Long-range correlations in prokaryotic chromosomes

One of the fascinating properties of the DNA sequences of prokaryotic and eukaryotic chromosomes is that they possess long-range order. Computational methods like spectral analysis, mutual information and DNA random walks have been used to probe long-range order via-long range correlations. This work attempts to show the advantage of using the Information Theoretic measure of mutual information for this purpose. A number Mu is found which indicates the existence of long-range order. Mu is the ratio between the value of mutual information function between two nucleotides of a DNA sequence separated by a large distance of 100 kilobases to the value expected from a randomized sequence of the same DNA. It is found that in spite of the constant shuffling of nucleotides due to insertion, deletion, inversion and recombination that occur during evolution, the chromosomal structure of prokaryotes is not always mosaic. While all archaeal chromosomes show mosaic structure and lack long-range order, a sizable fraction of the bacterial chromosomes do possess long-range order. A statistical multivariate analysis has been done to find which of the physical variables like genome size or GC% affects the organization of the chromosome or correlates with the long-range order. The existence of long-range order in bacterial chromosomes could be directly correlated to the degree of gene strand bias shown by it. Firmicutes which have low GC content also have pronounced strand bias and show long-range correlations. It is observed that the occurrence of long-range order in bacteria is independent of genome size, but depends on its GC content and gene strand bias.     

Origin of replication in circular prokaryotic chromosomes

To predict origins of replication in prokaryotic chromosomes, we analyse the leading and lagging strands of 200 chromosomes for differences in oligomer composition and show that these correlate strongly with taxonomic grouping, lifestyle and molecular details of the replication process. While all bacteria have a preference for Gs over Cs on the leading strand, we discover that the direction of the A/T skew is determined by the polymerase-alpha subunit that replicates the leading strand. The strength of the strand bias varies greatly between both phyla and environments and appears to correlate with growth rate. Finally we observe much greater diversity of skew among archaea than among bacteria. We have developed a program that accurately locates the origins of replication by measuring the differences between leading and lagging strand of all oligonucleotides up to 8 bp in length. The program and results for all publicly available genomes are available from http://www.cbs.dtu.dk/services/GenomeAtlas/suppl/origin.     

Fixation dynamics of beneficial alleles in prokaryotic polyploid chromosomes and plasmids

Theoretical population genetics has been mostly developed for sexually reproducing diploid and for monoploid (haploid) organisms, focusing on eukaryotes. The evolution of bacteria and archaea is often studied by models for the allele dynamics in monoploid populations. However, many prokaryotic organisms harbor multicopy replicons—chromosomes and plasmids—and theory for the allele dynamics in populations of polyploid prokaryotes remains lacking. Here, we present a population genetics model for replicons with multiple copies in the cell. Using this model, we characterize the fixation process of a dominant beneficial mutation at 2 levels: the phenotype and the genotype. Our results show that depending on the mode of replication and segregation, the fixation of the mutant phenotype may precede genotypic fixation by many generations; we term this time interval the heterozygosity window. We furthermore derive concise analytical expressions for the occurrence and length of the heterozygosity window, showing that it emerges if the copy number is high and selection strong. Within the heterozygosity window, the population is phenotypically adapted, while both alleles persist in the population. Replicon ploidy thus allows for the maintenance of genetic variation following phenotypic adaptation and consequently for reversibility in adaptation to fluctuating environmental conditions.     

Polarisation, key to good localisation

Polarisation of cells is crucial for vectorial transport of ions and solutes. In literature, however, proteins specifically targeted to the apical or basolateral membrane are often studied in non-polarised cells. To investigate whether these data can be extrapolated to expression in polarised cells, we studied several membrane-specific proteins. In polarised MDCK cells, the Aquaporin-2 water channel resides in intracellular vesicles and apical membrane, while the vasopressin-type 2 receptor, anion-exchanger 1 (AE1) protein and E-Cadherin mainly localise to the basolateral membrane. In non-polarised MDCK cells, however, Aquaporin-2 localises, besides plasma membrane, mainly in the Golgi complex, while the others show a dispersed staining throughout the cell. Moreover, while AQP2 mutants in dominant nephrogenic diabetes insipidus are missorted to different organelles in polarised cells, they all predominantly localise to the Golgi complex in non-polarised MDCK cells. Additionally, the maturation of V2R, and likely its missorting, is affected in transiently-transfected compared to stably-transfected cells. In conclusion, we show that the use of stably-transfected polarised cells is crucial in interpreting the processing and the localisation of membrane targeted proteins.     

Polarisation, key to good localisation

Polarisation of cells is crucial for vectorial transport of ions and solutes. In literature, however, proteins specifically targeted to the apical or basolateral membrane are often studied in non-polarised cells. To investigate whether these data can be extrapolated to expression in polarised cells, we studied several membrane-specific proteins. In polarised MDCK cells, the Aquaporin-2 water channel resides in intracellular vesicles and apical membrane, while the vasopressin-type 2 receptor, anion-exchanger 1 (AE1) protein and E-Cadherin mainly localise to the basolateral membrane. In non-polarised MDCK cells, however, Aquaporin-2 localises, besides plasma membrane, mainly in the Golgi complex, while the others show a dispersed staining throughout the cell. Moreover, while AQP2 mutants in dominant nephrogenic diabetes insipidus are missorted to different organelles in polarised cells, they all predominantly localise to the Golgi complex in non-polarised MDCK cells. Additionally, the maturation of V2R, and likely its missorting, is affected in transiently-transfected compared to stably-transfected cells. In conclusion, we show that the use of stably-transfected polarised cells is crucial in interpreting the processing and the localisation of membrane targeted proteins.     

Recombination of synthetic oligonucleotides with prokaryotic chromosomes: substrate requirements of the Escherichia coli/lambdaRed and Sulfolobus acidocaldarius recombination systems

In order to reveal functional properties of recombination involving short ssDNAs in hyperthermophilic archaea, we evaluated oligonucleotide-mediated transformation (OMT) in Sulfolobus acidocaldarius and Escherichia coli as a function of the molecular properties of the ssDNA substrates. Unmodified ssDNAs as short as 20–22 nt yielded recombinants in both organisms, as did longer DNAs forming as few as 2–5 base pairs on one side of the genomic mutation. The two OMT systems showed similar responses to certain end modifications of the oligonucleotides, but E. coli was found to require a 5' phosphate on 5'-limited ssDNA whereas this requirement was not evident in S. acidocaldarius . The ability of both E. coli and S. acidocaldarius to incorporate short, mismatched ssDNAs into their genomes raises questions about the biological significance of this capability, including its phylogenetic distribution among microorganisms and its impact on genome stability. These questions seem particularly relevant for S. acidocaldarius , as this archaeon has natural competence for OMT, encodes no MutSL homologues and thrives under environmental conditions that accelerate DNA decomposition.     

Deterministic Versus Stochastic Cell Polarisation Through Wave-Pinning

Cell polarization is an important part of the response of eukaryotic cells to stimuli, and forms a primary step in cell motility, differentiation, and many cellular functions. Among the important biochemical players implicated in the onset of intracellular asymmetries that constitute the early phases of polarization are the Rho GTPases, such as Cdc42, Rac, and Rho, which present high active concentration levels in a spatially localized manner. Rho GTPases exhibit positive feedback-driven interconversion between distinct active and inactive forms, the former residing on the cell membrane, and the latter predominantly in the cytosol. A?deterministic model of the dynamics of a single Rho GTPase described earlier by Mori et al.?exhibits sustained polarization by a wave-pinning mechanism. It remained, however, unclear how such polarization behaves at typically low cellular concentrations, as stochasticity could significantly affect the dynamics. We therefore study the low copy number dynamics of this model, using a stochastic kinetics framework based on the Gillespie algorithm, and propose statistical and analytic techniques which help us analyse the equilibrium behaviour of our stochastic system. We use local perturbation analysis to predict parameter regimes for initiation of polarity and wave-pinning in our deterministic system, and compare these predictions with deterministic and stochastic spatial simulations. Comparing the behaviour of the stochastic with the deterministic system, we determine the threshold number of molecules required for robust polarization in a given effective reaction volume. We show that when the molecule number is lowered wave-pinning behaviour is lost due to an increasingly large transition zone as well as increasing fluctuations in the pinning position, due to which a broadness can be reached that is unsustainable, causing the collapse of the wave, while the variations in the high and low equilibrium levels are much less affected.     

Evolutionary trends in the prokaryotic community and prokaryotic community-phage systems

The Evolutionary Constructor software has been used for computer simulation of the life and evolution of communities of unicellular haploid organisms (prokaryotic cells). Opposite trends of the community evolution (simplification and complication of the genome) have been studied. It has been demonstrated that species with reduced genomes tend to replace genetically and metabolically rich species under highly favorable environmental conditions. Under unfavorable conditions, the opposite tendency is observed. It has also been shown that introduction of phages capable of killing the cells into the system may radically change the current evolutionary trend.     

Ecology of prokaryotic viruses

The finding that total viral abundance is higher than total prokaryotic abundance and that a significant fraction of the prokaryotic community is infected with phages in aquatic systems has stimulated research on the ecology of prokaryotic viruses and their role in ecosystems. This review treats the ecology of prokaryotic viruses ('phages') in marine, freshwater and soil systems from a 'virus point of view'. The abundance of viruses varies strongly in different environments and is related to bacterial abundance or activity suggesting that the majority of the viruses found in the environment are typically phages. Data on phage diversity are sparse but indicate that phages are extremely diverse in natural systems. Lytic phages are predators of prokaryotes, whereas lysogenic and chronic infections represent a parasitic interaction. Some forms of lysogeny might be described best as mutualism. The little existing ecological data on phage populations indicate a large variety of environmental niches and survival strategies. The host cell is the main resource for phages and the resource quality, i.e., the metabolic state of the host cell, is a critical factor in all steps of the phage life cycle. Virus-induced mortality of prokaryotes varies strongly on a temporal and spatial scale and shows that phages can be important predators of bacterioplankton. This mortality and the release of cell lysis products into the environment can strongly influence microbial food web processes and biogeochemical cycles. Phages can also affect host diversity, e.g., by 'killing the winner' and keeping in check competitively dominant species or populations. Moreover, they mediate gene transfer between prokaryotes, but this remains largely unknown in the environment. Genomics or proteomics are providing us now with powerful tools in phage ecology, but final testing will have to be performed in the environment.     

Actin's prokaryotic homologs

Actin is one of the most abundant and conserved eukaryotic proteins. Remarkably, two prokaryotic homologs of actin, MreB and ParM, have only recently been identified. MreB and ParM polymerize into filaments and play important roles in the control of bacterial cell shape and plasmid segregation, respectively. Whereas the eukaryotic actins display a remarkable degree of conservation (e.g. no amino acid changes in muscle actin from chickens to humans), the two bacterial proteins have as much sequence similarity to each other ( approximately 11% sequence identity) as they do to actin. It is possible that the interesting properties of eukaryotic F-actin may account for the unusual degree of conservation among the actins, whereas the bacterial proteins have had fewer constraints over the course of evolution.     

原核微生物的多样性

微生物是一群以分解代谢为主的重要生物类群,其生物学多样性十分丰富。但由于它们的微观性,尤其原核微生物简单的单细胞结构、以无性方式进行快速地繁殖而造成的无准确的基线难以对其进行种群数目和数量的统计,因而对微生物的多样性研究远没有宏观生物那样深入和受到重视。本文根据原核微生物的特性,从其物种、所代表的进化分支、生理代谢类群及遗传背景几个方面简述了它们的多样性及重要意义,意在引起科学界和全社会对这类生物资源的认识和保护的重视。     

The prokaryotic selenoproteome

In the genetic code, the UGA codon has a dual function as it encodes selenocysteine (Sec) and serves as a stop signal. However, only the translation terminator function is used in gene annotation programs, resulting in misannotation of selenoprotein genes. Here, we applied two independent bioinformatics approaches to characterize a selenoprotein set in prokaryotic genomes. One method searched for selenoprotein genes by identifying RNA stem-loop structures, selenocysteine insertion sequence elements; the second approach identified Sec/Cys pairs in homologous sequences. These analyses identified all or almost all selenoproteins in completely sequenced bacterial and archaeal genomes and provided a view on the distribution and composition of prokaryotic selenoproteomes. In addition, lineage-specific and core selenoproteins were detected, which provided insights into the mechanisms of selenoprotein evolution. Characterization of selenoproteomes allows interpretation of other UGA codons in completed genomes of prokaryotes as terminators, addressing the UGA dual-function problem.     

Whole-genome prokaryotic phylogeny

Current understanding of the phylogeny of prokaryotes is based on the comparison of the highly conserved small ssu-rRNA subunit and similar regions. Although such molecules have proved to be very useful phylogenetic markers, mutational saturation is a problem, due to their restricted lengths. Now, a growing number of complete prokaryotic genomes are available. This paper addresses the problem of determining a prokaryotic phylogeny utilizing the comparison of complete genomes. We introduce a new strategy, GBDP, 'genome blast distance phylogeny', and show that different variants of this approach robustly produce phylogenies that are biologically sound, when applied to 91 prokaryotic genomes. In this approach, first Blast is used to compare genomes, then a distance matrix is computed, and finally a tree- or network-reconstruction method such as UPGMA, Neighbor-Joining, BioNJ or Neighbor-Net is applied.