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Sequence and structural aspects of functional diversification in class I alpha-mannosidase evolution.

MOTIVATION: Class I alpha-mannosidases comprise a homologous and functionally diverse family of glycoside hydrolases. Phylogenetic analysis based on an amino acid sequence alignment of the catalytic domain of class I alpha-mannosidases reveals four well-supported phylogenetic groups within this family. These groups include a number of paralogous members generated by gene duplications that occurred as far back as the initial divergence of the crown-group of eukaryotes. Three of the four phylogenetic groups consist of enzymes that have group-specific biochemical specificity and/or sites of activity. An attempt has been made to uncover the role that natural selection played in the sequence and structural divergence between the phylogenetically and functionally distinct Endoplasmic Reticulum (ER) and Golgi apparatus groups. RESULTS: Comparison of site-specific amino acid variability profiles for the ER and Golgi groups revealed statistically significant evidence for functional diversification at the sequence level and indicated a number of residues that are most likely to have played a role in the functional divergence between the two groups. The majority of these sites appear to contain residues that have been fixed within one organelle-specific group by positive selection. Somewhat surprisingly these selected residues map to the periphery of the alpha-mannosidase catalytic domain tertiary structure. Changes in these peripherally located residues would not seem to have a gross effect on protein function. Thus diversifying selection between the two groups may have acted in a gradual manner consistent with the Darwinian model of natural selection. CONTACT: bishogr@millsaps.edu.

Amino Acid Sequence↗

T lymphocyte surface antigens in primates.

We examined the binding of peripheral blood mononuclear cells from various primate species to a panel of monoclonal antibodies with specificity for human T lymphocytes. A remarkable structural conservation of the erythrocyte rosette receptor on T cells between such phylogenetically distant primates as lemurs and man was found. Furthermore, it was demonstrated that the degree of sharing of other T cell-specific surface determinants between man and other primates reflects the phylogenetic distance between them. Moreover, these studies suggest the potential use of monoclonal antibodies for further structural and functional elucidation of T lymphocyte-specific surface structures.

Animals↗

Molecular identification of a novel carnitine transporter specific to human testis. Insights into the mechanism of carnitine recognition.

l-Carnitine is an essential component of mitochondrial fatty acid beta-oxidation and plays a pivotal role in the maturation of spermatozoa within the male reproductive tract. Epididymal plasma contains the highest levels of l-carnitine found in the human body, and initiation of sperm motility occurs in parallel to l-carnitine increase in the epididymal lumen. Using a specific carrier, epididymal epithelium secretes l-carnitine into the lumen by an active transport mechanism; however, the structure-activity relationship comprising the carnitine-permeation pathway is poorly understood. We discovered a novel carnitine transporter (CT2) specifically located in human testis. Analyzing the primary structure of CT2 revealed that it is phylogenetically located between the organic cation transporter (OCT/OCTN) and anion transporter (OAT) families. Hence, CT2 represents a novel transporter family. When expressed in Xenopus oocytes, CT2 mediates the high affinity transport of l-carnitine but does not accept mainstream OCT/OCTN cationic or OAT anionic substrates. We synthesized and tested various carnitine-related compounds and investigated the physicochemical properties of substrate recognition by semi-empirical computational chemistry. The data suggest that the quaternary ammonium cation bulkiness and relative hydrophobicity be the most important factors that trigger CT2-substrate interactions. Immunohistochemistry showed that the CT2 protein is located in the luminal membrane of epididymal epithelium and within the Sertoli cells of the testis. The identification of CT2 represents an interesting evolutionary link between OCT/OCTNs and OATs, as well as provides us with an important insight into the maturation of human spermatozoa.

Amino Acid Sequence↗

Properties of phylogenetic trees generated by Yule-type speciation models.

We investigate some discrete structural properties of evolutionary trees generated under simple null models of speciation, such as the Yule model. These models have been used as priors in Bayesian approaches to phylogenetic analysis, and also to test hypotheses concerning the speciation process. In this paper we describe new results for three properties of trees generated under such models. Firstly, for a rooted tree generated by the Yule model we describe the probability distribution on the depth (number of edges from the root) of the most recent common ancestor of a random subset of k species. Next we show that, for trees generated under the Yule model, the approximate position of the root can be estimated from the associated unrooted tree, even for trees with a large number of leaves. Finally, we analyse a biologically motivated extension of the Yule model and describe its distribution on tree shapes when speciation occurs in rapid bursts.

Animals↗

The WRKY family of transcription factors in rice and Arabidopsis and their origins.

WRKY transcription factors, originally isolated from plants contain one or two conserved WRKY domains, about 60 amino acid residues with the WRKYGQK sequence followed by a C2H2 or C2HC zinc finger motif. Evidence is accumulating to suggest that the WRKY proteins play significant roles in responses to biotic and abiotic stresses, and in development. In this research, we identified 102 putative WRKY genes from the rice genome and compared them with those from Arabidopsis. The WRKY genes from rice and Arabidopsis were divided into three groups with several subgroups on the basis of phylogenies and the basic structure of the WRKY domains (WDs). The phylogenetic trees generated from the WDs and the genes indicate that the WRKY gene family arose during evolution through duplication and that the dramatic amplification of rice WRKY genes in group III is due to tandem and segmental gene duplication compared with those of Arabidopsis. The result suggests that some of the rice WRKY genes in group III are evolutionarily more active than those in Arabidopsis, and may have specific roles in monocotyledonous plants. Further, it was possible to identify the presence of WRKY-like genes in protists (Giardia lamblia and Dictyostelium discoideum) and green algae Chlamydomonas reinhardtii through database research, demonstrating the ancient origin of the gene family. The results obtained by alignments of the WDs from different species and other analysis imply that domain gain and loss is a divergent force for expansion of the WRKY gene family, and that a rapid amplification of the WRKY genes predate the divergence of monocots and dicots. On the basis of these results, we believe that genes encoding a single WD may have been derived from the C-terminal WD of the genes harboring two WDs. The conserved intron splicing positions in the WDs of higher plants offer clues about WRKY gene evolution, annotation, and classification.

Amino Acid Motifs↗

Molecular cloning and tissue-specific transcriptional regulation of the first peroxidase family member, Udp1, in stinging nettle (Urtica dioica).

A full-length cDNA clone, designated Udp1, was isolated from Urtica dioica (stinging nettle), using a polymerase chain reaction based strategy. The putative Udp1 protein is characterized by a cleavable N-terminal signal sequence, likely responsible for the rough endoplasmic reticulum entry and a 310 amino acids mature protein, containing all the important residues, which are evolutionary conserved among different members of the plant peroxidase family. A unique structural feature of the Udp1 peroxidase is defined into the short carboxyl-terminal extension, which could be associated with the vacuolar targeting process. Udp1 peroxidase is differentially regulated at the transcriptional level and is specifically expressed in the roots. Interestingly, wounding and ultraviolet radiation stress cause an ectopic induction of the Udp1 gene expression in the aerial parts of the plant. A genomic DNA fragment encoding the Udp1 peroxidase was also cloned and fully sequenced, revealing a structural organization of three exons and two introns. The phylogenetic relationships of the Udp1 protein to the Arabidopsis thaliana peroxidase family members were also examined and, in combination with the homology modelling approach, dictated the presence of distinct structural elements, which could be specifically involved in the determination of substrate recognition and subcellular localization of the Udp1 peroxidase.

Amino Acid Sequence↗

A novel zinc finger structure in the large subunit of human general transcription factor TFIIE.

The zinc finger domain in the large subunit of TFIIE (TFIIEalpha) is phylogenetically conserved and is essential for transcription. Here, we determined the solution structure of this domain by using NMR. It consisted of one alpha-helix and five beta-strands, showing novel features distinct from previously determined zinc-binding structures. We created point mutants of TFIIEalpha in this domain and examined their binding abilities to other general transcription factors as well as their transcription activities. Four Zn(2+)-ligand mutants, in which each of cysteine residues at positions 129, 132, 154, and 157 was replaced by alanine, possessed no transcription activities on a linearized template, whereas, on a supercoiled template, interesting functional asymmetry was observed: although the C-terminal two mutants abolished transcription activity (<5%), the N-terminal two mutants retained about 20% activities. The N-terminal two mutants bound stronger to the small subunit of TFIIF than the wild type and the C-terminal two mutants were impaired in their binding abilities to the XPB subunits of TFIIH. These suggest that the structural integrity of the zinc finger domain is essential for the TFIIE function, particularly in the transition from the transcription initiation to elongation and the conformational tuning of this domain for appropriate positioning of TFIIF, TFIIH, and polymerase II would be needed depending on the situation and timing.

Alanine↗

A conserved secondary structure for telomerase RNA.

The RNA moiety of the ribonucleoprotein enzyme telomerase contains the template for telomeric DNA synthesis. We present a secondary structure model for telomerase RNA, derived by a phylogenetic comparative analysis of telomerase RNAs from seven tetrahymenine ciliates. The telomerase RNA genes from Tetrahymena malaccensis, T. pyriformis, T. hyperangularis, T. pigmentosa, T. hegewishii, and Glaucoma chattoni were cloned, sequenced, and compared with the previously cloned RNA gene from T. thermophila and with each other. To define secondary structures of these RNAs, homologous complementary sequences were identified by the occurrence of covariation among putative base pairs. Although their primary sequences have diverged rapidly overall, a strikingly conserved secondary structure was identified for all these telomerase RNAs. Short regions of nucleotide conservation include a block of 22 totally conserved nucleotides that contains the telomeric templating region.

Animals↗

Phylogenetic distribution of extraintestinal virulence-associated traits in Escherichia coli.

The 72 member strains of the Escherichia coli Reference collection were assessed as to genotype for 31 putative extraintestinal virulence factor (VF) genes and DNA sequence for papA, the P fimbrial structural subunit gene. Although most VFs were concentrated in phylogenetic group B2 or jointly in groups B2 and D, others were concentrated primarily in group D, were broadly distributed (without group-specific associations), and/or occurred only outside of group B2. Statistical correlations among VFs suggested linkage on pathogenicity-associated islands or plasmids. Isolates from humans and nonhuman primates had more VFs than did isolates from other animals. Sequence diversity was minimal within each F type-specific papA allele group but was substantial among different papA allele groups. The distribution patterns of papA variants and other VFs suggested multiple horizontal transfer events. These findings provide new insights into the phylogenetic origins of extraintestinal VFs in E. coli.

Animals↗

Phylogenetic analysis of the cadherin superfamily.

Cadherins are a multigene family of proteins which mediate homophilic calcium-dependent cell adhesion and are thought to play an important role in morphogenesis by mediating specific intercellular adhesion. Different lines of experimental evidence have recently indicated that the site responsible for mediating adhesive interactions is localized to the first extracellular domain of cadherin. Based upon an analysis of the sequence of this domain, I show that cadherins can be classified into three groups with distinct structural features. Furthermore, using this sequence information a phylogenetic tree relating the known cadherins was assembled. This is the first such tree to be published for the cadherins. One cadherin subtype, neural cadherin (N-cadherin), shows very little sequence divergence between species, whereas all other cadherin subtypes show more substantial divergence, suggesting that selective pressure upon this domain may be greater for N-cadherin than for other cadherins. Phylogenetic analysis also suggests that the gene duplications which established the main branches leading to the different cadherin subtypes occurred very early in their history. These duplications set the stage for the diversified superfamily we now observe.

Amino Acid Sequence↗

Influence of phylogeny on posttranscriptional modification of rRNA in thermophilic prokaryotes: the complete modification map of 16S rRNA of Thermus thermophilus.

Posttranscriptional modification in RNA generally serves to fine-tune and regulate RNA structure and, in many cases, is relatively conserved and phylogenetically distinct. We report the complete modification map for SSU rRNA from Thermus thermophilus, determined primarily by HPLC/electrospray ionization MS-based methods. Thermus modification levels are significantly lower, and structures at the nucleoside level are very different from those of the archaeal thermophile Sulfolobus solfataricus growing in the same temperature range [Noon, K. R., et al. (1998) J. Bacteriol. 180, 2883-2888]. The Thermus modification map is unexpectedly similar to that of Escherichia coli (11 modified sites), with which it shares identity in 8 of the 14 modifications. Unlike the heavily methylated Sulfolobus SSU RNA, Thermus contains a single ribose-methylated residue, N(4),2'-O-dimethylcytidine-1402, suggesting that O-2'-ribose methylation in this bacterial thermophile plays a reduced role in thermostabilization compared with the thermophilic archaea. Adjacent pseudouridine residues were found in the single-stranded 3' tail of Thermus 16S rRNA at residues 1540 and 1541 (E. coli numbering) in the anti-Shine-Dalgarno mRNA binding sequence. The present results provide an example of the potential of LC/MS for extensive modification mapping in large RNAs.

Base Sequence↗

Sequences related to the ox pancreatic ribonuclease coding region in the genomic DNA of mammalian species.

Mammalian pancreatic ribonucleases form a family of homologous proteins that has been extensively investigated. The primary structures of these enzymes were used to derive phylogenetic trees. These analyses indicate that the presence of three strictly homologous enzymes in the bovine species (the pancreatic, seminal, and cerebral ribonucleases) is due to gene duplication events which occurred during the evolution of ancestral ruminants. In this paper we present evidence that confirms this finding and that suggests an overall structural conservation of the putative ribonuclease genes in ruminant species. We could also demonstrate that the sequences related to ox ribonuclease coding regions present in genomic DNA of the giraffe species are the orthologues of the bovine genes encoding the three ribonucleases mentioned above.

Amino Acid Sequence↗

Sebacinales form ectendomycorrhizas with Cavendishia nobilis, a member of the Andean clade of Ericaceae, in the mountain rain forest of southern Ecuador.

Cavendishia nobilis var. capitata is an endemic member of the Ericaceae growing as a hemiepiphyte in the tropical mountain rain forest of southern Ecuador. Mycorrhizas were collected from 20 individuals along an altitudinal gradient between 1850 and 2300 m. Transmission electron microscopy was used to study the symbiotic association in detail, and phylogenetic analyses based on nuclear rDNA coding for the ribosomal large subunit (nucLSU) were carried out to identify the associated mycorrhizal fungi. Microscopic and ultrastructural investigations showed the formation of a hyphal sheath, intercellular penetration of fine hyphae and colonization of the cortical cells by swollen hyphae of the same fungus. These structures were formed by hymenomycetes and ascomycetes. Molecular phylogenetic analysis detected seven groups of mycorrhizal fungi belonging to the Sebacinales. This is the first study to obtain evidence of ectendomycorrhizas in the Vaccinioideae. The ascomycetous nucLSU sequences belonged to members of the Leotiomycetes. The ectendomycorrhiza of C. nobilis with Sebacinales is discussed as a specific, hitherto undescribed mycorrhizal subcategory of ectomycorrhizas. We propose the term 'cavendishioid mycorrhiza'. This subcategory is most likely specific for the Andean clade of Ericaceae.

Ecosystem↗

The Polyvagal Theory: phylogenetic contributions to social behavior.

The scientific legacy of Paul MacLean provides important insights into the neural substrate of adaptive social behavior in mammals. Through his research and visionary conceptualizations, current investigators can legitimately study social behavior from a neurobiological perspective. His research and writings provided three important contributions. First, he emphasized the importance of evolution as an organizing principle that shaped both the structure of the nervous system and the adaptive social behavior. Second, by defining the limbic system, he legitimized the biological perspective in the study of emotion. Third, he recognized the important role of the vagal afferents in the regulation of higher brain structures. The paper will focus on the Polyvagal Theory. The Polyvagal Theory is a new conceptualization of the role of vagus and employs several features that MacLean emphasized including the importance of evolution, limbic structures and vagal afferents. The Polyvagal Theory builds on these early findings by MacLean and focuses on the link between phylogenetic changes in the autonomic nervous system and social behavior. By focusing on the phylogenetic changes in the structure of the vagus and the role that the vagus plays in the neural regulation of visceral state, new insights regarding social behavior emerge. Moreover, by articulating the phylogenetically organized hierarchy of neural circuits, insights into benefits of social behavior become evident as do an understanding of the behavioral and physiological features associated with stress and psychiatric disorders.

Animals↗

Modelling the secondary structures of slippage-prone hypervariable RNA regions: the example of the tiger beetle 18S rRNA variable region V4.

Variable regions within ribosomal RNAs frequently vary in length as a result of incorporating products of slippage. This makes constructing secondary structure models problematic because base homology is difficult or impossible to establish between species. Here, we model such a region by comparing the results of the MFOLD suboptimal folding algorithm for different species to identify conserved structures. Based on the reconstruction of base change on a phylogenetic tree of the species and comparison against null models of character change, we devise a statistical analysis to assess support of these structures from compensatory and semi-compensatory (i.e. G.C to G.U or A.U to G.U) mutations. As a model system we have used variable region V4 from cicindelid (tiger beetle) small subunit ribosomal RNAs (SSU rRNAs). This consists of a mixture of conserved and highly variable subregions and has been subject to extensive comparative analysis in the past. The model that results is similar to a previously described model of this variable region derived from a different set of species and contains a novel structure in the central, highly variable part. The method we describe may be useful in modelling other RNA regions that are subject to slippage.

Animals↗

Secondary and tertiary structure in the central domain of adenovirus type 2 VA RNA I.

The small (160 nt) adenovirus RNA, VA RNAI, antagonizes the activation of the cellular protein kinase PKR (also known as DAI), a key regulator of gene expression. VA RNA consists of two stems separated by a complex region, the central domain, that is essential for its function. A notable feature of the central domain is a pair of tetranucleotides, GGGU and ACCC, which are mutually complementary and phylogenetically conserved. To investigate their role in the structure and function of VA RNA, we generated three sets of mutations designed to disrupt the putative stem and to restore it with different nucleotides. Substitutions in either of the tetranucleotides abrogated VA RNA function in two independent PKR-based assays, demonstrating the importance of these sequences in vivo. Compensating mutants restored function, indicating that the two tetranucleotides pair in the cell, but all of the compensating mutants were less active than wild-type VA RNA. The effects of the mutations on RNA structure were probed by nuclease sensitivity analysis. Pronounced changes in two loops in the central domain correlated closely with the formation and disruption of the stem, suggesting that the tetranucleotide stem defines a critical element in the structure of the central domain through tertiary interactions with the two loops. A model for the central domain is presented that accommodates these findings and also accounts for the known sites of PKR interaction.

Adenoviruses, Human↗

Structure analysis of two Toxoplasma gondii and Neospora caninum satellite DNA families and evolution of their common monomeric sequence.

A family of repetitive DNA elements of approximately 350 bp-Sat350-that are members of Toxoplasma gondii satellite DNA was further analyzed. Sequence analysis identified at least three distinct repeat types within this family, called types A, B, and C. B repeats were divided into the subtypes B1 and B2. A search for internal repetitions within this family permitted the identification of conserved regions and the design of PCR primers that amplify almost all these repetitive elements. These primers amplified the expected 350-bp repeats and a novel 680-bp repetitive element (Sat680) related to this family. Two additional tandemly repeated high-order structures corresponding to this satellite DNA family were found by searching the Toxoplasma genome database with these sequences. These studies were confirmed by sequence analysis and identified: (1). an arrangement of AB1CB2 350-bp repeats and (2). an arrangement of two 350-bp-like repeats, resulting in a 680-bp monomer. Sequence comparison and phylogenetic analysis indicated that both high-order structures may have originated from the same ancestral 350-bp repeat. PCR amplification, sequence analysis and Southern blot showed that similar high-order structures were also found in the Toxoplasma-sister taxon Neospora caninum. The Toxoplasma genome database (http://ToxoDB.org ) permitted the assembly of a contig harboring Sat350 elements at one end and a long nonrepetitive DNA sequence flanking this satellite DNA. The region bordering the Sat350 repeats contained two differentially expressed sequence-related regions and interstitial telomeric sequences.

Animals↗

The nasopalatine ducts and associated structures in the rhesus monkey (Macaca mulatta): topography, prenatal development, function, and phylogeny.

Morphological and developmental characteristics of the rhesus monkey nasopalatine duct system and associated primary palatal structures are described along with functional and phylogenetic considerations. Examination of five adult palates and coronal sections of 13 fetal palates together with dissections of a sixth adult specimen and of a 119-day-old fetal palate reveal that the lateral lobes of the tripartate incisive papilla cover clefts leading into the ducts. The ducts pierce the bony palate to enter the nasal fossae in proximity to the incisive suture. The ontogenetic stability of the duct path reflects the retention of ancient duct and primitive choanae relationships and functionally maintains an optimal oral odorant-to-receptor channel. Sixteen timed pregnancy specimens (35-100 days) provided histological material for documenting rostral nasopalatal development. Duct primordia, identified at 35 days, had by 40 days formed the medial duct walls (conjoined septum-papilla from the primary medial palatal component), the lateral duct walls (maxillary processes), and the rostral walls (fused maxillary-intermaxillary components). The caudal walls derive from the fusion of palatal shelves with the papilla (45 days), thus distinguishing primary and secondary fusion modes. Duct epithelial maturation occurs between 70 and 100 days. The absence of a vomeronasal system is attributed to reduction of olfaction in reproductive behavior, while the presence of the coevolved nasopalatine ducts is linked to the persistence of epiglottal-velar valving. The ducts serve as oral food-odor conduits in otherwise functionally separated respiratory and digestive tracts.

Animals↗