins with a similar organization of domains outside of the SET domain. However, a majority of plant SET domain proteins do not have an animal homolog with similar domain organization, suggesting that plants have unique mechanisms to establish and maintain chromatin states. Although the domains present in plant and animal SET domain proteins often differ, the domains found in the plant proteins have been generally implicated in protein-protein interactions, indicating that most SET domain proteins operate in complexes. Combined analysis of the maize and Arabidopsis SET domain proteins reveals that duplication of SET domain proteins in plants is extensive and has occurred via multiple mechanisms that preceded the divergence of monocots and dicots.     

The minimal proteome in the reduced mitochondrion of the parasitic protist Giardia intestinalis

The mitosomes of Giardia intestinalis are thought to be mitochondria highly-reduced in response to the oxygen-poor niche. We performed a quantitative proteomic assessment of Giardia mitosomes to increase understanding of the function and evolutionary origin of these enigmatic organelles. Mitosome-enriched fractions were obtained from cell homogenate using Optiprep gradient centrifugation. To distinguish mitosomal proteins from contamination, we used a quantitative shot-gun strategy based on isobaric tagging of peptides with iTRAQ and tandem mass spectrometry. Altogether, 638 proteins were identified in mitosome-enriched fractions. Of these, 139 proteins had iTRAQ ratio similar to that of the six known mitosomal markers. Proteins were selected for expression in Giardia to verify their cellular localizations and the mitosomal localization of 20 proteins was confirmed. These proteins include nine components of the FeS cluster assembly machinery, a novel diflavo-protein with NADPH reductase activity, a novel VAMP-associated protein, and a key component of the outer membrane protein translocase. None of the novel mitosomal proteins was predicted by previous genome analyses. The small proteome of the Giardia mitosome reflects the reduction in mitochondrial metabolism, which is limited to the FeS cluster assembly pathway, and a simplicity in the protein import pathway required for organelle biogenesis.     

Phylogenomic assessment of the role of hybridization and introgression in trait evolution

Trait evolution among a set of species—a central theme in evolutionary biology—has long been understood and analyzed with respect to a species tree. However, the field of phylogenomics, which has been propelled by advances in sequencing technologies, has ushered in the era of species/gene tree incongruence and, consequently, a more nuanced understanding of trait evolution. For a trait whose states are incongruent with the branching patterns in the species tree, the same state could have arisen independently in different species (homoplasy) or followed the branching patterns of gene trees, incongruent with the species tree (hemiplasy). Another evolutionary process whose extent and significance are better revealed by phylogenomic studies is gene flow between different species. In this work, we present a phylogenomic method for assessing the role of hybridization and introgression in the evolution of polymorphic or monomorphic binary traits. We apply the method to simulated evolutionary scenarios to demonstrate the interplay between the parameters of the evolutionary history and the role of introgression in a binary trait’s evolution (which we call xenoplasy). Very importantly, we demonstrate, including on a biological data set, that inferring a species tree and using it for trait evolution analysis in the presence of gene flow could lead to misleading hypotheses about trait evolution.     

Phylogenomic analysis of vertebrate thrombospondins reveals fish-specific paralogues, ancestral gene relationships and a tetrapod innovation

Background  

Thrombospondins (TSPs) are evolutionarily-conserved, extracellular, calcium-binding glycoproteins with important roles in cell-extracellular matrix interactions, angiogenesis, synaptogenesis and connective tissue organisation. Five TSPs, designated TSP-1 through TSP-5, are encoded in the human genome. All but one have known roles in acquired or inherited human diseases. To further understand the roles of TSPs in human physiology and pathology, it would be advantageous to extend the repertoire of relevant vertebrate models. In general the zebrafish is proving an excellent model organism for vertebrate biology, therefore we set out to evaluate the status of TSPs in zebrafish and two species of pufferfish.