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PubMed2005

Characterization of Ty3-gypsy-like elements in Hibiscus syriacus.

Southern blot analysis revealed a ubiquitous distribu-tion and high copy number of Ty3-gypsy-like elements in the genome of Hibiscus syriacus. Comparative phylogenetic analysis of the large subunit of Rubisco and the integrase region of Ty3-gypsy elements in various plant species indicated that the retrotransposon-like sequences have different evolutionary histories and their own unique polymorphism in the H. syriacus population. Sequence-tagged site-restriction fragment length polymorphisms (STS-RFLP) analysis also indicated great variability in the numbers and sequences of Ty3-gypsy-like elements within H. syriacus varieties. Ty3-gypsy-like elements may still be active within H. syriacus, since Northern analysis of wounded leaves of H. syriacus variety Saehan with a probe for the integrase domain gave strong hybridization signals. The sequence heterogeneity and ubiquity of the Ty3-gypsy-like elements in H. syriacus genomes could provide reliable DNA markers for line identification as well for the analysis of genetic diversity in H. syriacus.

Amino Acid Sequence↗

PubMed1998

A maturase-encoding group III twintron is conserved in deeply rooted euglenoid species: are group III introns the chicken or the egg?

The fourth intron of the Euglena gracilis chloroplast photosystem II gene, psbCi4, is a 1,605-bp twintron composed of two group III introns and a coding locus for a 458-aa polypeptide, mat1, located in the internal intron. psbCi4 homologs have been identified in seven euglenoids, including E. myxocylindracea, E. viridis, E. deses, E. pisciformis, Cryptoglena pigra, Eutreptia sp., and Lepocinclis beutschlii. All of the species examined contain both the group III twintron and the mat1 locus, revealing a more widespread occurrence of group III introns than previously known. The L. beutschlii mat1 locus is interrupted by two novel mini-group II introns of 224 and 258 nt, the smallest group II introns yet identified. Reverse transcriptase polymerase chain reaction analysis confirmed the splicing boundaries of the external and internal E. myxocylindracea, E. viridis, and E. deses introns as well as the novel L. beutschlii mat1 introns. As determined by comparative phylogenetic analysis, group III introns contain a structural homolog of group II intron domain VI. The mat1 loci encode peptide motifs characteristic of group II intron maturases. A group III intron-encoded protein whose predicted sequence is similar to group II intron-encoded maturases and a bona fide domain VI within group III introns are compelling evidence for a common ancestor of group II and group III introns.

Amino Acid Sequence↗

PubMed1993

Prediction of common folding structures of homologous RNAs.

We have developed an algorithm and a computer program for simultaneously folding homologous RNA sequences. Given an alignment of M homologous sequences of length N, the program performs phylogenetic comparative analysis and predicts a common secondary structure conserved in the sequences. When the structure is not uniquely determined, it infers multiple structures which appear most plausible. This method is superior to energy minimization methods in the sense that it is not sensitive to point mutation of a sequence. It is also superior to usual phylogenetic comparative methods in that it does not require manual scrutiny for covariation or secondary structures. The most plausible 1-5 structures are produced in O(MN2 + N3) time and O(N2) space, which are the same requirements as those of widely used dynamic programs based on energy minimization for folding a single sequence. This is the first algorithm probably practical both in terms of time and space for finding secondary structures of homologous RNA sequences. The algorithm has been implemented in C on a Sun SparcStation, and has been verified by testing on tRNAs, 5S rRNAs, 16S rRNAs, TAR RNAs of human immunodeficiency virus type 1 (HIV-1), and RRE RNAs of HIV-1. We have also applied the program to cis-acting packaging sequences of HIV-1, for which no generally accepted structures yet exist, and propose potentially stable structures. Simulation of the program with random sequences with the same base composition and the same degree of similarity as the above sequences shows that structures common to homologous sequences are very unlikely to occur by chance in random sequences.

Algorithms↗

PubMed1996

Comparative analysis of ribonuclease P RNA structure in Archaea.

Although the structure of the catalytic RNA component of ribonuclease P has been well characterized in Bacteria, it has been little studied in other organisms, such as the Archaea. We have determined the sequences encoding RNase P RNA in eight euryarchaeal species: Halococcus morrhuae, Natronobacterium gregoryi, Halobacterium cutirubrum, Halobacteriurn trapanicum, Methanobacterium thermoautotrophicum strains deltaH and Marburg, Methanothermus fervidus and Thermococcus celer strain AL-1. On the basis of these and previously available sequences from Sulfolobus acidocaldarius, Haloferax volcanii and Methanosarcina barkeri the secondary structure of RNase P RNA in Archaea has been analyzed by phylogenetic comparative analysis. The archaeal RNAs are similar in both primary and secondary structure to bacterial RNase P RNAs, but unlike their bacterial counterparts these archaeal RNase P RNAs are not by themselves catalytically proficient in vitro.

Archaea↗

PubMed1995

Maintenance of pre-mRNA secondary structure by epistatic selection.

Linkage disequilibrium between polymorphisms in a natural population may result from various evolutionary forces, including random genetic drift due to sampling of gametes during reproduction, restricted migration between subpopulations in a subdivided population, or epistatic selection. In this report, we present evidence that the majority of significant linkage disequilibria observed in introns of the alcohol dehydrogenase locus (Adh) of Drosophila pseudoobscura are due to epistatic selection maintaining secondary structure of precursor mRNA (pre-mRNA). Based on phylogenetic-comparative analysis and a likelihood approach, we propose secondary structure models of Adh pre-mRNA for the regions of the adult intron and intron 2 where clustering of linkage disequilibria has been observed. Furthermore, we applied the likelihood ratio test to the phylogenetically predicted secondary structure in intron 1. In contrast to the other two structures, polymorphisms associated with the more conserved stem-loop structure of intron 1 are in low frequency, and linkage disequilibria have not been observed. These findings are qualitatively consistent with a model of compensatory fitness interactions. This model assumes that mutations disrupting pairing in a secondary structural element are individually deleterious if they destabilize a functionally important structure; a second "compensatory" mutation, however, may restabilize the structure and restore fitness.

Animals↗

PubMed1999

Conserved features of Y RNAs revealed by automated phylogenetic secondary structure analysis.

Y RNAs are small 'cytoplasmic' RNAs which are components of the Ro ribonucleoprotein (RNP) complex. The core of this complex, which is found in the cell nuclei of higher eukaryotes as well as the cytoplasm, is composed of a complex between the 60 kDa Ro protein and Y RNAs. Human cells contain four distinct Y RNAs (Y1, Y3, Y4 and Y5), while other eukaryotes contain a variable number of Y RNA homologues. When detected in a particular species, the Ro RNP has been present in every cell type within that particular organism. This characteristic, along with its high conservation among vertebrates, suggests an important function for Ro RNP in cellular metabolism; however, this function has not yet been definitively elucidated. In order to identify conserved features of Y RNA sequences and structures which may be directly involved in Ro RNP function, a phylogenetic comparative analysis of Y RNAs has been performed. Sequences of Y RNA homologues from five vertebrate species have been obtained and, together with previously published Y RNA sequences, used to predict Y RNA secondary structures. A novel RNA secondary structure comparison algorithm, the suboptimal RNA analysis program, has been developed and used in conjunction with available algorithms to find phylogenetically conserved secondary structure models for YI, Y3 and Y4 RNAs. Short, conserved sequences within the Y RNAs have been identified and are invariant among vertebrates, consistent with a direct role for Y RNAs in Ro function. A subset of these are located wholly or partially in looped regions in the Y3 and Y4 RNA predicted model structures, in accord with the possibility that these Y RNAs base pair with other cellular nucleic acids or are sites of interaction between the Ro RNP and other macromolecules.

Animals↗

PubMed1999

Genetic analysis of human immunodeficiency virus in Abidjan, Ivory Coast reveals predominance of HIV type 1 subtype A and introduction of subtype G.

To better understand the molecular epidemiology of HIV genetic diversity in Abidjan, Ivory Coast, we performed a genetic analysis of 170 HIV-1-seropositive specimens representing newly diagnosed tuberculosis patients (n = 143) and women monitored in a mother-to-child transmission cohort study (n = 27). Preliminary screening with RFLP presumptively classified 162 (95.3%) of these as subtype A. The envelope region of 108 specimens was subtyped by sequence analysis: 102 (94.4%) were subtype A, 2 (1.9%) were subtype D, and 4 (3.7%) were subtype G. Subtyping gag and env regions of the genome suggested that five of the six nonsubtype A isolates exhibited a potentially mosaic structure. A comparative phylogenetic analysis of HIV-1 subtype A C2V3 from 27 Ivory Coast and 21 Ugandan sequences revealed a striking clustering among Ivory Coast variants, and an independent segregation from Ugandan subtype A. Despite independent clustering with other subtype A specimens, limited variability of the V3 loop apex was observed; the globally predominant V3 motif, GPGQ, represented 90.1% of the HIV-1 strains. This study demonstrates that clade A is the predominant HIV-1 subtype in HIV-seropositive individuals in Abidjan, Ivory Coast and that these strains are phylogenetically distinct from other subtype A strains observed in East Africa.

Amino Acid Sequence↗

PubMed2002

Expression, purification, crystallization and preliminary diffraction analysis of RNase P protein from Thermotoga maritima.

Ribonuclease P (RNase P), the ubiquitous endonuclease that catalyzes maturation of the 5'-end of tRNA in bacteria, is a ribonucleoprotein particle composed of one large RNA and one small protein. Two major structural types of bacterial RNase P RNA have been identified by phylogenetic comparative analysis: the A (ancestral) and B (Bacillus) types. The RNase P protein from Thermotoga maritima, a hyperthermophilic bacterium with an A-type RNase P RNA, has been expressed in Escherichia coli. A purification strategy was developed to obtain a protein preparation suitable for crystallization. Protein crystals suitable for diffraction studies were obtained and characterized.

Crystallography, X-Ray↗

PubMed2001

A comparative analysis of the evolutionary patterning and mechanistic bases of lactate dehydrogenase thermal stability in porcelain crabs, genus Petrolisthes.

The kinetic properties of orthologous homologs (orthologs) of enzymes are typically correlated with environmental temperatures in species adapted to different thermal regimes, but correlations between adaptation temperature and enzyme thermal stability are less clear. Although the thermal stability of a protein is related chiefly to its primary structure (including post-translational modification), thermal stability can also be altered by extrinsic factors present in the intracellular milieu. Here, we present a comparative analysis of the thermal stability of lactate dehydrogenase (LDH) orthologs from 22 congeneric species of porcelain crab (genera Petrolisthes and Allopetrolisthes) from a broad range of thermal habitats. Interspecific diversity of LDH stability is high: temperatures required for a 50 % loss of activity in 10 min ranged from 65 to 75.5 degrees C, corresponding to half-lives of less than 1 min to more than 3 h at 70 degrees C. Although stability is positively correlated with maximal habitat temperature in some sister taxa, phylogenetic comparative analysis incorporating all 22 species does not indicate that the interspecific diversity of LDH stability represents an adaptive response to current thermal habitats. Examination of the mechanistic bases of LDH stabilization indicates that differences in stability are related both to properties of the LDH molecule itself (intrinsic stability) and to the effects of extrinsic protein(s). Intrinsic differences were shown by the unfolding of structure during heating, as measured by circular dichroism spectroscopy. Stabilizing effects of extrinsic proteins are implied by the results of cellular fractionation experiments that removed low-molecular-mass solutes and proteins from the muscle homogenates. We conclude that the overall structural stability and functional properties of proteins can evolve independently and that in vivo protein-protein interactions can provide another means to regulate protein stability selectively.

Animals↗

PubMed2004

Evolution and stability of the G-matrix on a landscape with a moving optimum.

In quantitative genetics, the genetic architecture of traits, described in terms of variances and covariances, plays a major role in determining the trajectory of evolutionary change. Hence, the genetic variance-covariance matrix (G-matrix) is a critical component of modern quantitative genetics theory. Considerable debate has surrounded the issue of G-matrix constancy because unstable G-matrices provide major difficulties for evolutionary inference. Empirical studies and analytical theory have not resolved the debate. Here we present the results of stochastic models of G-matrix evolution in a population responding to an adaptive landscape with an optimum that moves at a constant rate. This study builds on the previous results of stochastic simulations of G-matrix stability under stabilizing selection arising from a stationary optimum. The addition of a moving optimum leads to several important new insights. First, evolution along genetic lines of least resistance increases stability of the orientation of the G-matrix relative to stabilizing selection alone. Evolution across genetic lines of least resistance decreases G-matrix stability. Second, evolution in response to a continuously changing optimum can produce persistent maladaptation for a correlated trait, even if its optimum does not change. Third, the retrospective analysis of selection performs very well when the mean G-matrix (G) is known with certainty, indicating that covariance between G and the directional selection gradient beta is usually small enough in magnitude that it introduces only a small bias in estimates of the net selection gradient. Our results also show, however, that the contemporary G-matrix only serves as a rough guide to G. The most promising approach for the estimation of G is probably through comparative phylogenetic analysis. Overall, our results show that directional selection actually can increase stability of the G-matrix and that retrospective analysis of selection is inherently feasible. One major remaining challenge is to gain a sufficient understanding of the G-matrix to allow the confident estimation of G.

Adaptation, Biological↗

PubMed2004

Analysis and comparison of the 4b core protein gene of avipoxviruses from wild birds: evidence for interspecies spatial phylogenetic variation.

Avipoxviruses have been isolated from a wide variety of avian hosts, and yet little is known regarding the host-virus species variation of the genus Avipoxvirus. We have investigated the variations in the viral 4b core protein gene from six different avipoxviruses based on PCR, Southern blot and nucleotide sequence analysis to evaluate the suitability of this region for differentiation between avipoxvirus isolates. Southern blot and nucleotide sequence analysis revealed considerable interspecies variation between the different virus isolates. In the deduced amino acid sequences (of 142 residues) of the 4b core protein gene, fowlpox virus vaccine strain (FPV-VR250) was found to be similar to the three poxvirus isolates from great tit (GTV-A310, GTV-A311 and GTV-A256), sparrowpox virus (SPV-A468), and pigeonpox virus (PPV-B7) with similarities of 79.6%, 81%, 81%, 64.8% and 84.5%, respectively. Furthermore, comparative phylogenetic analysis of the aligned DNA sequences revealed divergence among the different viruses that can be consistently correlated to the host.

Amino Acid Sequence↗

PubMed1991

Predicting common foldings of homologous RNAs.

A new approach is proposed for determining common RNA secondary structures within a set of homologous RNAs. The approach is a combination of phylogenetic and thermodynamic methods which is based on the prediction of optimal and suboptimal secondary structures, topological similarity searches and phylogenetic comparative analysis. The optimal and suboptimal RNA secondary structures are predicted by energy minimization. Structural comparison of the predicted RNA secondary structures is used to find conserved structures that are topologically similar in all these homologous RNAs. The validity of the conserved structural elements found is then checked by phylogenetic comparison of the sequences. This procedure is used to predict common structures of ribonuclease P (RNAase P) RNAs.

Algorithms↗

PubMedDate not supplied

Analysis of the tertiary structure of bacterial RNase P RNA.

The ubiquitous occurrence of ribonuclease P (RNase P) as a ribonucleoprotein and the catalytic properties of bacterial RNase P RNAs indicate that RNA fulfills an ancient and important role in the function of this enzyme. This review focuses on efforts to determine the structure of the bacterial RNase P RNA ribozyme. Phylogenetic comparative analysis of a library of bacterial RNase P RNA sequences has resulted in a well-developed secondary structure model and allowed identification of some elements of tertiary structure. The native structure has been redesigned by circular permutation to facilitate intra- and inter-molecular crosslinking experiments in order to gain further structural information. The crosslinking constraints, together with the constraints provided by comparative analyses, have been incorporated into a first-order model of the structure of of the ribozyme-substrate complex. The developing structural perspective allows the design of self-cleaving pre-tRNA-RNase P RNA conjugates which are useful tools for additional structure-probing experiments.

Bacteria↗

PubMed1995

Analysis of the phylogenetic placement of different spumaretroviral genes reveals complex pattern of foamy virus evolution.

Foamy or spumaviruses are complex retroviruses. Phylogenetic trees have been constructed previously for either the polymerase or integrase domains showed a clustering of the foamy viruses relatively distant from other retroviruses. The most related retrovirus was found to be murine leukemia virus, irrespective of the method used or foamy viral gene analyzed. We analyze bel genes of different foamy viruses and compared the corresponding phylogenetic trees with those obtained from the pol genes that were constructed with refined computer programs. In addition, the nucleocapsid protein sequence of foamy viruses was used for a comparative phylogenetic analysis. Known biological properties of the individual FV protein domains are discussed to ascertain the apparent phylogenetic relatedness.

Amino Acid Sequence↗

PubMed2003

Molecular identification and expression analysis of tumor necrosis factor in channel catfish (Ictalurus punctatus).

A tumor necrosis factor (TNF) alpha-like gene, encoding a propeptide of 230 amino acids and a mature (soluble) peptide of 162 amino acids, was identified in channel catfish (Ictalurus punctatus). While the catfish protein shared features in common with both mammalian TNFalpha and TNFbeta homologs, overall sequence identity/similarity was slightly higher vs. TNFalpha genes when mature TNF sequences were compared. Phylogenetic analysis placed catfish and other fish TNF sequences within their own cluster apart from mammalian TNFalpha and beta genes, and supported the suggestion that TNFalpha and beta genes separated after the divergence of mammals and teleosts. In contrast to trout and carp, but similar to flounder, catfish TNF was present as a single copy gene. Expression studies demonstrated that catfish TNFalpha mRNA was present in all tested tissues (i.e. liver, spleen, head kidney, mesonephros, gill, thymus, and PBLs) from an unstimulated fish. Moreover, catfish TNF was constitutively expressed in actively proliferating, but otherwise unstimulated, macrophage (42TA) and T cell (G14D; TS32.17) lines, but not in B cell (1G8 or 3B11) or fibroblast lines. TNF expression was upregulated in PBLs, and in G14D and 42TA cells, but not in 3B11 cells, by PMA/calcium ionophore treatment. These results demonstrate that a catfish homolog of TNFalpha has been identified, and indicate that catfish TNFalpha is expressed in catfish in a manner similar to that seen in mammals.

Amino Acid Sequence↗

PubMed1996

Anoxygenic phototrophy across the phylogenetic spectrum: current understanding and future perspectives

The phylogenetic heterogeneity of anoxygenic phototrophic bacteria has been revealed by 16S rRNA sequence analysis, the results of which have led to extensive taxonomic rearrangements within previously defined taxa of phototrophs and stimulated interest in this group of organisms. Anoxygenic photosynthetic bacteria can be found within 4 of the 12 phylogenetic lineages, and in some cases are highly related to non-photosynthetic members of these groups. The largest number of phototrophs are found in the class Proteobacteria. Comparative phylogenetic analysis using 23S rDNA sequences generally supports the topology obtained from 16S rDNA sequences. The photosynthetic reaction centers are conserved in all photosynthetic bacteria, and are of two types. One is shared by the Proteobacteria and Chloroflexus aurantiacus and is similar to Photosystem II of cyanobacteria, while heliobacteria and Chlorobium and relatives possess a reaction center similar to the cyanobacterial Photosystem I. These similarities are supported by sequence analysis of core reaction center peptides, but contradict phylogenies reconstructed from rRNA sequence analysis. Genome analysis by means of physical mapping has been performed for only three species of anoxygenic phototrophs. Some conservation of operon structure and gene sequence has been found within the Proteobacteria, but does not extend to other phototrophs.

Journal Article↗

PubMed1997

The internal transcribed spacer 2 exhibits a common secondary structure in green algae and flowering plants.

Sequences of the Internal Transcribed Spacer 2 (ITS-2) regions of the nuclear rDNA repeats from 111 organisms of the family Volvocaceae (Chlorophyta) and unicellular organisms of the Volvocales, including Chlamydomonas reinhardtii, were determined. The use of thermodynamic energy optimization to generate secondary structures and phylogenetic comparative analysis of the spacer regions revealed a common secondary structure that is conserved despite wide intra- and interfamilial primary sequence divergence. The existence of this conserved higher-order structure is supported by the presence of numerous compensating basepair changes as well as by an evolutionary history of insertions and deletions that nevertheless maintains major aspects of the overall structure. Furthermore, this general structure is preserved across broad phylogenetic lines, as it is observed in the ITS-2s of other chlorophytes, including flowering plants; previous reports of common ITS-2 secondary structures in other eukaryotes were restricted to the order level. The reported ITS-2 structure possesses important conserved structural motifs which may help to mediate cleavages in the ITS-2 that occur during rRNA transcript processing. Their recognition can guide further studies of eukaryotic rRNA processing, and their application to sequence alignments may contribute significantly to the value of ITS-2 sequences in phylogenetic analyses at several taxonomic levels, but particularly in characterizing populations and species.

Base Sequence↗

PubMed1997

Identification and characterization of IS-like elements in Mycobacterium gordonae.

A new insertion sequence, IS1512, from Mycobacterium gordonae was cloned and sequenced. This element is present in up to 10 copies which provides a high diversity for restriction fragment length polymorphism analysis. We have also identified truncated IS1512-like elements, including a truncated IS1512 and another truncated insertion sequence which displays homology with IS1512 and was designated IS1511. Sequences homologous to the previously described transposon Tn554 are inserted into these truncated insertion sequences. Insertion loci of IS1511/IS1512 are shown to be highly related to those described for IS256-like elements in other species. Alignment of the putative transposases from Rhodococcus and Mycobacterium suggests these insertion sequences may form a distinct closely homologous subclass within the IS256 family. Analysis comparing phylogenetic divergence of these elements with that of 16S rRNA and superoxide dismutase genes suggests horizontal transfer of a IS1511/IS1512 precursor into M. gordonae, and the occurrence of horizontal transfer between Mycobacteriaceae and Rhodococcaceae.

Actinomyces↗