Structures of archaeal DNA segregation machinery reveal bacterial and eukaryotic linkages.

PubMed

Schumacher, Maria A; Tonthat, Nam K; Lee, Jeehyun; Rodriguez-Castañeda, Fernando A; Chinnam, Naga Babu; Kalliomaa-Sanford, Anne K; Ng, Irene W; Barge, Madhuri T; Shaw, Porsha L R; Barillà, Daniela

2015-09-04

Although recent studies have provided a wealth of information about archaeal biology, nothing is known about the molecular basis of DNA segregation in these organisms. Here, we unveil the machinery and assembly mechanism of the archaeal Sulfolobus pNOB8 partition system. This system uses three proteins: ParA; an atypical ParB adaptor; and a centromere-binding component, AspA. AspA utilizes a spreading mechanism to create a DNA superhelix onto which ParB assembles. This supercomplex links to the ParA motor, which contains a bacteria-like Walker motif. The C domain of ParB harbors structural similarity to CenpA, which dictates eukaryotic segregation. Thus, this archaeal system combines bacteria-like and eukarya-like components, which suggests the possible conservation of DNA segregation principles across the three domains of life. Copyright © 2015, American Association for the Advancement of Science.

A novel mechanism for direct real-time polymerase chain reaction that does not require DNA isolation from prokaryotic cells.

PubMed

Soejima, Takashi; Xiao, Jin-Zhong; Abe, Fumiaki

2016-06-23

Typically, polymerase chain reaction (PCR) is performed after DNA isolation. Real-time PCR (qPCR), also known as direct qPCR in mammalian cells with weak membranes, is a common technique using crude samples subjected to preliminary boiling to elute DNA. However, applying this methodology to prokaryotic cells, which have solid cell walls, in contrast to mammalian cells which immediately burst in water, can result in poor detection. We successfully achieved PCR elongation with the addition of 1.3 cfu of Cronobacter muytjensii to a newly developed direct qPCR master mix without performing any crude DNA extraction (detection limit of 1.6 × 10(0) cfu/ml for the test sample compared with a detection limit of 1.6 × 10(3) cfu/ml primarily for crude (boiling) or classical DNA isolation). We revealed that the chromosomal DNA retained in prokaryotic cells can function as a PCR template, similarly to the mechanism in in situ PCR. Elucidating this reaction mechanism may contribute to the development of an innovative master mix for direct qPCR to detect genes in a single bacterium with solid cell walls and might lead to numerous novel findings in prokaryotic genomics research.

A three-dimensional ParF meshwork assembles through the nucleoid to mediate plasmid segregation

PubMed Central

McLeod, Brett N.; Allison-Gamble, Gina E.; Barge, Madhuri T.; Tonthat, Nam K.; Schumacher, Maria A.; Hayes, Finbarr

2017-01-01

Abstract Genome segregation is a fundamental step in the life cycle of every cell. Most bacteria rely on dedicated DNA partition proteins to actively segregate chromosomes and low copy-number plasmids. Here, by employing super resolution microscopy, we establish that the ParF DNA partition protein of the ParA family assembles into a three-dimensional meshwork that uses the nucleoid as a scaffold and periodically shuttles between its poles. Whereas ParF specifies the territory for plasmid trafficking, the ParG partner protein dictates the tempo of ParF assembly cycles and plasmid segregation events by stimulating ParF adenosine triphosphate hydrolysis. Mutants in which this ParG temporal regulation is ablated show partition deficient phenotypes as a result of either altered ParF structure or dynamics and indicate that ParF nucleoid localization and dynamic relocation, although necessary, are not sufficient per se to ensure plasmid segregation. We propose a Venus flytrap model that merges the concepts of ParA polymerization and gradient formation and speculate that a transient, dynamic network of intersecting polymers that branches into the nucleoid interior is a widespread mechanism to distribute sizeable cargos within prokaryotic cells. PMID:28034957

DNA replication fidelity in Mycobacterium tuberculosis is mediated by an ancestral prokaryotic proofreader.

PubMed

Rock, Jeremy M; Lang, Ulla F; Chase, Michael R; Ford, Christopher B; Gerrick, Elias R; Gawande, Richa; Coscolla, Mireia; Gagneux, Sebastien; Fortune, Sarah M; Lamers, Meindert H

2015-06-01

The DNA replication machinery is an important target for antibiotic development in increasingly drug-resistant bacteria, including Mycobacterium tuberculosis. Although blocking DNA replication leads to cell death, disrupting the processes used to ensure replication fidelity can accelerate mutation and the evolution of drug resistance. In Escherichia coli, the proofreading subunit of the replisome, the ɛ exonuclease, is essential for high-fidelity DNA replication; however, we find that the corresponding subunit is completely dispensable in M. tuberculosis. Rather, the mycobacterial replicative polymerase DnaE1 itself encodes an editing function that proofreads DNA replication, mediated by an intrinsic 3'-5' exonuclease activity within its PHP domain. Inactivation of the DnaE1 PHP domain increases the mutation rate by more than 3,000-fold. Moreover, phylogenetic analysis of DNA replication proofreading in the bacterial kingdom suggests that E. coli is a phylogenetic outlier and that PHP domain-mediated proofreading is widely conserved and indeed may be the ancestral prokaryotic proofreader.

DNA replication fidelity in Mycobacterium tuberculosis is mediated by an ancestral prokaryotic proofreader

PubMed Central

Rock, Jeremy M.; Lang, Ulla F.; Chase, Michael R.; Ford, Christopher B.; Gerrick, Elias R.; Gawande, Richa; Coscolla, Mireia; Gagneux, Sebastien; Fortune, Sarah M.; Lamers, Meindert H.

2015-01-01

The DNA replication machinery is an important target for antibiotic development for increasingly drug resistant bacteria including Mycobacterium tuberculosis1. While blocking DNA replication leads to cell death, disrupting the processes used to ensure replication fidelity can accelerate mutation and the evolution of drug resistance. In E. coli, the proofreading subunit of the replisome, the ε-exonuclease, is essential for high fidelity DNA replication2; however, we find that it is completely dispensable in M. tuberculosis. Rather, the mycobacterial replicative polymerase, DnaE1, encodes a novel editing function that proofreads DNA replication, mediated by an intrinsic 3′-5′ exonuclease activity within its PHP domain. Inactivation of the DnaE1 PHP domain increases the mutation rate by greater than 3,000 fold. Moreover, phylogenetic analysis of DNA replication proofreading in the bacterial kingdom suggests that E. coli is a phylogenetic outlier and that PHP-domain mediated proofreading is widely conserved and indeed may be the ancestral prokaryotic proofreader. PMID:25894501

Stem cell identity and template DNA strand segregation.

PubMed

Tajbakhsh, Shahragim

2008-12-01

The quest for stem cell properties to distinguish their identity from that of committed daughters has led to a re-investigation of the notion that DNA strands are not equivalent, and 'immortal' DNA strands are retained in stem cells whereas newly replicated DNA strands segregate to the differentiating daughter cell during mitosis. Whether this process occurs only in stem cells, and also in all tissues, remains unclear. That individual chromosomes can be also partitioned non-randomly raises the question if this phenomenon is related to the immortal DNA hypothesis, and it underscores the need for high-resolution techniques to observe these events empirically. Although initially postulated as a mechanism to avoid DNA replication errors, alternative views including epigenetic regulation and sister chromatid silencing may provide insights into this process.

A three-dimensional ParF meshwork assembles through the nucleoid to mediate plasmid segregation.

PubMed

McLeod, Brett N; Allison-Gamble, Gina E; Barge, Madhuri T; Tonthat, Nam K; Schumacher, Maria A; Hayes, Finbarr; Barillà, Daniela

2017-04-07

Genome segregation is a fundamental step in the life cycle of every cell. Most bacteria rely on dedicated DNA partition proteins to actively segregate chromosomes and low copy-number plasmids. Here, by employing super resolution microscopy, we establish that the ParF DNA partition protein of the ParA family assembles into a three-dimensional meshwork that uses the nucleoid as a scaffold and periodically shuttles between its poles. Whereas ParF specifies the territory for plasmid trafficking, the ParG partner protein dictates the tempo of ParF assembly cycles and plasmid segregation events by stimulating ParF adenosine triphosphate hydrolysis. Mutants in which this ParG temporal regulation is ablated show partition deficient phenotypes as a result of either altered ParF structure or dynamics and indicate that ParF nucleoid localization and dynamic relocation, although necessary, are not sufficient per se to ensure plasmid segregation. We propose a Venus flytrap model that merges the concepts of ParA polymerization and gradient formation and speculate that a transient, dynamic network of intersecting polymers that branches into the nucleoid interior is a widespread mechanism to distribute sizeable cargos within prokaryotic cells. © The Author(s) 2016. Published by Oxford University Press on behalf of Nucleic Acids Research.

Computational Model for DNA Organization Mediated by Protein Interaction in Prokaryotes

NASA Astrophysics Data System (ADS)

Garimella, Karthik; Kharel, Savan

2016-03-01

In Escherichia Coli, there are several mechanisms that drive chromosomal organization. We know through experiments that the E. Coli chromosome is condensed into highly structured regions known as macrodomains (MDs). One of the regions known as the Terminus undergoes DNA-bridging condensation that form loops between distant DNA sites and it is known to be mediated by a Terminus specific protein, which binds to specific markers within the Terminus region. In the absence of Terminus specific protein, however, the Terminus region is known to not condense nearly as much, which will likely impede several biological processes including DNA replication. In order to understand the molecular basis of protein mediation in vivo several models of Terminus specific segregation have been constructed in silico which model DNA as polymer chains.

The bacterial segrosome: a dynamic nucleoprotein machine for DNA trafficking and segregation.

PubMed

Hayes, Finbarr; Barillà, Daniela

2006-02-01

The genomes of unicellular and multicellular organisms must be partitioned equitably in coordination with cytokinesis to ensure faithful transmission of duplicated genetic material to daughter cells. Bacteria use sophisticated molecular mechanisms to guarantee accurate segregation of both plasmids and chromosomes at cell division. Plasmid segregation is most commonly mediated by a Walker-type ATPase and one of many DNA-binding proteins that assemble on a cis-acting centromere to form a nucleoprotein complex (the segrosome) that mediates intracellular plasmid transport. Bacterial chromosome segregation involves a multipartite strategy in which several discrete protein complexes potentially participate. Shedding light on the basis of genome segregation in bacteria could indicate new strategies aimed at combating pathogenic and antibiotic-resistant bacteria.

Use of mariner transposases for one-step delivery and integration of DNA in prokaryotes and eukaryotes by transfection

PubMed Central

Michlewski, Gracjan; Finnegan, David J.; Elfick, Alistair; Rosser, Susan J.

2017-01-01

Abstract Delivery of DNA to cells and its subsequent integration into the host genome is a fundamental task in molecular biology, biotechnology and gene therapy. Here we describe an IP-free one-step method that enables stable genome integration into either prokaryotic or eukaryotic cells. A synthetic mariner transposon is generated by flanking a DNA sequence with short inverted repeats. When purified recombinant Mos1 or Mboumar-9 transposase is co-transfected with transposon-containing plasmid DNA, it penetrates prokaryotic or eukaryotic cells and integrates the target DNA into the genome. In vivo integrations by purified transposase can be achieved by electroporation, chemical transfection or Lipofection of the transposase:DNA mixture, in contrast to other published transposon-based protocols which require electroporation or microinjection. As in other transposome systems, no helper plasmids are required since transposases are not expressed inside the host cells, thus leading to generation of stable cell lines. Since it does not require electroporation or microinjection, this tool has the potential to be applied for automated high-throughput creation of libraries of random integrants for purposes including gene knock-out libraries, screening for optimal integration positions or safe genome locations in different organisms, selection of the highest production of valuable compounds for biotechnology, and sequencing. PMID:28204586

Combing Chromosomal DNA Mediated by the SMC Complex: Structure and Mechanisms.

PubMed

Kamada, Katsuhiko; Barillà, Daniela

2018-02-01

Genome maintenance requires various nucleoid-associated factors in prokaryotes. Among them, the SMC (Structural Maintenance of Chromosomes) protein has been thought to play a static role in the organization and segregation of the chromosome during cell division. However, recent studies have shown that the bacterial SMC is required to align left and right arms of the emerging chromosome and that the protein dynamically travels from origin to Ter region. A rod form of the SMC complex mediates DNA bridging and has been recognized as a machinery responsible for DNA loop extrusion, like eukaryotic condensin or cohesin complexes, which act as chromosome organizers. Attention is now turning to how the prototype of the complex is loaded on the entry site and translocated on chromosomal DNA, explaining its overall conformational changes at atomic levels. Here, we review and highlight recent findings concerning the prokaryotic SMC complex and discuss possible mechanisms from the viewpoint of protein architecture. © 2017 The Authors. BioEssays Published by WILEY Periodicals, Inc.

Single prokaryotic cell isolation and total transcript amplification protocol for transcriptomic analysis.

PubMed

Kang, Yun; McMillan, Ian; Norris, Michael H; Hoang, Tung T

2015-07-01

Until recently, transcriptome analyses of single cells have been confined to eukaryotes. The information obtained from single-cell transcripts can provide detailed insight into spatiotemporal gene expression, and it could be even more valuable if expanded to prokaryotic cells. Transcriptome analysis of single prokaryotic cells is a recently developed and powerful tool. Here we describe a procedure that allows amplification of the total transcript of a single prokaryotic cell for in-depth analysis. This is performed by using a laser-capture microdissection instrument for single-cell isolation, followed by reverse transcription via Moloney murine leukemia virus, degradation of chromosomal DNA with McrBC and DpnI restriction enzymes, single-stranded cDNA (ss-cDNA) ligation using T4 polynucleotide kinase and CircLigase, and polymerization of ss-cDNA to double-stranded cDNA (ds-cDNA) by Φ29 polymerase. This procedure takes ∼5 d, and sufficient amounts of ds-cDNA can be obtained from single-cell RNA template for further microarray analysis.

Dbl2 Regulates Rad51 and DNA Joint Molecule Metabolism to Ensure Proper Meiotic Chromosome Segregation

PubMed Central

Hyppa, Randy W.; Benko, Zsigmond; Misova, Ivana; Schleiffer, Alexander; Smith, Gerald R.; Gregan, Juraj

2016-01-01

To identify new proteins required for faithful meiotic chromosome segregation, we screened a Schizosaccharomyces pombe deletion mutant library and found that deletion of the dbl2 gene led to missegregation of chromosomes during meiosis. Analyses of both live and fixed cells showed that dbl2Δ mutant cells frequently failed to segregate homologous chromosomes to opposite poles during meiosis I. Removing Rec12 (Spo11 homolog) to eliminate meiotic DNA double-strand breaks (DSBs) suppressed the segregation defect in dbl2Δ cells, indicating that Dbl2 acts after the initiation of meiotic recombination. Analyses of DSBs and Holliday junctions revealed no significant defect in their formation or processing in dbl2Δ mutant cells, although some Rec12-dependent DNA joint molecules persisted late in meiosis. Failure to segregate chromosomes in the absence of Dbl2 correlated with persistent Rad51 foci, and deletion of rad51 or genes encoding Rad51 mediators also suppressed the segregation defect of dbl2Δ. Formation of foci of Fbh1, an F-box helicase that efficiently dismantles Rad51-DNA filaments, was impaired in dbl2Δ cells. Our results suggest that Dbl2 is a novel regulator of Fbh1 and thereby Rad51-dependent DSB repair required for proper meiotic chromosome segregation and viable sex cell formation. The wide conservation of these proteins suggests that our results apply to many species. PMID:27304859

Abundance, diversity and domain architecture variability in prokaryotic DNA-binding transcription factors.

PubMed

Perez-Rueda, Ernesto; Hernandez-Guerrero, Rafael; Martinez-Nuñez, Mario Alberto; Armenta-Medina, Dagoberto; Sanchez, Israel; Ibarra, J Antonio

2018-01-01

Gene regulation at the transcriptional level is a central process in all organisms, and DNA-binding transcription factors, known as TFs, play a fundamental role. This class of proteins usually binds at specific DNA sequences, activating or repressing gene expression. In general, TFs are composed of two domains: the DNA-binding domain (DBD) and an extra domain, which in this work we have named "companion domain" (CD). This latter could be involved in one or more functions such as ligand binding, protein-protein interactions or even with enzymatic activity. In contrast to DBDs, which have been widely characterized both experimentally and bioinformatically, information on the abundance, distribution, variability and possible role of the CDs is scarce. Here, we investigated these issues associated with the domain architectures of TFs in prokaryotic genomes. To this end, 19 families of TFs in 761 non-redundant bacterial and archaeal genomes were evaluated. In this regard we found four main groups based on the abundance and distribution in the analyzed genomes: i) LysR and TetR/AcrR; ii) AraC/XylS, SinR, and others; iii) Lrp, Fis, ArsR, and others; and iv) a group that included only two families, ArgR and BirA. Based on a classification of the organisms according to the life-styles, a major abundance of regulatory families in free-living organisms, in contrast with pathogenic, extremophilic or intracellular organisms, was identified. Finally, the protein architecture diversity associated to the 19 families considering a weight score for domain promiscuity evidenced which regulatory families were characterized by either a large diversity of CDs, here named as "promiscuous" families given the elevated number of variable domains found in those TFs, or a low diversity of CDs. Altogether this information helped us to understand the diversity and distribution of the 19 Prokaryotes TF families. Moreover, initial steps were taken to comprehend the variability of the extra domain

Rapid quantification and taxonomic classification of environmentalDNA from both prokaryotic and eukaryotic origins using a microarray

DOE Office of Scientific and Technical Information (OSTI.GOV)

DeSantis, Todd Z.; Stone, Carol E.; Murray, Sonya R.

2005-02-22

A microarray has been designed using 62,358 probes matched to both prokaryotic and eukaryotic small-subunit ribosomal RNA genes. The array categorized environmental DNA to specific phylogenetic clusters in under 9 h. To a background of DNA generated from natural outdoor aerosols, known quantities of rRNA gene copies from distinct organisms were added producing corresponding hybridization intensity scores that correlated well with their concentrations (r=0.917). Reproducible differences in microbial community composition were observed by altering the genomic DNA extraction method. Notably, gentle extractions produced peak intensities for Mycoplasmatales and Burkholderiales, whereas a vigorous disruption produced peak intensities for Vibrionales,Clostridiales, and Bacillales.

Chromosome segregation in Archaea mediated by a hybrid DNA partition machine

PubMed Central

Kalliomaa-Sanford, Anne K.; Rodriguez-Castañeda, Fernando A.; McLeod, Brett N.; Latorre-Roselló, Victor; Smith, Jasmine H.; Reimann, Julia; Albers, Sonja V.; Barillà, Daniela

2012-01-01

Eukarya and, more recently, some bacteria have been shown to rely on a cytoskeleton-based apparatus to drive chromosome segregation. In contrast, the factors and mechanisms underpinning this fundamental process are underexplored in archaea, the third domain of life. Here we establish that the archaeon Sulfolobus solfataricus harbors a hybrid segrosome consisting of two interacting proteins, SegA and SegB, that play a key role in genome segregation in this organism. SegA is an ortholog of bacterial, Walker-type ParA proteins, whereas SegB is an archaea-specific factor lacking sequence identity to either eukaryotic or bacterial proteins, but sharing homology with a cluster of uncharacterized factors conserved in both crenarchaea and euryarchaea, the two major archaeal sub-phyla. We show that SegA is an ATPase that polymerizes in vitro and that SegB is a site-specific DNA-binding protein contacting palindromic sequences located upstream of the segAB cassette. SegB interacts with SegA in the presence of nucleotides and dramatically affects its polymerization dynamics. Our data demonstrate that SegB strongly stimulates SegA polymerization, possibly by promoting SegA nucleation and accelerating polymer growth. Increased expression levels of segAB resulted in severe growth and chromosome segregation defects, including formation of anucleate cells, compact nucleoids confined to one half of the cell compartment and fragmented nucleoids. The overall picture emerging from our findings indicates that the SegAB complex fulfills a crucial function in chromosome segregation and is the prototype of a DNA partition machine widespread across archaea. PMID:22355141

Chromosome segregation in Archaea mediated by a hybrid DNA partition machine.

PubMed

Kalliomaa-Sanford, Anne K; Rodriguez-Castañeda, Fernando A; McLeod, Brett N; Latorre-Roselló, Victor; Smith, Jasmine H; Reimann, Julia; Albers, Sonja V; Barillà, Daniela

2012-03-06

Eukarya and, more recently, some bacteria have been shown to rely on a cytoskeleton-based apparatus to drive chromosome segregation. In contrast, the factors and mechanisms underpinning this fundamental process are underexplored in archaea, the third domain of life. Here we establish that the archaeon Sulfolobus solfataricus harbors a hybrid segrosome consisting of two interacting proteins, SegA and SegB, that play a key role in genome segregation in this organism. SegA is an ortholog of bacterial, Walker-type ParA proteins, whereas SegB is an archaea-specific factor lacking sequence identity to either eukaryotic or bacterial proteins, but sharing homology with a cluster of uncharacterized factors conserved in both crenarchaea and euryarchaea, the two major archaeal sub-phyla. We show that SegA is an ATPase that polymerizes in vitro and that SegB is a site-specific DNA-binding protein contacting palindromic sequences located upstream of the segAB cassette. SegB interacts with SegA in the presence of nucleotides and dramatically affects its polymerization dynamics. Our data demonstrate that SegB strongly stimulates SegA polymerization, possibly by promoting SegA nucleation and accelerating polymer growth. Increased expression levels of segAB resulted in severe growth and chromosome segregation defects, including formation of anucleate cells, compact nucleoids confined to one half of the cell compartment and fragmented nucleoids. The overall picture emerging from our findings indicates that the SegAB complex fulfills a crucial function in chromosome segregation and is the prototype of a DNA partition machine widespread across archaea.

Specific and non-specific interactions of ParB with DNA: implications for chromosome segregation

PubMed Central

Taylor, James A.; Pastrana, Cesar L.; Butterer, Annika; Pernstich, Christian; Gwynn, Emma J.; Sobott, Frank; Moreno-Herrero, Fernando; Dillingham, Mark S.

2015-01-01

The segregation of many bacterial chromosomes is dependent on the interactions of ParB proteins with centromere-like DNA sequences called parS that are located close to the origin of replication. In this work, we have investigated the binding of Bacillus subtilis ParB to DNA in vitro using a variety of biochemical and biophysical techniques. We observe tight and specific binding of a ParB homodimer to the parS sequence. Binding of ParB to non-specific DNA is more complex and displays apparent positive co-operativity that is associated with the formation of larger, poorly defined, nucleoprotein complexes. Experiments with magnetic tweezers demonstrate that non-specific binding leads to DNA condensation that is reversible by protein unbinding or force. The condensed DNA structure is not well ordered and we infer that it is formed by many looping interactions between neighbouring DNA segments. Consistent with this view, ParB is also able to stabilize writhe in single supercoiled DNA molecules and to bridge segments from two different DNA molecules in trans. The experiments provide no evidence for the promotion of non-specific DNA binding and/or condensation events by the presence of parS sequences. The implications of these observations for chromosome segregation are discussed. PMID:25572315

Occurrence of lignin degradation genotypes and phenotypes among prokaryotes.

PubMed

Tian, Jiang-Hao; Pourcher, Anne-Marie; Bouchez, Théodore; Gelhaye, Eric; Peu, Pascal

2014-12-01

A number of prokaryotes actively contribute to lignin degradation in nature and their activity could be of interest for many applications including the production of biogas/biofuel from lignocellulosic biomass and biopulping. This review compares the reliability and efficiency of the culture-dependent screening methods currently used for the isolation of ligninolytic prokaryotes. Isolated prokaryotes exhibiting lignin-degrading potential are presented according to their phylogenetic groups. With the development of bioinformatics, culture-independent techniques are emerging that allow larger-scale data mining for ligninolytic prokaryotic functions but today, these techniques still have some limits. In this work, two phylogenetic affiliations of isolated prokaryotes exhibiting ligninolytic potential and laccase-encoding prokaryotes were determined on the basis of 16S rDNA sequences, providing a comparative view of results obtained by the two types of screening techniques. The combination of laboratory culture and bioinformatics approaches is a promising way to explore lignin-degrading prokaryotes.

In vivo analysis of polyadenylation in prokaryotes.

PubMed

Mohanty, Bijoy K; Kushner, Sidney R

2014-01-01

Polyadenylation at the 3' ends of mRNAs, tRNAs, rRNAs, and sRNAs plays important roles in RNA metabolism in both prokaryotes and eukaryotes. However, the nature of poly(A) tails in prokaryotes is distinct compared to their eukaryotic counterparts. Specifically, depending on the organism, eukaryotic poly(A) tails average between 50 and >200 nt and can easily be isolated by several techniques involving oligo(dT)-dependent cDNA amplification. In contrast, the bulk of the poly(A) tails present on prokaryotic transcripts is relatively short (<10 nt) and is difficult to characterize using similar techniques. This chapter describes methods that can circumvent these problems. For example, we discuss how to isolate total RNA and characterize its overall polyadenylation status employing a poly(A) sizing assay. Furthermore, we describe a technique involving RNase H treatment of total RNA followed by northern analysis in order to distinguish length of poly(A) tails on various types of transcripts. Finally, we outline a useful procedure to clone the poly(A) tails of specific transcripts using 5'-3' end-ligated RNA, which is independent of oligo(dT)-dependent cDNA amplification. These approaches are particularly helpful in analyzing transcripts with either short or long poly(A) tails both in prokaryotes and eukaryotes.

Treadmilling of a prokaryotic tubulin-like protein, TubZ, required for plasmid stability in Bacillus thuringiensis

PubMed Central

Larsen, Rachel A.; Cusumano, Christina; Fujioka, Akina; Lim-Fong, Grace; Patterson, Paula; Pogliano, Joe

2007-01-01

Prokaryotes rely on a distant tubulin homolog, FtsZ, for assembling the cytokinetic ring essential for cell division, but are otherwise generally thought to lack tubulin-like polymers that participate in processes such as DNA segregation. Here we characterize a protein (TubZ) from the Bacillus thuringiensis virulence plasmid pBtoxis, which is a member of the tubulin/FtsZ GTPase superfamily but is only distantly related to both FtsZ and tubulin. TubZ assembles dynamic, linear polymers that exhibit directional polymerization with plus and minus ends, movement by treadmilling, and a critical concentration for assembly. A point mutation (D269A) that alters a highly conserved catalytic residue within the T7 loop completely eliminates treadmilling and allows the formation of stable polymers at a much lower protein concentration than the wild-type protein. When expressed in trans, TubZ(D269A) coassembles with wild-type TubZ and significantly reduces the stability of pBtoxis, demonstrating a direct correlation between TubZ dynamics and plasmid maintenance. The tubZ gene is in an operon with tubR, which encodes a putative DNA-binding protein that regulates TubZ levels. Our results suggest that TubZ is representative of a novel class of prokaryotic cytoskeletal proteins important for plasmid stability that diverged long ago from the ancient tubulin/FtsZ ancestor. PMID:17510284

Analysis of T-DNA integration and generative segregation in transgenic winter triticale (x Triticosecale Wittmack)

PubMed Central

2012-01-01

Background While the genetic transformation of the major cereal crops has become relatively routine, to date only a few reports were published on transgenic triticale, and robust data on T-DNA integration and segregation have not been available in this species. Results Here, we present a comprehensive analysis of stable transgenic winter triticale cv. Bogo carrying the selectable marker gene HYGROMYCIN PHOSPHOTRANSFERASE (HPT) and a synthetic green fluorescent protein gene (gfp). Progeny of four independent transgenic plants were comprehensively investigated with regard to the number of integrated T-DNA copies, the number of plant genomic integration loci, the integrity and functionality of individual T-DNA copies, as well as the segregation of transgenes in T1 and T2 generations, which also enabled us to identify homozygous transgenic lines. The truncation of some integrated T-DNAs at their left end along with the occurrence of independent segregation of multiple T-DNAs unintendedly resulted in a single-copy segregant that is selectable marker-free and homozygous for the gfp gene. The heritable expression of gfp driven by the maize UBI-1 promoter was demonstrated by confocal laser scanning microscopy. Conclusions The used transformation method is a valuable tool for the genetic engineering of triticale. Here we show that comprehensive molecular analyses are required for the correct interpretation of phenotypic data collected from the transgenic plants. PMID:23006412

Analysis of T-DNA integration and generative segregation in transgenic winter triticale (x Triticosecale Wittmack).

PubMed

Hensel, Goetz; Oleszczuk, Sylwia; Daghma, Diaa Eldin S; Zimny, Janusz; Melzer, Michael; Kumlehn, Jochen

2012-09-25

While the genetic transformation of the major cereal crops has become relatively routine, to date only a few reports were published on transgenic triticale, and robust data on T-DNA integration and segregation have not been available in this species. Here, we present a comprehensive analysis of stable transgenic winter triticale cv. Bogo carrying the selectable marker gene HYGROMYCIN PHOSPHOTRANSFERASE (HPT) and a synthetic green fluorescent protein gene (gfp). Progeny of four independent transgenic plants were comprehensively investigated with regard to the number of integrated T-DNA copies, the number of plant genomic integration loci, the integrity and functionality of individual T-DNA copies, as well as the segregation of transgenes in T1 and T2 generations, which also enabled us to identify homozygous transgenic lines. The truncation of some integrated T-DNAs at their left end along with the occurrence of independent segregation of multiple T-DNAs unintendedly resulted in a single-copy segregant that is selectable marker-free and homozygous for the gfp gene. The heritable expression of gfp driven by the maize UBI-1 promoter was demonstrated by confocal laser scanning microscopy. The used transformation method is a valuable tool for the genetic engineering of triticale. Here we show that comprehensive molecular analyses are required for the correct interpretation of phenotypic data collected from the transgenic plants.

Keeping the Wolves at Bay: Antitoxins of Prokaryotic Type II Toxin-Antitoxin Systems.

PubMed

Chan, Wai Ting; Espinosa, Manuel; Yeo, Chew Chieng

2016-01-01

In their initial stages of discovery, prokaryotic toxin-antitoxin (TA) systems were confined to bacterial plasmids where they function to mediate the maintenance and stability of usually low- to medium-copy number plasmids through the post-segregational killing of any plasmid-free daughter cells that developed. Their eventual discovery as nearly ubiquitous and repetitive elements in bacterial chromosomes led to a wealth of knowledge and scientific debate as to their diversity and functionality in the prokaryotic lifestyle. Currently categorized into six different types designated types I-VI, type II TA systems are the best characterized. These generally comprised of two genes encoding a proteic toxin and its corresponding proteic antitoxin, respectively. Under normal growth conditions, the stable toxin is prevented from exerting its lethal effect through tight binding with the less stable antitoxin partner, forming a non-lethal TA protein complex. Besides binding with its cognate toxin, the antitoxin also plays a role in regulating the expression of the type II TA operon by binding to the operator site, thereby repressing transcription from the TA promoter. In most cases, full repression is observed in the presence of the TA complex as binding of the toxin enhances the DNA binding capability of the antitoxin. TA systems have been implicated in a gamut of prokaryotic cellular functions such as being mediators of programmed cell death as well as persistence or dormancy, biofilm formation, as defensive weapons against bacteriophage infections and as virulence factors in pathogenic bacteria. It is thus apparent that these antitoxins, as DNA-binding proteins, play an essential role in modulating the prokaryotic lifestyle whilst at the same time preventing the lethal action of the toxins under normal growth conditions, i.e., keeping the proverbial wolves at bay. In this review, we will cover the diversity and characteristics of various type II TA antitoxins. We shall

Keeping the Wolves at Bay: Antitoxins of Prokaryotic Type II Toxin-Antitoxin Systems

PubMed Central

Chan, Wai Ting; Espinosa, Manuel; Yeo, Chew Chieng

2016-01-01

In their initial stages of discovery, prokaryotic toxin-antitoxin (TA) systems were confined to bacterial plasmids where they function to mediate the maintenance and stability of usually low- to medium-copy number plasmids through the post-segregational killing of any plasmid-free daughter cells that developed. Their eventual discovery as nearly ubiquitous and repetitive elements in bacterial chromosomes led to a wealth of knowledge and scientific debate as to their diversity and functionality in the prokaryotic lifestyle. Currently categorized into six different types designated types I–VI, type II TA systems are the best characterized. These generally comprised of two genes encoding a proteic toxin and its corresponding proteic antitoxin, respectively. Under normal growth conditions, the stable toxin is prevented from exerting its lethal effect through tight binding with the less stable antitoxin partner, forming a non-lethal TA protein complex. Besides binding with its cognate toxin, the antitoxin also plays a role in regulating the expression of the type II TA operon by binding to the operator site, thereby repressing transcription from the TA promoter. In most cases, full repression is observed in the presence of the TA complex as binding of the toxin enhances the DNA binding capability of the antitoxin. TA systems have been implicated in a gamut of prokaryotic cellular functions such as being mediators of programmed cell death as well as persistence or dormancy, biofilm formation, as defensive weapons against bacteriophage infections and as virulence factors in pathogenic bacteria. It is thus apparent that these antitoxins, as DNA-binding proteins, play an essential role in modulating the prokaryotic lifestyle whilst at the same time preventing the lethal action of the toxins under normal growth conditions, i.e., keeping the proverbial wolves at bay. In this review, we will cover the diversity and characteristics of various type II TA antitoxins. We shall

The prokaryotic enhancer binding protein NTRC has an ATPase activity which is phosphorylation and DNA dependent.

PubMed Central

Austin, S; Dixon, R

1992-01-01

The prokaryotic activator protein NTRC binds to enhancer-like elements and activates transcription in response to nitrogen limitation by catalysing open complex formation by sigma 54 RNA polymerase holoenzyme. Formation of open complexes requires the phosphorylated form of NTRC and the reaction is ATP dependent. We find that NTRC has an ATPase activity which is activated by phosphorylation and is strongly stimulated by the presence of DNA containing specific NTRC binding sites. Images PMID:1534752

Segregation of mtDNA Throughout Human Embryofetal Development: m.3243A > G as a Model System

PubMed Central

Monnot, Sophie; Gigarel, Nadine; Samuels, David C; Burlet, Philippe; Hesters, Laetitia; Frydman, Nelly; Frydman, René; Kerbrat, Violaine; Funalot, Benoit; Martinovic, Jelena; Benachi, Alexandra; Feingold, Josué; Munnich, Arnold; Bonnefont, Jean-Paul; Steffann, Julie

2011-01-01

Mitochondrial DNA (mtDNA) mutations cause a wide range of serious diseases with high transmission risk and maternal inheritance. Tissue heterogeneity of the heteroplasmy rate (“mutant load”) accounts for the wide phenotypic spectrum observed in carriers. Owing to the absence of therapy, couples at risk to transmit such disorders commonly ask for prenatal (PND) or preimplantation diagnosis (PGD). The lack of data regarding heteroplasmy distribution throughout intrauterine development, however, hampers the implementation of such procedures. We tracked the segregation of the m.3243A > G mutation (MT-TL1 gene) responsible for the MELAS syndrome in the developing embryo/fetus, using tissues and cells from eight carrier females, their 38 embryos and 12 fetuses. Mutant mtDNA segregation was found to be governed by random genetic drift, during oogenesis and somatic tissue development. The size of the bottleneck operating for m.3243A > G during oogenesis was shown to be individual-dependent. Comparison with data we achieved for the m.8993T > G mutation (MT-ATP6 gene), responsible for the NARP/Leigh syndrome, indicates that these mutations differentially influence mtDNA segregation during oogenesis, while their impact is similar in developing somatic tissues. These data have major consequences for PND and PGD procedures in mtDNA inherited disorders. Hum Mutat 32:116–125, 2011. © 2010 Wiley-Liss, Inc. PMID:21120938

Viral Diversity Threshold for Adaptive Immunity in Prokaryotes

PubMed Central

Weinberger, Ariel D.; Wolf, Yuri I.; Lobkovsky, Alexander E.; Gilmore, Michael S.; Koonin, Eugene V.

2012-01-01

ABSTRACT Bacteria and archaea face continual onslaughts of rapidly diversifying viruses and plasmids. Many prokaryotes maintain adaptive immune systems known as clustered regularly interspaced short palindromic repeats (CRISPR) and CRISPR-associated genes (Cas). CRISPR-Cas systems are genomic sensors that serially acquire viral and plasmid DNA fragments (spacers) that are utilized to target and cleave matching viral and plasmid DNA in subsequent genomic invasions, offering critical immunological memory. Only 50% of sequenced bacteria possess CRISPR-Cas immunity, in contrast to over 90% of sequenced archaea. To probe why half of bacteria lack CRISPR-Cas immunity, we combined comparative genomics and mathematical modeling. Analysis of hundreds of diverse prokaryotic genomes shows that CRISPR-Cas systems are substantially more prevalent in thermophiles than in mesophiles. With sequenced bacteria disproportionately mesophilic and sequenced archaea mostly thermophilic, the presence of CRISPR-Cas appears to depend more on environmental temperature than on bacterial-archaeal taxonomy. Mutation rates are typically severalfold higher in mesophilic prokaryotes than in thermophilic prokaryotes. To quantitatively test whether accelerated viral mutation leads microbes to lose CRISPR-Cas systems, we developed a stochastic model of virus-CRISPR coevolution. The model competes CRISPR-Cas-positive (CRISPR-Cas+) prokaryotes against CRISPR-Cas-negative (CRISPR-Cas−) prokaryotes, continually weighing the antiviral benefits conferred by CRISPR-Cas immunity against its fitness costs. Tracking this cost-benefit analysis across parameter space reveals viral mutation rate thresholds beyond which CRISPR-Cas cannot provide sufficient immunity and is purged from host populations. These results offer a simple, testable viral diversity hypothesis to explain why mesophilic bacteria disproportionately lack CRISPR-Cas immunity. More generally, fundamental limits on the adaptability of biological

Assembling the bacterial segrosome.

PubMed

Hayes, Finbarr; Barillà, Daniela

2006-05-01

Genome segregation in prokaryotes is a highly ordered process that integrates with DNA replication, cytokinesis and other fundamental facets of the bacterial cell cycle. The segrosome is the nucleoprotein complex that mediates DNA segregation in bacteria, its assembly and organization is best understood for plasmid partition. The recent elucidation of structures of the ParB plasmid segregation protein bound to centromeric DNA, and of the tertiary structures of other segregation proteins, are key milestones in the path to deciphering the molecular basis of bacterial DNA segregation.

DNA and origin region segregation are not affected by the transition from rod to sphere after inhibition of Escherichia coli MreB by A22.

PubMed

Karczmarek, Aneta; Martínez-Arteaga, Rocío; Baselga, Rocío Martínez-Arteaga; Alexeeva, Svetlana; Hansen, Flemming G; Vicente, Miguel; Nanninga, Nanne; den Blaauwen, Tanneke

2007-07-01

The bacterial actin homologue MreB forms a helix underneath the cytoplasmic membrane and was shown to be essential in the morphogenesis of the rod-shaped cells. Additionally, MreB was implicated to be involved in DNA segregation. However, in our hands the mreBCD deletion strain (PA340-678) grew without apparent DNA segregation defect, suggesting that the reported chromosome segregation inhibition could be caused by a temporarily effect of MreB inhibition or depletion. To assess the involvement of MreB in DNA segregation during the transition from rod to sphere, we compared the effect of A22 and the PBP2 inhibitor mecillinam on the percentage of cells with segregated nucleoids and the number of oriC foci in wild-type Escherichia coli cells. Cells became spherical in the same time window during both treatments and we could not detect any difference in the chromosome or oriC segregation between these two treatments. Additionally, flow cytometric analyses showed that A22 and mecillinam treatment gave essentially the same chromosome segregation pattern. We conclude that MreB is not directly involved in DNA segregation of E. coli.

Deep sub-seafloor prokaryotes stimulated at interfaces over geological time.

PubMed

Parkes, R John; Webster, Gordon; Cragg, Barry A; Weightman, Andrew J; Newberry, Carole J; Ferdelman, Timothy G; Kallmeyer, Jens; Jørgensen, Bo B; Aiello, Ivano W; Fry, John C

2005-07-21

The sub-seafloor biosphere is the largest prokaryotic habitat on Earth but also a habitat with the lowest metabolic rates. Modelled activity rates are very low, indicating that most prokaryotes may be inactive or have extraordinarily slow metabolism. Here we present results from two Pacific Ocean sites, margin and open ocean, both of which have deep, subsurface stimulation of prokaryotic processes associated with geochemical and/or sedimentary interfaces. At 90 m depth in the margin site, stimulation was such that prokaryote numbers were higher (about 13-fold) and activity rates higher than or similar to near-surface values. Analysis of high-molecular-mass DNA confirmed the presence of viable prokaryotes and showed changes in biodiversity with depth that were coupled to geochemistry, including a marked community change at the 90-m interface. At the open ocean site, increases in numbers of prokaryotes at depth were more restricted but also corresponded to increased activity; however, this time they were associated with repeating layers of diatom-rich sediments (about 9 Myr old). These results show that deep sedimentary prokaryotes can have high activity, have changing diversity associated with interfaces and are active over geological timescales.

A septal chromosome segregator protein evolved into a conjugative DNA-translocator protein

PubMed Central

Sepulveda, Edgardo; Vogelmann, Jutta

2011-01-01

Streptomycetes, Gram-positive soil bacteria well known for the production of antibiotics feature a unique conjugative DNA transfer system. In contrast to classical conjugation which is characterized by the secretion of a pilot protein covalently linked to a single-stranded DNA molecule, in Streptomyces a double-stranded DNA molecule is translocated during conjugative transfer. This transfer involves a single plasmid encoded protein, TraB. A detailed biochemical and biophysical characterization of TraB, revealed a close relationship to FtsK, mediating chromosome segregation during bacterial cell division. TraB translocates plasmid DNA by recognizing 8-bp direct repeats located in a specific plasmid region clt. Similar sequences accidentally also occur on chromosomes and have been shown to be bound by TraB. We suggest that TraB mobilizes chromosomal genes by the interaction with these chromosomal clt-like sequences not relying on the integration of the conjugative plasmid into the chromosome. PMID:22479692

Development of a Prokaryotic Universal Primer for Simultaneous Analysis of Bacteria and Archaea Using Next-Generation Sequencing

PubMed Central

Takahashi, Shunsuke; Tomita, Junko; Nishioka, Kaori; Hisada, Takayoshi; Nishijima, Miyuki

2014-01-01

For the analysis of microbial community structure based on 16S rDNA sequence diversity, sensitive and robust PCR amplification of 16S rDNA is a critical step. To obtain accurate microbial composition data, PCR amplification must be free of bias; however, amplifying all 16S rDNA species with equal efficiency from a sample containing a large variety of microorganisms remains challenging. Here, we designed a universal primer based on the V3-V4 hypervariable region of prokaryotic 16S rDNA for the simultaneous detection of Bacteria and Archaea in fecal samples from crossbred pigs (Landrace×Large white×Duroc) using an Illumina MiSeq next-generation sequencer. In-silico analysis showed that the newly designed universal prokaryotic primers matched approximately 98.0% of Bacteria and 94.6% of Archaea rRNA gene sequences in the Ribosomal Database Project database. For each sequencing reaction performed with the prokaryotic universal primer, an average of 69,330 (±20,482) reads were obtained, of which archaeal rRNA genes comprised approximately 1.2% to 3.2% of all prokaryotic reads. In addition, the detection frequency of Bacteria belonging to the phylum Verrucomicrobia, including members of the classes Verrucomicrobiae and Opitutae, was higher in the NGS analysis using the prokaryotic universal primer than that performed with the bacterial universal primer. Importantly, this new prokaryotic universal primer set had markedly lower bias than that of most previously designed universal primers. Our findings demonstrate that the prokaryotic universal primer set designed in the present study will permit the simultaneous detection of Bacteria and Archaea, and will therefore allow for a more comprehensive understanding of microbial community structures in environmental samples. PMID:25144201

Genomics of Bacterial and Archaeal Viruses: Dynamics within the Prokaryotic Virosphere

PubMed Central

Krupovic, Mart; Prangishvili, David; Hendrix, Roger W.; Bamford, Dennis H.

2011-01-01

Summary: Prokaryotes, bacteria and archaea, are the most abundant cellular organisms among those sharing the planet Earth with human beings (among others). However, numerous ecological studies have revealed that it is actually prokaryotic viruses that predominate on our planet and outnumber their hosts by at least an order of magnitude. An understanding of how this viral domain is organized and what are the mechanisms governing its evolution is therefore of great interest and importance. The vast majority of characterized prokaryotic viruses belong to the order Caudovirales, double-stranded DNA (dsDNA) bacteriophages with tails. Consequently, these viruses have been studied (and reviewed) extensively from both genomic and functional perspectives. However, albeit numerous, tailed phages represent only a minor fraction of the prokaryotic virus diversity. Therefore, the knowledge which has been generated for this viral system does not offer a comprehensive view of the prokaryotic virosphere. In this review, we discuss all families of bacterial and archaeal viruses that contain more than one characterized member and for which evolutionary conclusions can be attempted by use of comparative genomic analysis. We focus on the molecular mechanisms of their genome evolution as well as on the relationships between different viral groups and plasmids. It becomes clear that evolutionary mechanisms shaping the genomes of prokaryotic viruses vary between different families and depend on the type of the nucleic acid, characteristics of the virion structure, as well as the mode of the life cycle. We also point out that horizontal gene transfer is not equally prevalent in different virus families and is not uniformly unrestricted for diverse viral functions. PMID:22126996

Comparison of Methods of Detection of Exceptional Sequences in Prokaryotic Genomes.

PubMed

Rusinov, I S; Ershova, A S; Karyagina, A S; Spirin, S A; Alexeevski, A V

2018-02-01

Many proteins need recognition of specific DNA sequences for functioning. The number of recognition sites and their distribution along the DNA might be of biological importance. For example, the number of restriction sites is often reduced in prokaryotic and phage genomes to decrease the probability of DNA cleavage by restriction endonucleases. We call a sequence an exceptional one if its frequency in a genome significantly differs from one predicted by some mathematical model. An exceptional sequence could be either under- or over-represented, depending on its frequency in comparison with the predicted one. Exceptional sequences could be considered biologically meaningful, for example, as targets of DNA-binding proteins or as parts of abundant repetitive elements. Several methods to predict frequency of a short sequence in a genome, based on actual frequencies of certain its subsequences, are used. The most popular are methods based on Markov chain models. But any rigorous comparison of the methods has not previously been performed. We compared three methods for the prediction of short sequence frequencies: the maximum-order Markov chain model-based method, the method that uses geometric mean of extended Markovian estimates, and the method that utilizes frequencies of all subsequences including discontiguous ones. We applied them to restriction sites in complete genomes of 2500 prokaryotic species and demonstrated that the results depend greatly on the method used: lists of 5% of the most under-represented sites differed by up to 50%. The method designed by Burge and coauthors in 1992, which utilizes all subsequences of the sequence, showed a higher precision than the other two methods both on prokaryotic genomes and randomly generated sequences after computational imitation of selective pressure. We propose this method as the first choice for detection of exceptional sequences in prokaryotic genomes.

Condensin suppresses recombination and regulates double-strand break processing at the repetitive ribosomal DNA array to ensure proper chromosome segregation during meiosis in budding yeast

PubMed Central

Li, Ping; Jin, Hui; Yu, Hong-Guo

2014-01-01

During meiosis, homologues are linked by crossover, which is required for bipolar chromosome orientation before chromosome segregation at anaphase I. The repetitive ribosomal DNA (rDNA) array, however, undergoes little or no meiotic recombination. Hyperrecombination can cause chromosome missegregation and rDNA copy number instability. We report here that condensin, a conserved protein complex required for chromosome organization, regulates double-strand break (DSB) formation and repair at the rDNA gene cluster during meiosis in budding yeast. Condensin is highly enriched at the rDNA region during prophase I, released at the prophase I/metaphase I transition, and reassociates with rDNA before anaphase I onset. We show that condensin plays a dual role in maintaining rDNA stability: it suppresses the formation of Spo11-mediated rDNA breaks, and it promotes DSB processing to ensure proper chromosome segregation. Condensin is unnecessary for the export of rDNA breaks outside the nucleolus but required for timely repair of meiotic DSBs. Our work reveals that condensin coordinates meiotic recombination with chromosome segregation at the repetitive rDNA sequence, thereby maintaining genome integrity. PMID:25103240

BubR1- and Polo-Coated DNA Tethers Facilitate Poleward Segregation of Acentric Chromatids

PubMed Central

Royou, Anne; Gagou, Mary E.; Karess, Roger; Sullivan, William

2010-01-01

Summary The mechanisms that safeguard cells against chromosomal instability (CIN) are of great interest, as CIN contributes to tumorigenesis. To gain insight into these mechanisms, we studied the behavior of cells entering mitosis with damaged chromosomes. We used the endonuclease I-CreI to generate acentric chromosomes in Drosophila larvae. While I-CreI expression produces acentric chromosomes in the majority of neuronal stem cells, remarkably, it has no effect on adult survival. Our live studies reveal that acentric chromatids segregate efficiently to opposite poles. The acentric chromatid poleward movement is mediated through DNA tethers decorated with BubR1, Polo, INCENP, and Aurora-B. Reduced BubR1 or Polo function results in abnormal segregation of acentric chromatids, a decrease in acentric chromosome tethering, and a great reduction in adult survival. We propose that BubR1 and Polo facilitate the accurate segregation of acentric chromatids by maintaining the integrity of the tethers that connect acentric chromosomes to their centric partners. PMID:20141837

Template DNA-strand co-segregation and asymmetric cell division in skeletal muscle stem cells.

PubMed

Shinin, Vasily; Gayraud-Morel, Barbara; Tajbakhsh, Shahragim

2009-01-01

Stem cells are present in all tissues and organs, and are crucial for normal regulated growth. How the pool size of stem cells and their progeny is regulated to establish the tissue prenatally, then maintain it throughout life, is a key question in biology and medicine. The ability to precisely locate stem and progenitors requires defining lineage progression from stem to differentiated cells, assessing the mode of cell expansion and self-renewal and identifying markers to assess the different cell states within the lineage. We have shown that during lineage progression from a quiescent adult muscle satellite cell to a differentiated myofibre, both symmetric and asymmetric divisions take place. Furthermore, we provide evidence that a sub-population of label retaining satellite cells co-segregate template DNA strands to one daughter cell. These findings provide a means of identifying presumed stem and progenitor cells within the lineage. In addition, asymmetric segregation of template DNA and the cytoplasmic protein Numb provides a landmark to define cell behaviour as self-renewal and differentiation decisions are being executed.

Multi-scale Modeling of Chromosomal DNA in Living Cells

NASA Astrophysics Data System (ADS)

Spakowitz, Andrew

The organization and dynamics of chromosomal DNA play a pivotal role in a range of biological processes, including gene regulation, homologous recombination, replication, and segregation. Establishing a quantitative theoretical model of DNA organization and dynamics would be valuable in bridging the gap between the molecular-level packaging of DNA and genome-scale chromosomal processes. Our research group utilizes analytical theory and computational modeling to establish a predictive theoretical model of chromosomal organization and dynamics. In this talk, I will discuss our efforts to develop multi-scale polymer models of chromosomal DNA that are both sufficiently detailed to address specific protein-DNA interactions while capturing experimentally relevant time and length scales. I will demonstrate how these modeling efforts are capable of quantitatively capturing aspects of behavior of chromosomal DNA in both prokaryotic and eukaryotic cells. This talk will illustrate that capturing dynamical behavior of chromosomal DNA at various length scales necessitates a range of theoretical treatments that accommodate the critical physical contributions that are relevant to in vivo behavior at these disparate length and time scales. National Science Foundation, Physics of Living Systems Program (PHY-1305516).

Argonaute Proteins and Mechanisms of RNA Interference in Eukaryotes and Prokaryotes.

PubMed

Olina, A V; Kulbachinskiy, A V; Aravin, A A; Esyunina, D M

2018-05-01

Noncoding RNAs play essential roles in genetic regulation in all organisms. In eukaryotic cells, many small noncoding RNAs act in complex with Argonaute proteins and regulate gene expression by recognizing complementary RNA targets. The complexes of Argonaute proteins with small RNAs also play a key role in silencing of mobile genetic elements and, in some cases, viruses. These processes are collectively called RNA interference. RNA interference is a powerful tool for specific gene silencing in both basic research and therapeutic applications. Argonaute proteins are also found in prokaryotic organisms. Recent studies have shown that prokaryotic Argonautes can also cleave their target nucleic acids, in particular DNA. This activity of prokaryotic Argonautes might potentially be used to edit eukaryotic genomes. However, the molecular mechanisms of small nucleic acid biogenesis and the functions of Argonaute proteins, in particular in bacteria and archaea, remain largely unknown. Here we briefly review available data on the RNA interference processes and Argonaute proteins in eukaryotes and prokaryotes.

Clustered regularly interspaced short palindromic repeats (CRISPRs): the hallmark of an ingenious antiviral defense mechanism in prokaryotes.

PubMed

Al-Attar, Sinan; Westra, Edze R; van der Oost, John; Brouns, Stan J J

2011-04-01

Many prokaryotes contain the recently discovered defense system against mobile genetic elements. This defense system contains a unique type of repetitive DNA stretches, termed Clustered Regularly Interspaced Short Palindromic Repeats (CRISPRs). CRISPRs consist of identical repeated DNA sequences (repeats), interspaced by highly variable sequences referred to as spacers. The spacers originate from either phages or plasmids and comprise the prokaryotes' 'immunological memory'. CRISPR-associated (cas) genes encode conserved proteins that together with CRISPRs make-up the CRISPR/Cas system, responsible for defending the prokaryotic cell against invaders. CRISPR-mediated resistance has been proposed to involve three stages: (i) CRISPR-Adaptation, the invader DNA is encountered by the CRISPR/Cas machinery and an invader-derived short DNA fragment is incorporated in the CRISPR array. (ii) CRISPR-Expression, the CRISPR array is transcribed and the transcript is processed by Cas proteins. (iii) CRISPR-Interference, the invaders' nucleic acid is recognized by complementarity to the crRNA and neutralized. An application of the CRISPR/Cas system is the immunization of industry-relevant prokaryotes (or eukaryotes) against mobile-genetic invasion. In addition, the high variability of the CRISPR spacer content can be exploited for phylogenetic and evolutionary studies. Despite impressive progress during the last couple of years, the elucidation of several fundamental details will be a major challenge in future research.

Condensin suppresses recombination and regulates double-strand break processing at the repetitive ribosomal DNA array to ensure proper chromosome segregation during meiosis in budding yeast.

PubMed

Li, Ping; Jin, Hui; Yu, Hong-Guo

2014-10-01

During meiosis, homologues are linked by crossover, which is required for bipolar chromosome orientation before chromosome segregation at anaphase I. The repetitive ribosomal DNA (rDNA) array, however, undergoes little or no meiotic recombination. Hyperrecombination can cause chromosome missegregation and rDNA copy number instability. We report here that condensin, a conserved protein complex required for chromosome organization, regulates double-strand break (DSB) formation and repair at the rDNA gene cluster during meiosis in budding yeast. Condensin is highly enriched at the rDNA region during prophase I, released at the prophase I/metaphase I transition, and reassociates with rDNA before anaphase I onset. We show that condensin plays a dual role in maintaining rDNA stability: it suppresses the formation of Spo11-mediated rDNA breaks, and it promotes DSB processing to ensure proper chromosome segregation. Condensin is unnecessary for the export of rDNA breaks outside the nucleolus but required for timely repair of meiotic DSBs. Our work reveals that condensin coordinates meiotic recombination with chromosome segregation at the repetitive rDNA sequence, thereby maintaining genome integrity. © 2014 Li et al. This article is distributed by The American Society for Cell Biology under license from the author(s). Two months after publication it is available to the public under an Attribution–Noncommercial–Share Alike 3.0 Unported Creative Commons License (http://creativecommons.org/licenses/by-nc-sa/3.0).

Theory of prokaryotic genome evolution.

PubMed

Sela, Itamar; Wolf, Yuri I; Koonin, Eugene V

2016-10-11

Bacteria and archaea typically possess small genomes that are tightly packed with protein-coding genes. The compactness of prokaryotic genomes is commonly perceived as evidence of adaptive genome streamlining caused by strong purifying selection in large microbial populations. In such populations, even the small cost incurred by nonfunctional DNA because of extra energy and time expenditure is thought to be sufficient for this extra genetic material to be eliminated by selection. However, contrary to the predictions of this model, there exists a consistent, positive correlation between the strength of selection at the protein sequence level, measured as the ratio of nonsynonymous to synonymous substitution rates, and microbial genome size. Here, by fitting the genome size distributions in multiple groups of prokaryotes to predictions of mathematical models of population evolution, we show that only models in which acquisition of additional genes is, on average, slightly beneficial yield a good fit to genomic data. These results suggest that the number of genes in prokaryotic genomes reflects the equilibrium between the benefit of additional genes that diminishes as the genome grows and deletion bias (i.e., the rate of deletion of genetic material being slightly greater than the rate of acquisition). Thus, new genes acquired by microbial genomes, on average, appear to be adaptive. The tight spacing of protein-coding genes likely results from a combination of the deletion bias and purifying selection that efficiently eliminates nonfunctional, noncoding sequences.

Survival and Evolution of CRISPR–Cas System in Prokaryotes and Its Applications

PubMed Central

Shabbir, Muhammad Abu Bakr; Hao, Haihong; Shabbir, Muhammad Zubair; Hussain, Hafiz Iftikhar; Iqbal, Zahid; Ahmed, Saeed; Sattar, Adeel; Iqbal, Mujahid; Li, Jun; Yuan, Zonghui

2016-01-01

Prokaryotes have developed numerous innate immune mechanisms in order to fend off bacteriophage or plasmid attack. One of these immune systems is clustered regularly interspaced short palindromic repeats (CRISPR). CRISPR-associated proteins play a key role in survival of prokaryotes against invaders, as these systems cleave DNA of foreign genetic elements. Beyond providing immunity, these systems have significant impact in altering the bacterial physiology in term of its virulence and pathogenicity, as well as evolution. Also, due to their diverse nature of functionality, cas9 endoribonuclease can be easily reprogrammed with the help of guide RNAs, showing unprecedented potential and significance for gene editing in treating genetic diseases. Here, we also discuss the use of NgAgo–gDNA system in genome editing of human cells. PMID:27725818

Detecting the borders between coding and non-coding DNA regions in prokaryotes based on recursive segmentation and nucleotide doublets statistics

PubMed Central

2012-01-01

Background Detecting the borders between coding and non-coding regions is an essential step in the genome annotation. And information entropy measures are useful for describing the signals in genome sequence. However, the accuracies of previous methods of finding borders based on entropy segmentation method still need to be improved. Methods In this study, we first applied a new recursive entropic segmentation method on DNA sequences to get preliminary significant cuts. A 22-symbol alphabet is used to capture the differential composition of nucleotide doublets and stop codon patterns along three phases in both DNA strands. This process requires no prior training datasets. Results Comparing with the previous segmentation methods, the experimental results on three bacteria genomes, Rickettsia prowazekii, Borrelia burgdorferi and E.coli, show that our approach improves the accuracy for finding the borders between coding and non-coding regions in DNA sequences. Conclusions This paper presents a new segmentation method in prokaryotes based on Jensen-Rényi divergence with a 22-symbol alphabet. For three bacteria genomes, comparing to A12_JR method, our method raised the accuracy of finding the borders between protein coding and non-coding regions in DNA sequences. PMID:23282225

Bacterial chromosome organization and segregation

PubMed Central

Badrinarayanan, Anjana; Le, Tung BK; Laub, Michael T

2016-01-01

If fully stretched out, a typical bacterial chromosome would be nearly one millimeter long, or approximately 1000 times the length of a cell. Not only must cells massively compact their genetic material, but they must also organize their DNA in a manner that is compatible with a range of cellular processes, including DNA replication, DNA repair, homologous recombination, and horizontal gene transfer. Recent work, driven in part by technological advances, has begun to reveal the general principles of chromosome organization in bacteria. Here, drawing on studies of many different organisms, we review the emerging picture of how bacterial chromosomes are structured at multiple length-scales, highlighting the functions of various DNA-binding proteins and impact of physical forces. Additionally, we discuss the spatial dynamics of chromosomes, particularly during their segregation to daughter cells. Although there has been tremendous progress, we also highlight gaps that remain in understanding chromosome organization and segregation. PMID:26566111

[Prokaryotic expression systems].

PubMed

Porowińska, Dorota; Wujak, Magdalena; Roszek, Katarzyna; Komoszyński, Michał

2013-03-01

For overproduction of recombinant proteins both eukaryotic and prokaryotic expression systems are used. Choosing the right system depends, among other things, on the growth rate and culture of host cells, level of the target gene expression and posttranslational processing of the synthesized protein. Regardless of the type of expression system, its basic elements are the vector and the expression host. The most widely used system for protein overproduction, both on a laboratory and industrial scale, is the prokaryotic system. This system is based primarily on the bacteria E. coli, although increasingly often Bacillus species are used. The prokaryotic system allows one to obtain large quantities of recombinant proteins in a short time. A simple and inexpensive bacterial cell culture and well-known mechanisms of transcription and translation facilitate the use of these microorganisms. The simplicity of genetic modifications and the availability of many bacterial mutants are additional advantages of the prokaryotic system. In this article we characterize the structural elements of prokaryotic expression vectors. Also strategies for preparation of the target protein gene that increase productivity, facilitate detection and purification of recombinant protein and provide its activity are discussed. Bacterial strains often used as host cells in expression systems as well as the potential location of heterologous proteins are characterized. Knowledge of the basic elements of the prokaryotic expression system allows for production of biologically active proteins in a short time and in satisfactory quantities. 

Duplication and segregation of the actin (MreB) cytoskeleton during the prokaryotic cell cycle.

PubMed

Vats, Purva; Rothfield, Lawrence

2007-11-06

The bacterial actin homolog MreB exists as a single-copy helical cytoskeletal structure that extends between the two poles of rod-shaped bacteria. In this study, we show that equipartition of the MreB cytoskeleton into daughter cells is accomplished by division and segregation of the helical MreB array into two equivalent structures located in opposite halves of the predivisional cell. This process ensures that each daughter cell inherits one copy of the MreB cytoskeleton. The process is triggered by the membrane association of the FtsZ cell division protein. The cytoskeletal division and segregation events occur before and independently of cytokinesis and involve specialized MreB structures that appear to be intermediates in this process.

Structural properties of prokaryotic promoter regions correlate with functional features.

PubMed

Meysman, Pieter; Collado-Vides, Julio; Morett, Enrique; Viola, Roberto; Engelen, Kristof; Laukens, Kris

2014-01-01

The structural properties of the DNA molecule are known to play a critical role in transcription. In this paper, the structural profiles of promoter regions were studied within the context of their diversity and their function for eleven prokaryotic species; Escherichia coli, Klebsiella pneumoniae, Salmonella Typhimurium, Pseudomonas auroginosa, Geobacter sulfurreducens Helicobacter pylori, Chlamydophila pneumoniae, Synechocystis sp., Synechoccocus elongates, Bacillus anthracis, and the archaea Sulfolobus solfataricus. The main anchor point for these promoter regions were transcription start sites identified through high-throughput experiments or collected within large curated databases. Prokaryotic promoter regions were found to be less stable and less flexible than the genomic mean across all studied species. However, direct comparison between species revealed differences in their structural profiles that can not solely be explained by the difference in genomic GC content. In addition, comparison with functional data revealed that there are patterns in the promoter structural profiles that can be linked to specific functional loci, such as sigma factor regulation or transcription factor binding. Interestingly, a novel structural element clearly visible near the transcription start site was found in genes associated with essential cellular functions and growth in several species. Our analyses reveals the great diversity in promoter structural profiles both between and within prokaryotic species. We observed relationships between structural diversity and functional features that are interesting prospects for further research to yet uncharacterized functional loci defined by DNA structural properties.

PTEN in the maintenance of genome integrity: From DNA replication to chromosome segregation.

PubMed

Hou, Sheng-Qi; Ouyang, Meng; Brandmaier, Andrew; Hao, Hongbo; Shen, Wen H

2017-10-01

Faithful DNA replication and accurate chromosome segregation are the key machineries of genetic transmission. Disruption of these processes represents a hallmark of cancer and often results from loss of tumor suppressors. PTEN is an important tumor suppressor that is frequently mutated or deleted in human cancer. Loss of PTEN has been associated with aneuploidy and poor prognosis in cancer patients. In mice, Pten deletion or mutation drives genomic instability and tumor development. PTEN deficiency induces DNA replication stress, confers stress tolerance, and disrupts mitotic spindle architecture, leading to accumulation of structural and numerical chromosome instability. Therefore, PTEN guards the genome by controlling multiple processes of chromosome inheritance. Here, we summarize current understanding of the PTEN function in promoting high-fidelity transmission of genetic information. We also discuss the PTEN pathways of genome maintenance and highlight potential targets for cancer treatment. © 2017 WILEY Periodicals, Inc.

Halophilic & halotolerant prokaryotes in humans.

PubMed

Seck, El Hadji; Dufour, Jean-Charles; Raoult, Didier; Lagier, Jean-Christophe

2018-05-04

Halophilic prokaryotes are described as microorganisms living in hypersaline environments. Here, we list the halotolerant and halophilic bacteria which have been isolated in humans. Of the 52 halophilic prokaryotes, 32 (61.54%) were moderately halophilic, 17 (32.69%) were slightly halophilic and three (5.76%) were extremely halophilic prokaryotes. At the phylum level, 29 (54.72%) belong to Firmicutes, 15 (28.84%) to Proteobacteria, four (7.69%) to Actinobacteria, three (5.78%) to Euryarchaeota and one (1.92%) belongs to Bacteroidetes. Halophilic prokaryotes are rarely pathogenic: of these 52 halophilic prokaryotes only two (3.92%) species were classified in Risk Group 2 (Vibrio cholerae, Vibrio parahaemolyticus) and one (1.96%), species in Risk Group 3 (Bacillus anthracis).

The structural role of the zinc ion can be dispensable in prokaryotic zinc-finger domains

PubMed Central

Baglivo, Ilaria; Russo, Luigi; Esposito, Sabrina; Malgieri, Gaetano; Renda, Mario; Salluzzo, Antonio; Di Blasio, Benedetto; Isernia, Carla; Fattorusso, Roberto; Pedone, Paolo V.

2009-01-01

The recent characterization of the prokaryotic Cys2His2 zinc-finger domain, identified in Ros protein from Agrobacterium tumefaciens, has demonstrated that, although possessing a similar zinc coordination sphere, this domain is structurally very different from its eukaryotic counterpart. A search in the databases has identified ≈300 homologues with a high sequence identity to the Ros protein, including the amino acids that form the extensive hydrophobic core in Ros. Surprisingly, the Cys2His2 zinc coordination sphere is generally poorly conserved in the Ros homologues, raising the question of whether the zinc ion is always preserved in these proteins. Here, we present a functional and structural study of a point mutant of Ros protein, Ros56–142C82D, in which the second coordinating cysteine is replaced by an aspartate, 5 previously-uncharacterized representative Ros homologues from Mesorhizobium loti, and 2 mutants of the homologues. Our results indicate that the prokaryotic zinc-finger domain, which in Ros protein tetrahedrally coordinates Zn(II) through the typical Cys2His2 coordination, in Ros homologues can either exploit a CysAspHis2 coordination sphere, previously never described in DNA binding zinc finger domains to our knowledge, or lose the metal, while still preserving the DNA-binding activity. We demonstrate that this class of prokaryotic zinc-finger domains is structurally very adaptable, and surprisingly single mutations can transform a zinc-binding domain into a nonzinc-binding domain and vice versa, without affecting the DNA-binding ability. In light of our findings an evolutionary link between the prokaryotic and eukaryotic zinc-finger domains, based on bacteria-to-eukaryota horizontal gene transfer, is discussed. PMID:19369210

Functional advantages conferred by extracellular prokaryotic membrane vesicles.

PubMed

Manning, Andrew J; Kuehn, Meta J

2013-01-01

The absence of subcellular organelles is a characteristic typically used to distinguish prokaryotic from eukaryotic cells. But recent discoveries do not support this dogma. Over the past 50 years, researchers have begun to appreciate and characterize Gram-negative bacterial outer membrane-derived vesicles and Gram-positive and archaeal membrane vesicles. These extracellular, membrane-bound organelles can perform a variety of functions, including binding and delivery of DNA, transport of virulence factors, protection of the cell from outer membrane targeting antimicrobials and ridding the cell of toxic envelope proteins. Here, we review the contributions of these extracellular organelles to prokaryotic physiology and compare these with the contributions of the bacterial interior membrane-bound organelles responsible for harvesting light energy and for generating magnetic crystals of heavy metals. Understanding the roles of these multifunctional extracellular vesicle organelles as microbial tools will help us to better realize the diverse interactions that occur in our polymicrobial world. Copyright © 2013 S. Karger AG, Basel.

The RECG1 DNA Translocase Is a Key Factor in Recombination Surveillance, Repair, and Segregation of the Mitochondrial DNA in Arabidopsis[OPEN

PubMed Central

Le Ret, Monique; Bergdoll, Marc; Bichara, Marc; Dietrich, André

2015-01-01

The mitochondria of flowering plants have considerably larger and more complex genomes than the mitochondria of animals or fungi, mostly due to recombination activities that modulate their genomic structures. These activities most probably participate in the repair of mitochondrial DNA (mtDNA) lesions by recombination-dependent processes. Rare ectopic recombination across short repeats generates new genomic configurations that contribute to mtDNA heteroplasmy, which drives rapid evolution of the sequence organization of plant mtDNAs. We found that Arabidopsis thaliana RECG1, an ortholog of the bacterial RecG translocase, is an organellar protein with multiple roles in mtDNA maintenance. RECG1 targets to mitochondria and plastids and can complement a bacterial recG mutant that shows defects in repair and replication control. Characterization of Arabidopsis recG1 mutants showed that RECG1 is required for recombination-dependent repair and for suppression of ectopic recombination in mitochondria, most likely because of its role in recovery of stalled replication forks. The analysis of alternative mitotypes present in a recG1 line and of their segregation following backcross allowed us to build a model to explain how a new stable mtDNA configuration, compatible with normal plant development, can be generated by stoichiometric shift. PMID:26462909

Helicobacter pylori shows asymmetric and polar cell divisome assembly associated with DNA replisome.

PubMed

Kamran, Mohammad; Dubey, Priyanka; Verma, Vijay; Dasgupta, Santanu; Dhar, Suman K

2018-05-09

DNA replication and cell division are two fundamental processes in the life cycle of a cell. The majority of prokaryotic cells undergo division by means of binary fission in coordination with replication of the genome. Both processes, but especially their coordination, are poorly understood in Helicobacter pylori. Here, we studied the cell divisome assembly and the subsequent processes of membrane and peptidoglycan synthesis in the bacterium. To our surprise, we found the cell divisome assembly to be polar, which was well-corroborated by the asymmetric membrane and peptidoglycan synthesis at the poles. The divisome components showed its assembly to be synchronous with that of the replisome and the two remained associated throughout the cell cycle, demonstrating a tight coordination among chromosome replication, segregation and cell division in H. pylori. To our knowledge, this is the first report where both DNA replication and cell division along with their possible association have been demonstrated for this pathogenic bacterium. © 2018 Federation of European Biochemical Societies.

Bioinformatics of prokaryotic RNAs

PubMed Central

Backofen, Rolf; Amman, Fabian; Costa, Fabrizio; Findeiß, Sven; Richter, Andreas S; Stadler, Peter F

2014-01-01

The genome of most prokaryotes gives rise to surprisingly complex transcriptomes, comprising not only protein-coding mRNAs, often organized as operons, but also harbors dozens or even hundreds of highly structured small regulatory RNAs and unexpectedly large levels of anti-sense transcripts. Comprehensive surveys of prokaryotic transcriptomes and the need to characterize also their non-coding components is heavily dependent on computational methods and workflows, many of which have been developed or at least adapted specifically for the use with bacterial and archaeal data. This review provides an overview on the state-of-the-art of RNA bioinformatics focusing on applications to prokaryotes. PMID:24755880

Minimization of extracellular space as a driving force in prokaryote association and the origin of eukaryotes.

PubMed

Hooper, Scott L; Burstein, Helaine J

2014-11-18

Internalization-based hypotheses of eukaryotic origin require close physical association of host and symbiont. Prior hypotheses of how these associations arose include chance, specific metabolic couplings between partners, and prey-predator/parasite interactions. Since these hypotheses were proposed, it has become apparent that mixed-species, close-association assemblages (biofilms) are widespread and predominant components of prokaryotic ecology. Which forces drove prokaryotes to evolve the ability to form these assemblages are uncertain. Bacteria and archaea have also been found to form membrane-lined interconnections (nanotubes) through which proteins and RNA pass. These observations, combined with the structure of the nuclear envelope and an energetic benefit of close association (see below), lead us to propose a novel hypothesis of the driving force underlying prokaryotic close association and the origin of eukaryotes. Respiratory proton transport does not alter external pH when external volume is effectively infinite. Close physical association decreases external volume. For small external volumes, proton transport decreases external pH, resulting in each transported proton increasing proton motor force to a greater extent. We calculate here that in biofilms this effect could substantially decrease how many protons need to be transported to achieve a given proton motor force. Based as it is solely on geometry, this energetic benefit would occur for all prokaryotes using proton-based respiration. This benefit may be a driving force in biofilm formation. Under this hypothesis a very wide range of prokaryotic species combinations could serve as eukaryotic progenitors. We use this observation and the discovery of prokaryotic nanotubes to propose that eukaryotes arose from physically distinct, functionally specialized (energy factory, protein factory, DNA repository/RNA factory), obligatorily symbiotic prokaryotes in which the protein factory and DNA repository

A Partial Least Squares Based Procedure for Upstream Sequence Classification in Prokaryotes.

PubMed

Mehmood, Tahir; Bohlin, Jon; Snipen, Lars

2015-01-01

The upstream region of coding genes is important for several reasons, for instance locating transcription factor, binding sites, and start site initiation in genomic DNA. Motivated by a recently conducted study, where multivariate approach was successfully applied to coding sequence modeling, we have introduced a partial least squares (PLS) based procedure for the classification of true upstream prokaryotic sequence from background upstream sequence. The upstream sequences of conserved coding genes over genomes were considered in analysis, where conserved coding genes were found by using pan-genomics concept for each considered prokaryotic species. PLS uses position specific scoring matrix (PSSM) to study the characteristics of upstream region. Results obtained by PLS based method were compared with Gini importance of random forest (RF) and support vector machine (SVM), which is much used method for sequence classification. The upstream sequence classification performance was evaluated by using cross validation, and suggested approach identifies prokaryotic upstream region significantly better to RF (p-value < 0.01) and SVM (p-value < 0.01). Further, the proposed method also produced results that concurred with known biological characteristics of the upstream region.

Stygofauna enhance prokaryotic transport in groundwater ecosystems.

PubMed

Smith, Renee J; Paterson, James S; Launer, Elise; Tobe, Shanan S; Morello, Eliesa; Leijs, Remko; Marri, Shashikanth; Mitchell, James G

2016-09-06

More than 97% of the world's freshwater reserves are found in aquifers, making groundwater one of the most important resources on the planet. Prokaryotic communities in groundwater underpin the turnover of energy and matter while also maintaining groundwater purity. Thus, knowledge of microbial transport in the subsurface is crucial for maintaining groundwater health. Here, we describe for the first time the importance of stygofauna as vectors for prokaryotes. The "hitch-hiking" prokaryotes associated with stygofauna may be up to 5 orders of magnitude higher in abundance and transported up to 34× faster than bulk groundwater flow. We also demonstrate that prokaryotic diversity associated with stygofauna may be higher than that of the surrounding groundwater. Stygofauna are a newly recognized prokaryotic niche in groundwater ecosystems that have the potential to transport remediating, water purifying and pathogenic prokaryotes. Therefore, stygofauna may influence ecosystem dynamics and health at a microbial level, and at a larger scale could be a new source of prokaryotic diversity in groundwater ecosystems.

The prokaryotic zinc-finger: structure, function and comparison with the eukaryotic counterpart.

PubMed

Malgieri, Gaetano; Palmieri, Maddalena; Russo, Luigi; Fattorusso, Roberto; Pedone, Paolo V; Isernia, Carla

2015-12-01

Classical zinc finger (ZF) domains were thought to be confined to the eukaryotic kingdom until the transcriptional regulator Ros protein was identified in Agrobacterium tumefaciens. The Ros Cys2 His2 ZF binds DNA in a peculiar mode and folds in a domain significantly larger than its eukaryotic counterpart consisting of 58 amino acids (the 9-66 region) arranged in a βββαα topology, and stabilized by a conserved, extensive, 15-residue hydrophobic core. The prokaryotic ZF domain, then, shows some intriguing new features that make it interestingly different from its eukaryotic counterpart. This review will focus on the prokaryotic ZFs, summarizing and discussing differences and analogies with the eukaryotic domains and providing important insights into their structure/function relationships. © 2015 FEBS.

[Prokaryotic expression, purification and antigenicity identification of recombinant human survivin protein].

PubMed

Yin, Xiaotao; Wang, Wei; Tian, Renli; Xu, Yuanji; Yan, Jinqi; Zhang, Wei; Gao, Jiangping; Yu, Jiyun

2013-08-01

To construct a prokaryotic expression plasmid pET28a-survivin, optimize the recombinant protein expression conditions in E.coli, and purify the survivin recombinant protein and identify its antigenicity. Survivin cDNA segment was amplified by PCR and cloned into prokaryotic expression vector pET28a(+) to construct the recombinant expression vector pET28a-survivin. The expression vector was transformed into BL21 (DE3) and the fusion protein survivin/His was induced by IPTG. The fusion protein was purified through Ni affinity chromatography. The antigenicity of the purified survivin protein was identified by Western blotting and ELISA. The recombinant expression vector was verified successfully by BamHI and HindIII. The fusion protein induced by IPTG was obtained with Mr; about 24 000. The purity of the purified protein reached 90% by SDS-PAGE analysis. And the antigenicity of the survivin protein was validated by Western blotting and ELISA. The prokaryotic expression plasmid pET28a-survivin was successfully constructed and the survivin protein was expressed and purified in E.coli. The antigenicity of the purified survivin protein was demonstrated desirable.

Stygofauna enhance prokaryotic transport in groundwater ecosystems

PubMed Central

Smith, Renee J.; Paterson, James S.; Launer, Elise; Tobe, Shanan S.; Morello, Eliesa; Leijs, Remko; Marri, Shashikanth; Mitchell, James G.

2016-01-01

More than 97% of the world’s freshwater reserves are found in aquifers, making groundwater one of the most important resources on the planet. Prokaryotic communities in groundwater underpin the turnover of energy and matter while also maintaining groundwater purity. Thus, knowledge of microbial transport in the subsurface is crucial for maintaining groundwater health. Here, we describe for the first time the importance of stygofauna as vectors for prokaryotes. The “hitch-hiking” prokaryotes associated with stygofauna may be up to 5 orders of magnitude higher in abundance and transported up to 34× faster than bulk groundwater flow. We also demonstrate that prokaryotic diversity associated with stygofauna may be higher than that of the surrounding groundwater. Stygofauna are a newly recognized prokaryotic niche in groundwater ecosystems that have the potential to transport remediating, water purifying and pathogenic prokaryotes. Therefore, stygofauna may influence ecosystem dynamics and health at a microbial level, and at a larger scale could be a new source of prokaryotic diversity in groundwater ecosystems. PMID:27597322

[Norrie syndrome: identification of carriers by segregation analysis with flanking DNA markers].

PubMed

Körner, J; Uhlhaas, S; Neugebauer, M; Gal, A

1989-01-01

Norrie disease is an X-linked recessive disorder. Affected males present with congenital blindness. Additionally, hearing loss and psychotic behavior may occur at any time. Since carriers are clinically healthy, they can only be identified by genetic means. Daughters of carriers or sisters of affected males have an à priori 50% risk of being carriers themselves. Close linkage has been found between the Norrie disease locus (NDP) and the DNA locus DXS7 mapped to Xp11.3. For genetic counselling, this linkage relationship allows carriers of the disease to be identified in informative families. We describe a large pedigree with Norrie disease. Segregation analysis was carried out with DXS7 and a second flanking marker, DXS255, both linked to NDP. In this way, three females at risk were identified who had a high probability of being carriers for Norrie disease.

The tail of the ParG DNA segregation protein remodels ParF polymers and enhances ATP hydrolysis via an arginine finger-like motif

PubMed Central

Barillà, Daniela; Carmelo, Emma; Hayes, Finbarr

2007-01-01

The ParF protein of plasmid TP228 belongs to the ubiquitous superfamily of ParA ATPases that drive DNA segregation in bacteria. ATP-bound ParF polymerizes into multistranded filaments. The partner protein ParG is dimeric, consisting of C-termini that interweave into a ribbon–helix–helix domain contacting the centromeric DNA and unstructured N-termini. ParG stimulates ATP hydrolysis by ParF ≈30-fold. Here, we establish that the mobile tails of ParG are crucial for this enhancement and that arginine R19 within the tail is absolutely required for activation of ParF nucleotide hydrolysis. R19 is part of an arginine finger-like loop in ParG that is predicted to intercalate into the ParF nucleotide-binding pocket thereby promoting ATP hydrolysis. Significantly, mutations of R19 abrogated DNA segregation in vivo, proving that intracellular stimulation of ATP hydrolysis by ParG is a key regulatory process for partitioning. Furthermore, ParG bundles ParF-ATP filaments as well as promoting nucleotide-independent polymerization. The N-terminal flexible tail is required for both activities, because N-terminal ΔParG polypeptides are defective in both functions. Strikingly, the critical arginine finger-like residue R19 is dispensable for ParG-mediated remodeling of ParF polymers, revealing that the ParG N-terminal tail possesses two separable activities in the interplay with ParF: a catalytic function during ATP hydrolysis and a mechanical role in modulation of polymerization. We speculate that activation of nucleotide hydrolysis via an arginine finger loop may be a conserved, regulatory mechanism of ParA family members and their partner proteins, including ParA-ParB and Soj-Spo0J that mediate DNA segregation and MinD-MinE that determine septum localization. PMID:17261809

The tail of the ParG DNA segregation protein remodels ParF polymers and enhances ATP hydrolysis via an arginine finger-like motif.

PubMed

Barillà, Daniela; Carmelo, Emma; Hayes, Finbarr

2007-02-06

The ParF protein of plasmid TP228 belongs to the ubiquitous superfamily of ParA ATPases that drive DNA segregation in bacteria. ATP-bound ParF polymerizes into multistranded filaments. The partner protein ParG is dimeric, consisting of C-termini that interweave into a ribbon-helix-helix domain contacting the centromeric DNA and unstructured N-termini. ParG stimulates ATP hydrolysis by ParF approximately 30-fold. Here, we establish that the mobile tails of ParG are crucial for this enhancement and that arginine R19 within the tail is absolutely required for activation of ParF nucleotide hydrolysis. R19 is part of an arginine finger-like loop in ParG that is predicted to intercalate into the ParF nucleotide-binding pocket thereby promoting ATP hydrolysis. Significantly, mutations of R19 abrogated DNA segregation in vivo, proving that intracellular stimulation of ATP hydrolysis by ParG is a key regulatory process for partitioning. Furthermore, ParG bundles ParF-ATP filaments as well as promoting nucleotide-independent polymerization. The N-terminal flexible tail is required for both activities, because N-terminal DeltaParG polypeptides are defective in both functions. Strikingly, the critical arginine finger-like residue R19 is dispensable for ParG-mediated remodeling of ParF polymers, revealing that the ParG N-terminal tail possesses two separable activities in the interplay with ParF: a catalytic function during ATP hydrolysis and a mechanical role in modulation of polymerization. We speculate that activation of nucleotide hydrolysis via an arginine finger loop may be a conserved, regulatory mechanism of ParA family members and their partner proteins, including ParA-ParB and Soj-Spo0J that mediate DNA segregation and MinD-MinE that determine septum localization.

Molecular model of the mitochondrial genome segregation machinery in Trypanosoma brucei

PubMed Central

Hoffmann, Anneliese; Käser, Sandro; Jakob, Martin; Amodeo, Simona; Peitsch, Camille; Týč, Jiří; Vaughan, Sue; Schneider, André

2018-01-01

In almost all eukaryotes, mitochondria maintain their own genome. Despite the discovery more than 50 y ago, still very little is known about how the genome is correctly segregated during cell division. The protozoan parasite Trypanosoma brucei contains a single mitochondrion with a singular genome, the kinetoplast DNA (kDNA). Electron microscopy studies revealed the tripartite attachment complex (TAC) to physically connect the kDNA to the basal body of the flagellum and to ensure correct segregation of the mitochondrial genome via the basal bodies movement, during the cell cycle. Using superresolution microscopy, we precisely localize each of the currently known TAC components. We demonstrate that the TAC is assembled in a hierarchical order from the base of the flagellum toward the mitochondrial genome and that the assembly is not dependent on the kDNA itself. Based on the biochemical analysis, the TAC consists of several nonoverlapping subcomplexes, suggesting an overall size of the TAC exceeding 2.8 mDa. We furthermore demonstrate that the TAC is required for correct mitochondrial organelle positioning but not for organelle biogenesis or segregation. PMID:29434039

Prokaryote genome fluidity: toward a system approach of the mobilome.

PubMed

Toussaint, Ariane; Chandler, Mick

2012-01-01

The importance of horizontal/lateral gene transfer (LGT) in shaping the genomes of prokaryotic organisms has been recognized in recent years as a result of analysis of the increasing number of available genome sequences. LGT is largely due to the transfer and recombination activities of mobile genetic elements (MGEs). Bacterial and archaeal genomes are mosaics of vertically and horizontally transmitted DNA segments. This generates reticulate relationships between members of the prokaryotic world that are better represented by networks than by "classical" phylogenetic trees. In this review we summarize the nature and activities of MGEs, and the problems that presently limit their analysis on a large scale. We propose routes to improve their annotation in the flow of genomic and metagenomic sequences that currently exist and those that become available. We describe network analysis of evolutionary relationships among some MGE categories and sketch out possible developments of this type of approach to get more insight into the role of the mobilome in bacterial adaptation and evolution.

The evolution of ecological tolerance in prokaryotes

NASA Technical Reports Server (NTRS)

Knoll, A. H.; Bauld, J.

1989-01-01

The ecological ranges of Archaeobacteria and Eubacteria are constrained by a requirement for liquid water and the physico-chemical stability limits of biomolecules, but within this broad envelope, prokaryotes have evolved adaptations that permit them to tolerate a remarkable spectrum of habitats. Laboratory experiments indicate that prokaryotes can adapt rapidly to novel environmental conditions, yet geological studies suggest early diversification and long-term stasis within the prokaryotic kingdoms. These apparently contradictory perspectives can be reconciled by understanding that, in general, rates and patterns of prokaryotic evolution reflect the developmental history of the Earth's surface environments. Our understanding of modern microbial ecology provides a lens through which our accumulating knowledge of physiology, molecular phylogeny and the Earth's history can be integrated and focussed on the phenomenon of prokaryotic evolution.

Stygofauna enhance prokaryotic transport in groundwater ecosystems

DOE Office of Scientific and Technical Information (OSTI.GOV)

Smith, Renee J.; Paterson, James S.; Launer, Elise

More than 97% of the world’s freshwater reserves are found in aquifers, making groundwater one of the most important resources on the planet. Prokaryotic communities in groundwater underpin the turnover of energy and matter while also maintaining groundwater purity. Thus, knowledge of microbial transport in the subsurface is crucial for maintaining groundwater health. Here, we describe for the first time the importance of stygofauna as vectors for prokaryotes. The “hitch-hiking” prokaryotes associated with stygofauna may be up to 5 orders of magnitude higher in abundance and transported up to 34× faster than bulk groundwater flow. We also demonstrate that prokaryoticmore » diversity associated with stygofauna may be higher than that of the surrounding groundwater. Stygofauna are a newly recognized prokaryotic niche in groundwater ecosystems that have the potential to transport remediating, water purifying and pathogenic prokaryotes. Furthermore, stygofauna may influence ecosystem dynamics and health at a microbial level, and at a larger scale could be a new source of prokaryotic diversity in groundwater ecosystems.« less

Stygofauna enhance prokaryotic transport in groundwater ecosystems

DOE PAGES

Smith, Renee J.; Paterson, James S.; Launer, Elise; ...

2016-09-06

More than 97% of the world’s freshwater reserves are found in aquifers, making groundwater one of the most important resources on the planet. Prokaryotic communities in groundwater underpin the turnover of energy and matter while also maintaining groundwater purity. Thus, knowledge of microbial transport in the subsurface is crucial for maintaining groundwater health. Here, we describe for the first time the importance of stygofauna as vectors for prokaryotes. The “hitch-hiking” prokaryotes associated with stygofauna may be up to 5 orders of magnitude higher in abundance and transported up to 34× faster than bulk groundwater flow. We also demonstrate that prokaryoticmore » diversity associated with stygofauna may be higher than that of the surrounding groundwater. Stygofauna are a newly recognized prokaryotic niche in groundwater ecosystems that have the potential to transport remediating, water purifying and pathogenic prokaryotes. Furthermore, stygofauna may influence ecosystem dynamics and health at a microbial level, and at a larger scale could be a new source of prokaryotic diversity in groundwater ecosystems.« less

Modulation of DNA binding by gene-specific transcription factors.

PubMed

Schleif, Robert F

2013-10-01

The transcription of many genes, particularly in prokaryotes, is controlled by transcription factors whose activity can be modulated by controlling their DNA binding affinity. Understanding the molecular mechanisms by which DNA binding affinity is regulated is important, but because forming definitive conclusions usually requires detailed structural information in combination with data from extensive biophysical, biochemical, and sometimes genetic experiments, little is truly understood about this topic. This review describes the biological requirements placed upon DNA binding transcription factors and their consequent properties, particularly the ways that DNA binding affinity can be modulated and methods for its study. What is known and not known about the mechanisms modulating the DNA binding affinity of a number of prokaryotic transcription factors, including CAP and lac repressor, is provided.

Prokaryotic carbonic anhydrases of Earth's environment.

PubMed

Kumar, R Siva Sai; Ferry, James G

2014-01-01

Carbonic anhydrase is a metalloenzyme catalyzing the reversible hydration of carbon dioxide to bicarbonate. Five independently evolved classes have been described for which one or more are found in nearly every cell type underscoring the general importance of this ubiquitous enzyme in Nature. The bulk of research to date has centered on the enzymes from mammals and plants with less emphasis on prokaryotes. Prokaryotic carbonic anhydrases play important roles in the ecology of Earth's biosphere including acquisition of CO2 for photosynthesis and the physiology of aerobic and anaerobic prokaryotes decomposing the photosynthate back to CO2 thereby closing the global carbon cycle. This review focuses on the physiology and biochemistry of carbonic anhydrases from prokaryotes belonging to the domains Bacteria and Archaea that play key roles in the ecology of Earth's biosphere.

Cospeciation of Psyllids and Their Primary Prokaryotic Endosymbionts

PubMed Central

Thao, MyLo L.; Moran, Nancy A.; Abbot, Patrick; Brennan, Eric B.; Burckhardt, Daniel H.; Baumann, Paul

2000-01-01

Psyllids are plant sap-feeding insects that harbor prokaryotic endosymbionts in specialized cells within the body cavity. Four-kilobase DNA fragments containing 16S and 23S ribosomal DNA (rDNA) were amplified from the primary (P) endosymbiont of 32 species of psyllids representing three psyllid families and eight subfamilies. In addition, 0.54-kb fragments of the psyllid nuclear gene wingless were also amplified from 26 species. Phylogenetic trees derived from 16S-23S rDNA and from the host wingless gene are very similar, and tests of compatibility of the data sets show no significant conflict between host and endosymbiont phylogenies. This result is consistent with a single infection of a shared psyllid ancestor and subsequent cospeciation of the host and the endosymbiont. In addition, the phylogenies based on DNA sequences generally agreed with psyllid taxonomy based on morphology. The 3′ end of the 16S rDNA of the P endosymbionts differs from that of other members of the domain Bacteria in the lack of a sequence complementary to the mRNA ribosome binding site. The rate of sequence change in the 16S-23S rDNA of the psyllid P endosymbiont was considerably higher than that of other bacteria, including other fast-evolving insect endosymbionts. The lineage consisting of the P endosymbionts of psyllids was given the designation Candidatus Carsonella (gen. nov.) with a single species, Candidatus Carsonella ruddii (sp. nov.). PMID:10877784

Direct detection of methylation in genomic DNA

PubMed Central

Bart, A.; van Passel, M. W. J.; van Amsterdam, K.; van der Ende, A.

2005-01-01

The identification of methylated sites on bacterial genomic DNA would be a useful tool to study the major roles of DNA methylation in prokaryotes: distinction of self and nonself DNA, direction of post-replicative mismatch repair, control of DNA replication and cell cycle, and regulation of gene expression. Three types of methylated nucleobases are known: N6-methyladenine, 5-methylcytosine and N4-methylcytosine. The aim of this study was to develop a method to detect all three types of DNA methylation in complete genomic DNA. It was previously shown that N6-methyladenine and 5-methylcytosine in plasmid and viral DNA can be detected by intersequence trace comparison of methylated and unmethylated DNA. We extended this method to include N4-methylcytosine detection in both in vitro and in vivo methylated DNA. Furthermore, application of intersequence trace comparison was extended to bacterial genomic DNA. Finally, we present evidence that intrasequence comparison suffices to detect methylated sites in genomic DNA. In conclusion, we present a method to detect all three natural types of DNA methylation in bacterial genomic DNA. This provides the possibility to define the complete methylome of any prokaryote. PMID:16091626

A framework for classification of prokaryotic protein kinases.

PubMed

Tyagi, Nidhi; Anamika, Krishanpal; Srinivasan, Narayanaswamy

2010-05-26

Overwhelming majority of the Serine/Threonine protein kinases identified by gleaning archaeal and eubacterial genomes could not be classified into any of the well known Hanks and Hunter subfamilies of protein kinases. This is owing to the development of Hanks and Hunter classification scheme based on eukaryotic protein kinases which are highly divergent from their prokaryotic homologues. A large dataset of prokaryotic Serine/Threonine protein kinases recognized from genomes of prokaryotes have been used to develop a classification framework for prokaryotic Ser/Thr protein kinases. We have used traditional sequence alignment and phylogenetic approaches and clustered the prokaryotic kinases which represent 72 subfamilies with at least 4 members in each. Such a clustering enables classification of prokaryotic Ser/Thr kinases and it can be used as a framework to classify newly identified prokaryotic Ser/Thr kinases. After series of searches in a comprehensive sequence database we recognized that 38 subfamilies of prokaryotic protein kinases are associated to a specific taxonomic level. For example 4, 6 and 3 subfamilies have been identified that are currently specific to phylum proteobacteria, cyanobacteria and actinobacteria respectively. Similarly subfamilies which are specific to an order, sub-order, class, family and genus have also been identified. In addition to these, we also identify organism-diverse subfamilies. Members of these clusters are from organisms of different taxonomic levels, such as archaea, bacteria, eukaryotes and viruses. Interestingly, occurrence of several taxonomic level specific subfamilies of prokaryotic kinases contrasts with classification of eukaryotic protein kinases in which most of the popular subfamilies of eukaryotic protein kinases occur diversely in several eukaryotes. Many prokaryotic Ser/Thr kinases exhibit a wide variety of modular organization which indicates a degree of complexity and protein-protein interactions in the

Restriction-Modification systems interplay causes avoidance of GATC site in prokaryotic genomes.

PubMed

Ershova, Anna; Rusinov, Ivan; Vasiliev, Mikhail; Spirin, Sergey; Karyagina, Anna

2016-04-01

Palindromes are frequently underrepresented in prokaryotic genomes. Palindromic 5[Formula: see text]-GATC-3[Formula: see text] site is a recognition site of different Restriction-Modification (R-M) systems, as well as solitary methyltransferase Dam. Classical GATC-specific R-M systems methylate GATC and cleave unmethylated GATC. On the contrary, methyl-directed Type II restriction endonucleases cleave methylated GATC. Methylation of GATC by Dam methyltransferase is involved in the regulation of different cellular processes. The diversity of functions of GATC-recognizing proteins makes GATC sequence a good model for studying the reasons of palindrome avoidance in prokaryotic genomes. In this work, the influence of R-M systems and solitary proteins on the GATC site avoidance is described by a mathematical model. GATC avoidance is strongly associated with the presence of alternate (methyl-directed or classical Type II R-M system) genes in different strains of the same species, as we have shown for Streptococcus pneumoniae, Neisseria meningitidis, Eubacterium rectale, and Moraxella catarrhalis. We hypothesize that GATC avoidance can result from a DNA exchange between strains with different methylation status of GATC site within the process of natural transformation. If this hypothesis is correct, the GATC avoidance is a sign of a DNA exchange between bacteria with different methylation status in a mixed population.

A comparison of stable-isotope probing of DNA and phospholipid fatty acids to study prokaryotic functional diversity in sulfate-reducing marine sediment enrichment slurries.

PubMed

Webster, Gordon; Watt, Lynsey C; Rinna, Joachim; Fry, John C; Evershed, Richard P; Parkes, R John; Weightman, Andrew J

2006-09-01

Marine sediment slurries enriched for anaerobic, sulfate-reducing prokaryotic communities utilizing glucose and acetate were used to provide the first comparison between stable-isotope probing (SIP) of phospholipid fatty acids (PLFA) and DNA (16S rRNA and dsrA genes) biomarkers. Different 13C-labelled substrates (glucose, acetate and pyruvate) at low concentrations (100 microM) were used over a 7-day incubation to follow and identify carbon flow into different members of the community. Limited changes in total PLFA and bacterial 16S rRNA gene DGGE profiles over 7 days suggested the presence of a stable bacterial community. A broad range of PLFA were rapidly labelled (within 12 h) in the 13C-glucose slurry but this changed with time, suggesting the presence of an active glucose-utilizing population and later development of another population able to utilize glucose metabolites. The identity of the major glucose-utilizers was unclear as 13C-enriched PLFA were common (16:0, 16:1, 18:1omega7, highest incorporation) and there was little difference between 12C- and 13C-DNA 16S rRNA gene denaturing gradient gel electrophoresis (DGGE) profiles. Seemingly glucose, a readily utilizable substrate, resulted in widespread incorporation consistent with the higher extent of 13C-incorporation (approximately 10 times) into PLFA compared with 13C-acetate or 13C-pyruvate. 13C-PLFA in the 13C-acetate and 13C-pyruvate slurries were similar to each other and to those that developed in the 13C-glucose slurry after 4 days. These were more diagnostic, with branched odd-chain fatty acids (i15:0, a15:0 and 15:1omega6) possibly indicating the presence of Desulfococcus or Desulfosarcina sulfate-reducing bacteria (SRB) and sequences related to these SRB were in the 13C-acetate-DNA dsrA gene library. The 13C-acetate-DNA 16S rRNA gene library also contained sequences closely related to SRB, but these were the acetate-utilizing Desulfobacter sp., as well as a broad range of uncultured Bacteria. In

Desiccation tolerance of prokaryotes.

PubMed Central

Potts, M

1994-01-01

The removal of cell-bound water through air drying and the addition of water to air-dried cells are forces that have played a pivotal role in the evolution of the prokaryotes. In bacterial cells that have been subjected to air drying, the evaporation of free cytoplasmic water (Vf) can be instantaneous, and an equilibrium between cell-bound water (Vb) and the environmental water (vapor) potential (psi wv) may be achieved rapidly. In the air-dried state some bacteria survive only for seconds whereas others can tolerate desiccation for thousands, perhaps millions, of years. The desiccated (anhydrobiotic) cell is characterized by its singular lack of water--with contents as low as 0.02 g of H2O g (dry weight)-1. At these levels the monolayer coverage by water of macromolecules, including DNA and proteins, is disturbed. As a consequence the mechanisms that confer desiccation tolerance upon air-dried bacteria are markedly different from those, such as the mechanism of preferential exclusion of compatible solutes, that preserve the integrity of salt-, osmotically, and freeze-thaw-stressed cells. Desiccation tolerance reflects a complex array of interactions at the structural, physiological, and molecular levels. Many of the mechanisms remain cryptic, but it is clear that they involve interactions, such as those between proteins and co-solvents, that derive from the unique properties of the water molecule. A water replacement hypothesis accounts for how the nonreducing disaccharides trehalose and sucrose preserve the integrity of membranes and proteins. Nevertheless, we have virtually no insight into the state of the cytoplasm of an air-dried cell. There is no evidence for any obvious adaptations of proteins that can counter the effects of air drying or for the occurrence of any proteins that provide a direct and a tangible contribution to cell stability. Among the prokaryotes that can exist as anhydrobiotic cells, the cyanobacteria have a marked capacity to do so. One

Biophysical Adaptations of Prokaryotic Voltage-Gated Sodium Channels.

PubMed

Vien, T N; DeCaen, P G

2016-01-01

This chapter describes the adaptive features found in voltage-gated sodium channels (NaVs) of prokaryotes and eukaryotes. These two families are distinct, having diverged early in evolutionary history but maintain a surprising degree of convergence in function. While prokaryotic NaVs are required for growth and motility, eukaryotic NaVs selectively conduct fast electrical currents for short- and long-range signaling across cell membranes in mammalian organs. Current interest in prokaryotic NaVs is stoked by their resolved high-resolution structures and functional features which are reminiscent of eukaryotic NaVs. In this chapter, comparisons between eukaryotic and prokaryotic NaVs are made to highlight the shared and unique aspects of ion selectivity, voltage sensitivity, and pharmacology. Examples of prokaryotic and eukaryotic NaV convergent evolution will be discussed within the context of their structural features. Copyright © 2016 Elsevier Inc. All rights reserved.

Brownian Ratchet Mechanism for Faithful Segregation of Low-Copy-Number Plasmids.

PubMed

Hu, Longhua; Vecchiarelli, Anthony G; Mizuuchi, Kiyoshi; Neuman, Keir C; Liu, Jian

2017-04-11

Bacterial plasmids are extrachromosomal DNA that provides selective advantages for bacterial survival. Plasmid partitioning can be remarkably robust. For high-copy-number plasmids, diffusion ensures that both daughter cells inherit plasmids after cell division. In contrast, most low-copy-number plasmids need to be actively partitioned by a conserved tripartite ParA-type system. ParA is an ATPase that binds to chromosomal DNA; ParB is the stimulator of the ParA ATPase and specifically binds to the plasmid at a centromere-like site, parS. ParB stimulation of the ParA ATPase releases ParA from the bacterial chromosome, after which it takes a long time to reset its DNA-binding affinity. We previously demonstrated in vitro that the ParA system can exploit this biochemical asymmetry for directed cargo transport. Multiple ParA-ParB bonds can bridge a parS-coated cargo to a DNA carpet, and they can work collectively as a Brownian ratchet that directs persistent cargo movement with a ParA-depletion zone trailing behind. By extending this model, we suggest that a similar Brownian ratchet mechanism recapitulates the full range of actively segregated plasmid motilities observed in vivo. We demonstrate that plasmid motility is tuned as the replenishment rate of the ParA-depletion zone progressively increases relative to the cargo speed, evolving from diffusion to pole-to-pole oscillation, local excursions, and, finally, immobility. When the plasmid replicates, the daughters largely display motilities similar to that of their mother, except that when the single-focus progenitor is locally excursive, the daughter foci undergo directed segregation. We show that directed segregation maximizes the fidelity of plasmid partition. Given that local excursion and directed segregation are the most commonly observed modes of plasmid motility in vivo, we suggest that the operation of the ParA-type partition system has been shaped by evolution for high fidelity of plasmid segregation

LPSN—list of prokaryotic names with standing in nomenclature

PubMed Central

Parte, Aidan C.

2014-01-01

The List of Prokaryotic Names with Standing in Nomenclature (LPSN; http://www.bacterio.net) is a database that lists the names of prokaryotes (Bacteria and Archaea) that have been validly published in the International Journal of Systematic and Evolutionary Microbiology directly or by inclusion in a Validation List, under the Rules of International Code of Nomenclature of Bacteria. Currently there are 15 974 taxa listed. In addition, LPSN has an up-to-date classification of prokaryotes and information on prokaryotic nomenclature and culture collections. PMID:24243842

Insights into structural variations and genome rearrangements in prokaryotic genomes.

PubMed

Periwal, Vinita; Scaria, Vinod

2015-01-01

Structural variations (SVs) are genomic rearrangements that affect fairly large fragments of DNA. Most of the SVs such as inversions, deletions and translocations have been largely studied in context of genetic diseases in eukaryotes. However, recent studies demonstrate that genome rearrangements can also have profound impact on prokaryotic genomes, leading to altered cell phenotype. In contrast to single-nucleotide variations, SVs provide a much deeper insight into organization of bacterial genomes at a much better resolution. SVs can confer change in gene copy number, creation of new genes, altered gene expression and many other functional consequences. High-throughput technologies have now made it possible to explore SVs at a much refined resolution in bacterial genomes. Through this review, we aim to highlight the importance of the less explored field of SVs in prokaryotic genomes and their impact. We also discuss its potential applicability in the emerging fields of synthetic biology and genome engineering where targeted SVs could serve to create sophisticated and accurate genome editing. © The Author 2014. Published by Oxford University Press. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.

DNA-damage response during mitosis induces whole-chromosome missegregation.

PubMed

Bakhoum, Samuel F; Kabeche, Lilian; Murnane, John P; Zaki, Bassem I; Compton, Duane A

2014-11-01

Many cancers display both structural (s-CIN) and numerical (w-CIN) chromosomal instabilities. Defective chromosome segregation during mitosis has been shown to cause DNA damage that induces structural rearrangements of chromosomes (s-CIN). In contrast, whether DNA damage can disrupt mitotic processes to generate whole chromosomal instability (w-CIN) is unknown. Here, we show that activation of the DNA-damage response (DDR) during mitosis selectively stabilizes kinetochore-microtubule (k-MT) attachments to chromosomes through Aurora-A and PLK1 kinases, thereby increasing the frequency of lagging chromosomes during anaphase. Inhibition of DDR proteins, ATM or CHK2, abolishes the effect of DNA damage on k-MTs and chromosome segregation, whereas activation of the DDR in the absence of DNA damage is sufficient to induce chromosome segregation errors. Finally, inhibiting the DDR during mitosis in cancer cells with persistent DNA damage suppresses inherent chromosome segregation defects. Thus, the DDR during mitosis inappropriately stabilizes k-MTs, creating a link between s-CIN and w-CIN. The genome-protective role of the DDR depends on its ability to delay cell division until damaged DNA can be fully repaired. Here, we show that when DNA damage is induced during mitosis, the DDR unexpectedly induces errors in the segregation of entire chromosomes, thus linking structural and numerical chromosomal instabilities. ©2014 American Association for Cancer Research.

Primer design for a prokaryotic differential display RT-PCR.

PubMed Central

Fislage, R; Berceanu, M; Humboldt, Y; Wendt, M; Oberender, H

1997-01-01

We have developed a primer set for a prokaryotic differential display of mRNA in the Enterobacteriaceae group. Each combination of ten 10mer and ten 11mer primers generates up to 85 bands from total Escherichia coli RNA, thus covering expressed sequences of a complete bacterial genome. Due to the lack of polyadenylation in prokaryotic RNA the type T11VN anchored oligonucleotides for the reverse transcriptase reaction had to be replaced with respect to the original method described by Liang and Pardee [ Science , 257, 967-971 (1992)]. Therefore, the sequences of both the 10mer and the new 11mer oligonucleotides were determined by a statistical evaluation of species-specific coding regions extracted from the EMBL database. The 11mer primers used for reverse transcription were selected for localization in the 3'-region of the bacterial RNA. The 10mer primers preferentially bind to the 5'-end of the RNA. None of the primers show homology to rRNA or other abundant small RNA species. Randomly sampled cDNA bands were checked for their bacterial origin either by re-amplification, cloning and sequencing or by re-amplification and direct sequencing with 10mer and 11mer primers after asymmetric PCR. PMID:9108168

Primer design for a prokaryotic differential display RT-PCR.

PubMed

Fislage, R; Berceanu, M; Humboldt, Y; Wendt, M; Oberender, H

1997-05-01

We have developed a primer set for a prokaryotic differential display of mRNA in the Enterobacteriaceae group. Each combination of ten 10mer and ten 11mer primers generates up to 85 bands from total Escherichia coli RNA, thus covering expressed sequences of a complete bacterial genome. Due to the lack of polyadenylation in prokaryotic RNA the type T11VN anchored oligonucleotides for the reverse transcriptase reaction had to be replaced with respect to the original method described by Liang and Pardee [ Science , 257, 967-971 (1992)]. Therefore, the sequences of both the 10mer and the new 11mer oligonucleotides were determined by a statistical evaluation of species-specific coding regions extracted from the EMBL database. The 11mer primers used for reverse transcription were selected for localization in the 3'-region of the bacterial RNA. The 10mer primers preferentially bind to the 5'-end of the RNA. None of the primers show homology to rRNA or other abundant small RNA species. Randomly sampled cDNA bands were checked for their bacterial origin either by re-amplification, cloning and sequencing or by re-amplification and direct sequencing with 10mer and 11mer primers after asymmetric PCR.

Prokaryotic diversity, composition structure, and phylogenetic analysis of microbial communities in leachate sediment ecosystems.

PubMed

Liu, Jingjing; Wu, Weixiang; Chen, Chongjun; Sun, Faqian; Chen, Yingxu

2011-09-01

In order to obtain insight into the prokaryotic diversity and community in leachate sediment, a culture-independent DNA-based molecular phylogenetic approach was performed with archaeal and bacterial 16S rRNA gene clone libraries derived from leachate sediment of an aged landfill. A total of 59 archaeal and 283 bacterial rDNA phylotypes were identified in 425 archaeal and 375 bacterial analyzed clones. All archaeal clones distributed within two archaeal phyla of the Euryarchaeota and Crenarchaeota, and well-defined methanogen lineages, especially Methanosaeta spp., are the most numerically dominant species of the archaeal community. Phylogenetic analysis of the bacterial library revealed a variety of pollutant-degrading and biotransforming microorganisms, including 18 distinct phyla. A substantial fraction of bacterial clones showed low levels of similarity with any previously documented sequences and thus might be taxonomically new. Chemical characteristics and phylogenetic inferences indicated that (1) ammonium-utilizing bacteria might form consortia to alleviate or avoid the negative influence of high ammonium concentration on other microorganisms, and (2) members of the Crenarchaeota found in the sediment might be involved in ammonium oxidation. This study is the first to report the composition of the microbial assemblages and phylogenetic characteristics of prokaryotic populations extant in leachate sediment. Additional work on microbial activity and contaminant biodegradation remains to be explored.

Evolutionary dynamics of adult stem cells: comparison of random and immortal-strand segregation mechanisms.

PubMed

Tannenbaum, Emmanuel; Sherley, James L; Shakhnovich, Eugene I

2005-04-01

This paper develops a point-mutation model describing the evolutionary dynamics of a population of adult stem cells. Such a model may prove useful for quantitative studies of tissue aging and the emergence of cancer. We consider two modes of chromosome segregation: (1) random segregation, where the daughter chromosomes of a given parent chromosome segregate randomly into the stem cell and its differentiating sister cell and (2) "immortal DNA strand" co-segregation, for which the stem cell retains the daughter chromosomes with the oldest parent strands. Immortal strand co-segregation is a mechanism, originally proposed by [Cairns Nature (London) 255, 197 (1975)], by which stem cells preserve the integrity of their genomes. For random segregation, we develop an ordered strand pair formulation of the dynamics, analogous to the ordered strand pair formalism developed for quasispecies dynamics involving semiconservative replication with imperfect lesion repair (in this context, lesion repair is taken to mean repair of postreplication base-pair mismatches). Interestingly, a similar formulation is possible with immortal strand co-segregation, despite the fact that this segregation mechanism is age dependent. From our model we are able to mathematically show that, when lesion repair is imperfect, then immortal strand co-segregation leads to better preservation of the stem cell lineage than random chromosome segregation. Furthermore, our model allows us to estimate the optimal lesion repair efficiency for preserving an adult stem cell population for a given period of time. For human stem cells, we obtain that mispaired bases still present after replication and cell division should be left untouched, to avoid potentially fixing a mutation in both DNA strands.

Evolutionary dynamics of adult stem cells: Comparison of random and immortal-strand segregation mechanisms

NASA Astrophysics Data System (ADS)

Tannenbaum, Emmanuel; Sherley, James L.; Shakhnovich, Eugene I.

2005-04-01

This paper develops a point-mutation model describing the evolutionary dynamics of a population of adult stem cells. Such a model may prove useful for quantitative studies of tissue aging and the emergence of cancer. We consider two modes of chromosome segregation: (1) random segregation, where the daughter chromosomes of a given parent chromosome segregate randomly into the stem cell and its differentiating sister cell and (2) “immortal DNA strand” co-segregation, for which the stem cell retains the daughter chromosomes with the oldest parent strands. Immortal strand co-segregation is a mechanism, originally proposed by [Cairns Nature (London) 255, 197 (1975)], by which stem cells preserve the integrity of their genomes. For random segregation, we develop an ordered strand pair formulation of the dynamics, analogous to the ordered strand pair formalism developed for quasispecies dynamics involving semiconservative replication with imperfect lesion repair (in this context, lesion repair is taken to mean repair of postreplication base-pair mismatches). Interestingly, a similar formulation is possible with immortal strand co-segregation, despite the fact that this segregation mechanism is age dependent. From our model we are able to mathematically show that, when lesion repair is imperfect, then immortal strand co-segregation leads to better preservation of the stem cell lineage than random chromosome segregation. Furthermore, our model allows us to estimate the optimal lesion repair efficiency for preserving an adult stem cell population for a given period of time. For human stem cells, we obtain that mispaired bases still present after replication and cell division should be left untouched, to avoid potentially fixing a mutation in both DNA strands.

Nonstructural NSs protein of rift valley fever virus interacts with pericentromeric DNA sequences of the host cell, inducing chromosome cohesion and segregation defects.

PubMed

Mansuroglu, Z; Josse, T; Gilleron, J; Billecocq, A; Leger, P; Bouloy, M; Bonnefoy, E

2010-01-01

Rift Valley fever virus (RVFV) is an emerging, highly pathogenic virus; RVFV infection can lead to encephalitis, retinitis, or fatal hepatitis associated with hemorrhagic fever in humans, as well as death, abortions, and fetal deformities in animals. RVFV nonstructural NSs protein, a major factor of the virulence, forms filamentous structures in the nuclei of infected cells. In order to further understand RVFV pathology, we investigated, by chromatin immunoprecipitation, immunofluorescence, fluorescence in situ hybridization, and confocal microscopy, the capacity of NSs to interact with the host genome. Our results demonstrate that even though cellular DNA is predominantly excluded from NSs filaments, NSs interacts with some specific DNA regions of the host genome such as clusters of pericentromeric gamma-satellite sequence. Targeting of these sequences by NSs was correlated with the induction of chromosome cohesion and segregation defects in RVFV-infected murine, as well as sheep cells. Using recombinant nonpathogenic virus rZHDeltaNSs210-230, expressing a NSs protein deleted of its region of interaction with cellular factor SAP30, we showed that the NSs-SAP30 interaction was essential for NSs to target pericentromeric sequences, as well as for induction of chromosome segregation defects. The effect of RVFV upon the inheritance of genetic information is discussed with respect to the pathology associated with fetal deformities and abortions, highlighting the main role played by cellular cofactor SAP30 on the establishment of NSs interactions with host DNA sequences and RVFV pathogenesis.

Eukaryotic and Prokaryotic Cytoskeletons: Structure and Mechanics

NASA Astrophysics Data System (ADS)

Gopinathan, Ajay

2013-03-01

The eukaryotic cytoskeleton is an assembly of filamentous proteins and a host of associated proteins that collectively serve functional needs ranging from spatial organization and transport to the production and transmission of forces. These systems can exhibit a wide variety of non-equilibrium, self-assembled phases depending on context and function. While much recent progress has been made in understanding the self-organization, rheology and nonlinear mechanical properties of such active systems, in this talk, we will concentrate on some emerging aspects of cytoskeletal physics that are promising. One such aspect is the influence of cytoskeletal network topology and its dynamics on both active and passive intracellular transport. Another aspect we will highlight is the interplay between chirality of filaments, their elasticity and their interactions with the membrane that can lead to novel conformational states with functional implications. Finally we will consider homologs of cytoskeletal proteins in bacteria, which are involved in templating cell growth, segregating genetic material and force production, which we will discuss with particular reference to contractile forces during cell division. These prokaryotic structures function in remarkably similar yet fascinatingly different ways from their eukaryotic counterparts and can enrich our understanding of cytoskeletal functioning as a whole.

Effects of atmospheric pressure plasmas on isolated and cellular DNA-a review.

PubMed

Arjunan, Krishna Priya; Sharma, Virender K; Ptasinska, Sylwia

2015-01-29

Atmospheric Pressure Plasma (APP) is being used widely in a variety of biomedical applications. Extensive research in the field of plasma medicine has shown the induction of DNA damage by APP in a dose-dependent manner in both prokaryotic and eukaryotic systems. Recent evidence suggests that APP-induced DNA damage shows potential benefits in many applications, such as sterilization and cancer therapy. However, in several other applications, such as wound healing and dentistry, DNA damage can be detrimental. This review reports on the extensive investigations devoted to APP interactions with DNA, with an emphasis on the critical role of reactive species in plasma-induced damage to DNA. The review consists of three main sections dedicated to fundamental knowledge of the interactions of reactive oxygen species (ROS)/reactive nitrogen species (RNS) with DNA and its components, as well as the effects of APP on isolated and cellular DNA in prokaryotes and eukaryotes.

Repair of 8-oxo-7,8-dihydroguanine in prokaryotic and eukaryotic cells: Properties and biological roles of the Fpg and OGG1 DNA N-glycosylases.

PubMed

Boiteux, Serge; Coste, Franck; Castaing, Bertrand

2017-06-01

Oxidatively damaged DNA results from the attack of sugar and base moieties by reactive oxygen species (ROS), which are formed as byproducts of normal cell metabolism and during exposure to endogenous or exogenous chemical or physical agents. Guanine, having the lowest redox potential, is the DNA base the most susceptible to oxidation, yielding products such as 8-oxo-7,8-dihydroguanine (8-oxoG) and 2-6-diamino-4-hydroxy-5-formamidopyrimidine (FapyG). In DNA, 8-oxoG was shown to be mutagenic yielding GC to TA transversions upon incorporation of dAMP opposite this lesion by replicative DNA polymerases. In prokaryotic and eukaryotic cells, 8-oxoG is primarily repaired by the base excision repair pathway (BER) initiated by a DNA N-glycosylase, Fpg and OGG1, respectively. In Escherichia coli, Fpg cooperates with MutY and MutT to prevent 8-oxoG-induced mutations, the "GO-repair system". In Saccharomyces cerevisiae, OGG1 cooperates with nucleotide excision repair (NER), mismatch repair (MMR), post-replication repair (PRR) and DNA polymerase η to prevent mutagenesis. Human and mouse cells mobilize all these pathways using OGG1, MUTYH (MutY-homolog also known as MYH), MTH1 (MutT-homolog also known as NUDT1), NER, MMR, NEILs and DNA polymerases η and λ, to prevent 8-oxoG-induced mutations. In fact, mice deficient in both OGG1 and MUTYH develop cancer in different organs at adult age, which points to the critical impact of 8-oxoG repair on genetic stability in mammals. In this review, we will focus on Fpg and OGG1 proteins, their biochemical and structural properties as well as their biological roles. Other DNA N-glycosylases able to release 8-oxoG from damaged DNA in various organisms will be discussed. Finally, we will report on the role of OGG1 in human disease and the possible use of 8-oxoG DNA N-glycosylases as therapeutic targets. Copyright © 2017 Elsevier Inc. All rights reserved.

[Construction and prokaryotic expression of recombinant gene EGFRvIII HBcAg and immunogenicity analysis of the fusion protein].

PubMed

Duan, Xiao-yi; Wang, Jian-sheng; Guo, You-min; Han, Jun-li; Wang, Quan-ying; Yang, Guang-xiao

2007-01-01

To construct recombinant prokaryotic expression plasmid pET28a(+)/c-PEP-3-c and evaluate the immunogenicity of the fusion protein. cDNA fragment encoding PEP-3 was obtained from pGEM-T Easy/PEP-3 and inserted into recombinant plasmid pGEMEX/HBcAg. Then it was subcloned in prokaryotic expression vector and transformed into E.coli BL21(DE3). The fusion protein was expressed by inducing IPTG and purified by Ni(2+)-NTA affinity chromatography. BALB/c mice were immunized with fusion protein and the antibody titre was determined by indirect ELISA. The recombinant gene was confirmed to be correct by restriction enzyme digestion and DNA sequencing. After prokaryotic expression, fusion protein existed in sediment and accounted for 56% of all bacterial lysate. The purified product accounted for 92% of all protein and its concentration was 8 g/L. The antibody titre in blood serum reached 1:16 000 after the fourth immunization and reached 1:2.56x10(5) after the sixth immunization. The titre of anti-PEP-3 antibody reached 1:1.28x10(5) and the titre of anti-HBcAg antibody was less than 1:4x10(3). Fusion gene PEP-3-HBcAg is highly expressed in E.coli BL21. The expressed fusion protein can induce neutralizing antibody with high titer and specificity, which lays a foundation for the study of genetically engineering vaccine for malignant tumors with the high expression of EGFRvIII.

Further evidence that aberrant segregation and crossing over in Sordaria brevicollis may be discrete, though associated, events.

PubMed

Theivendirarajah, K; Whitehouse, H L

1983-01-01

Crosses were made between buff spore colour mutants in Sordaria brevicollis in the presence of flanking markers. Recombinant asci with one or more wild-type spores were isolated and the spores germinated and scored for buff and flanking marker genotype. The buff genotype was determined by back-crossing to each parent and looking for recombinants. It was found that the majority of the recombinant asci had aberrant segregation at one or other mutant site but not both. It was inferred that in the recombinants hybrid DNA rarely extended to both sites. When the aberrant segregation was associated with crossing-over, the crossovers were situated at either end of the gene rather than between the allelic sites where the hybrid DNA was believed to terminate. Thus, some of the crossovers were separated from the site of the aberrant segregation by a site apparently not involved in hybrid DNA and none was in the position predicted by the Meselson-Radding model, that is, where the hybrid DNA terminates.

Higher 5-hydroxymethylcytosine identifies immortal DNA strand chromosomes in asymmetrically self-renewing distributed stem cells.

PubMed

Huh, Yang Hoon; Cohen, Justin; Sherley, James L

2013-10-15

Immortal strands are the targeted chromosomal DNA strands of nonrandom sister chromatid segregation, a mitotic chromosome segregation pattern unique to asymmetrically self-renewing distributed stem cells (DSCs). By nonrandom segregation, immortal DNA strands become the oldest DNA strands in asymmetrically self-renewing DSCs. Nonrandom segregation of immortal DNA strands may limit DSC mutagenesis, preserve DSC fate, and contribute to DSC aging. The mechanisms responsible for specification and maintenance of immortal DNA strands are unknown. To discover clues to these mechanisms, we investigated the 5-methylcytosine and 5-hydroxymethylcytosine (5hmC) content on chromosomes in mouse hair follicle DSCs during nonrandom segregation. Although 5-methylcytosine content did not differ significantly, the relative content of 5hmC was significantly higher in chromosomes containing immortal DNA strands than in opposed mitotic chromosomes containing younger mortal DNA strands. The difference in relative 5hmC content was caused by the loss of 5hmC from mortal chromosomes. These findings implicate higher 5hmC as a specific molecular determinant of immortal DNA strand chromosomes. Because 5hmC is an intermediate during DNA demethylation, we propose a ten-eleven translocase enzyme mechanism for both the specification and maintenance of nonrandomly segregated immortal DNA strands. The proposed mechanism reveals a means by which DSCs "know" the generational age of immortal DNA strands. The mechanism is supported by molecular expression data and accounts for the selection of newly replicated DNA strands when nonrandom segregation is initiated. These mechanistic insights also provide a possible basis for another characteristic property of immortal DNA strands, their guanine ribonucleotide dependency.

DNA synthesis arrest sites at the right terminus of rat long interspersed repeated (LINE or L1Rn) DNA family members.

PubMed Central

d'Ambrosio, E; Furano, A V

1987-01-01

An approximately equal to 150-bp GC-rich (approximately equal to 60%) region is at the right end of rat long interspersed repeated DNA (LINE or L1Rn) family members. We report here that one of the DNA strands from this region contains several non-palindromic sites that strongly arrest DNA synthesis in vitro by the prokaryotic Klenow and T4 DNA polymerases, the eukaryotic alpha polymerase, and AMV reverse transcriptase. The strongest arrest sites are G-rich (approximately equal to 70%) homopurine stretches of 18 or more residues. Shorter homopurine stretches (12 residues or fewer) did not arrest DNA synthesis even if the stretch contains 11/12 G residues. Arrest of the prokaryotic polymerases was not affected by their respective single strand binding proteins or polymerase accessory proteins. The region of duplex DNA which contains DNA synthesis arrest sites reacts with bromoacetaldehyde when present in negatively supercoiled molecules. By contrast, homopurine stretches that do not arrest DNA synthesis do not react with bromoacetaldehyde. The presence of bromoacetaldehyde-reactive bases in a G-rich homopurine-containing duplex under torsional stress is thought to be caused by base stacking in the homopurine strand. Therefore, we suggest that base-stacked regions of the template arrest DNA synthesis. Images PMID:2436148

Capturing prokaryotic dark matter genomes.

PubMed

Gasc, Cyrielle; Ribière, Céline; Parisot, Nicolas; Beugnot, Réjane; Defois, Clémence; Petit-Biderre, Corinne; Boucher, Delphine; Peyretaillade, Eric; Peyret, Pierre

2015-12-01

Prokaryotes are the most diverse and abundant cellular life forms on Earth. Most of them, identified by indirect molecular approaches, belong to microbial dark matter. The advent of metagenomic and single-cell genomic approaches has highlighted the metabolic capabilities of numerous members of this dark matter through genome reconstruction. Thus, linking functions back to the species has revolutionized our understanding of how ecosystem function is sustained by the microbial world. This review will present discoveries acquired through the illumination of prokaryotic dark matter genomes by these innovative approaches. Copyright © 2015 Institut Pasteur. Published by Elsevier Masson SAS. All rights reserved.

Distribution of putative xenogeneic silencers in prokaryote genomes.

PubMed

Perez-Rueda, Ernesto; Ibarra, J Antonio

2015-10-01

Gene silencing is an important function as it keeps newly acquired foreign DNA repressed, thereby avoiding possible deleterious effects in the host organism. Known transcriptional regulators associated with this process are called xenogeneic silencers (XS) and belong to either the H-NS, Lsr2, MvaT or Rok families. In the work described here we looked for XS-like regulators and their distribution in prokaryotic organisms was evaluated. Our analysis showed that putative XS regulators similar to H-NS, Lsr2, MvaT or Rok are present only in bacteria (31.7%). This does not exclude the existence of alternative XS in the rest of the organisms analyzed. Additionally, of the four XS groups evaluated in this work, those from the H-NS family have diversified more than the other groups. In order to compare the distribution of these putative XS regulators we also searched for other nucleoid-associated proteins (NAPs) not included in this group such as Fis, EbfC/YbaB, HU/IHF and Alba. Results showed that NAPs from the Fis, EbfC/YbaB, HU/IHF and Alba families are widely (94%) distributed among prokaryotes. These NAPs were found in multiple combinations with or without XS-like proteins. In regard with XS regulators, results showed that only XS proteins from one family were found in those organisms containing them. This suggests specificity for this type of regulators and their corresponding genomes. Copyright © 2015 Elsevier Ltd. All rights reserved.

Heterozygous SSBP1 start loss mutation co-segregates with hearing loss and the m.1555A>G mtDNA variant in a large multigenerational family.

PubMed

Kullar, Peter J; Gomez-Duran, Aurora; Gammage, Payam A; Garone, Caterina; Minczuk, Michal; Golder, Zoe; Wilson, Janet; Montoya, Julio; Häkli, Sanna; Kärppä, Mikko; Horvath, Rita; Majamaa, Kari; Chinnery, Patrick F

2018-01-01

The m.1555A>G mtDNA variant causes maternally inherited deafness, but the reasons for the highly variable clinical penetrance are not known. Exome sequencing identified a heterozygous start loss mutation in SSBP1, encoding the single stranded binding protein 1 (SSBP1), segregating with hearing loss in a multi-generational family transmitting m.1555A>G, associated with mtDNA depletion and multiple deletions in skeletal muscle. The SSBP1 mutation reduced steady state SSBP1 levels leading to a perturbation of mtDNA metabolism, likely compounding the intra-mitochondrial translation defect due to m.1555A>G in a tissue-specific manner. This family demonstrates the importance of rare trans-acting genetic nuclear modifiers in the clinical expression of mtDNA disease. © The Author (2017). Published by Oxford University Press on behalf of the Guarantors of Brain.

Higher 5-hydroxymethylcytosine identifies immortal DNA strand chromosomes in asymmetrically self-renewing distributed stem cells

PubMed Central

Huh, Yang Hoon; Cohen, Justin; Sherley, James L.

2013-01-01

Immortal strands are the targeted chromosomal DNA strands of nonrandom sister chromatid segregation, a mitotic chromosome segregation pattern unique to asymmetrically self-renewing distributed stem cells (DSCs). By nonrandom segregation, immortal DNA strands become the oldest DNA strands in asymmetrically self-renewing DSCs. Nonrandom segregation of immortal DNA strands may limit DSC mutagenesis, preserve DSC fate, and contribute to DSC aging. The mechanisms responsible for specification and maintenance of immortal DNA strands are unknown. To discover clues to these mechanisms, we investigated the 5-methylcytosine and 5-hydroxymethylcytosine (5hmC) content on chromosomes in mouse hair follicle DSCs during nonrandom segregation. Although 5-methylcytosine content did not differ significantly, the relative content of 5hmC was significantly higher in chromosomes containing immortal DNA strands than in opposed mitotic chromosomes containing younger mortal DNA strands. The difference in relative 5hmC content was caused by the loss of 5hmC from mortal chromosomes. These findings implicate higher 5hmC as a specific molecular determinant of immortal DNA strand chromosomes. Because 5hmC is an intermediate during DNA demethylation, we propose a ten-eleven translocase enzyme mechanism for both the specification and maintenance of nonrandomly segregated immortal DNA strands. The proposed mechanism reveals a means by which DSCs “know” the generational age of immortal DNA strands. The mechanism is supported by molecular expression data and accounts for the selection of newly replicated DNA strands when nonrandom segregation is initiated. These mechanistic insights also provide a possible basis for another characteristic property of immortal DNA strands, their guanine ribonucleotide dependency. PMID:24082118

[Role and alterations of DNA methylation during the aging and cancer].

PubMed

Szigeti, Krisztina Andrea; Galamb, Orsolya; Kalmár, Alexandra; Barták, Barbara Kinga; Nagy, Zsófia Brigitta; Márkus, Eszter; Igaz, Péter; Tulassay, Zsolt; Molnár, Béla

2018-01-01

Besides the genetic research, increasing number of scientific studies focus on epigenetic phenomena - such as DNA methylation - regulating the expression of genes behind the phenotype, thus can be related to the pathomechanism of several diseases. In this review, we aim to summarize the current knowledge about the evolutionary appearance and functional diversity of DNA methylation as one of the epigenetic mechanisms and to demonstrate its role in aging and cancerous diseases. DNA methylation is also characteristic/also appear to prokaryotes, eukaryotes and viruses. In prokaryotes and viruses, it provides defence mechanisms against extragenous DNA. DNA methylation in prokaryotes plays a significant role in the regulation of transcription, the initiation of replication and in Dam-directed mismatch repair. In viruses, it participates not only in defence mechanisms, but in the assembly of capsids as well which is necessary for spreading. In eukaryotes, DNA methylation is involved in recombination, replication, X chromosome inactivation, transposon control, regulation of chromatin structure and transcription, and it also contributes to the imprinting phenomenon. Besides the above-mentioned aspects, DNA methylation also has an evolutionary role as it can change DNA mutation rate. Global hypomethylation appearing during aging and in cancerous diseases can lead to genetic instablility and spontaneous mutations through its role in the regulation of transposable elements. Local hypermethylated alterations such as hypermethylation of SFRP1, SFRP2, DKK1 and APC gene promoters can cause protein expression changes, thus contribute to development of cancer phenotype. DNA methylation alterations during aging in cancerous diseases support the importance of epigenetic research focusing on disease diagnostics and prognostics. Orv Hetil. 2018; 159(1): 3-15.

A Proposed Genus Boundary for the Prokaryotes Based on Genomic Insights

PubMed Central

Qin, Qi-Long; Xie, Bin-Bin; Zhang, Xi-Ying; Chen, Xiu-Lan; Zhou, Bai-Cheng; Zhou, Jizhong; Oren, Aharon

2014-01-01

Genomic information has already been applied to prokaryotic species definition and classification. However, the contribution of the genome sequence to prokaryotic genus delimitation has been less studied. To gain insights into genus definition for the prokaryotes, we attempted to reveal the genus-level genomic differences in the current prokaryotic classification system and to delineate the boundary of a genus on the basis of genomic information. The average nucleotide sequence identity between two genomes can be used for prokaryotic species delineation, but it is not suitable for genus demarcation. We used the percentage of conserved proteins (POCP) between two strains to estimate their evolutionary and phenotypic distance. A comprehensive genomic survey indicated that the POCP can serve as a robust genomic index for establishing the genus boundary for prokaryotic groups. Basically, two species belonging to the same genus would share at least half of their proteins. In a specific lineage, the genus and family/order ranks showed slight or no overlap in terms of POCP values. A prokaryotic genus can be defined as a group of species with all pairwise POCP values higher than 50%. Integration of whole-genome data into the current taxonomy system can provide comprehensive information for prokaryotic genus definition and delimitation. PMID:24706738

Evolution of RNA- and DNA-guided antivirus defense systems in prokaryotes and eukaryotes: common ancestry vs convergence.

PubMed

Koonin, Eugene V

2017-02-10

Complementarity between nucleic acid molecules is central to biological information transfer processes. Apart from the basal processes of replication, transcription and translation, complementarity is also employed by multiple defense and regulatory systems. All cellular life forms possess defense systems against viruses and mobile genetic elements, and in most of them some of the defense mechanisms involve small guide RNAs or DNAs that recognize parasite genomes and trigger their inactivation. The nucleic acid-guided defense systems include prokaryotic Argonaute (pAgo)-centered innate immunity and CRISPR-Cas adaptive immunity as well as diverse branches of RNA interference (RNAi) in eukaryotes. The archaeal pAgo machinery is the direct ancestor of eukaryotic RNAi that, however, acquired additional components, such as Dicer, and enormously diversified through multiple duplications. In contrast, eukaryotes lack any heritage of the CRISPR-Cas systems, conceivably, due to the cellular toxicity of some Cas proteins that would get activated as a result of operon disruption in eukaryotes. The adaptive immunity function in eukaryotes is taken over partly by the PIWI RNA branch of RNAi and partly by protein-based immunity. In this review, I briefly discuss the interplay between homology and analogy in the evolution of RNA- and DNA-guided immunity, and attempt to formulate some general evolutionary principles for this ancient class of defense systems. This article was reviewed by Mikhail Gelfand and Bojan Zagrovic.

Do the same traffic rules apply? Directional chromosome segregation by SpoIIIE and FtsK.

PubMed

Besprozvannaya, Marina; Burton, Briana M

2014-08-01

Over a decade of studies have tackled the question of how FtsK/SpoIIIE translocases establish and maintain directional DNA translocation during chromosome segregation in bacteria. FtsK/SpoIIIE translocases move DNA in a highly processive, directional manner, where directionality is facilitated by sequences on the substrate DNA molecules that are being transported. In recent years, structural, biochemical, single-molecule and high-resolution microscopic studies have provided new insight into the mechanistic details of directional DNA segregation. Out of this body of work, a series of models have emerged and, ultimately, yielded two seemingly opposing models: the loading model and the target search model. We review these recent mechanistic insights into directional DNA movement and discuss the data that may serve to unite these suggested models, as well as propose future directions that may ultimately solve the debate. © 2014 John Wiley & Sons Ltd.

ASM Conference on Prokaryotic Development

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kaplan, H. B.

2005-07-13

Support was provided by DOE for the 2nd ASM Conference on Prokaryotic Development. The final conference program and abstracts book is attached. The conference presentations are organized around topics that are central to the current research areas in prokaryotic development. The program starts with topics that involve relatively simple models systems and ends with systems that are more complex. The topics are: i) the cell cycle, ii) the cytoskeleton, iii) morphogenesis, iv) developmental transcription, v) signaling, vi) multicellularity, and vii) developmental diversity and symbiosis. The best-studied prokaryotic development model systems will be highlighted at the conference through research presentations bymore » leaders in the field. Many of these systems are also model systems of relevance to the DOE mission including carbon sequestration (Bradyrizobium, Synechococcus), energy production (Anabaena, Rhodobacter) and bioremediation (Caulobacter, Mesorhizobium). In addition, many of the highlighted organisms have important practical applications; the actinomycetes and myxobacteria produce antimicrobials that are of commercial interest. It is certain that the cutting-edge science presented at the conference will be applicable to the large group of bacteria relevant to the DOE mission.« less

An Estimate of the Total DNA in the Biosphere

PubMed Central

Landenmark, Hanna K. E.; Forgan, Duncan H.; Cockell, Charles S.

2015-01-01

Modern whole-organism genome analysis, in combination with biomass estimates, allows us to estimate a lower bound on the total information content in the biosphere: 5.3 × 1031 (±3.6 × 1031) megabases (Mb) of DNA. Given conservative estimates regarding DNA transcription rates, this information content suggests biosphere processing speeds exceeding yottaNOPS values (1024 Nucleotide Operations Per Second). Although prokaryotes evolved at least 3 billion years before plants and animals, we find that the information content of prokaryotes is similar to plants and animals at the present day. This information-based approach offers a new way to quantify anthropogenic and natural processes in the biosphere and its information diversity over time. PMID:26066900

An Estimate of the Total DNA in the Biosphere.

PubMed

Landenmark, Hanna K E; Forgan, Duncan H; Cockell, Charles S

2015-06-01

Modern whole-organism genome analysis, in combination with biomass estimates, allows us to estimate a lower bound on the total information content in the biosphere: 5.3 × 1031 (±3.6 × 1031) megabases (Mb) of DNA. Given conservative estimates regarding DNA transcription rates, this information content suggests biosphere processing speeds exceeding yottaNOPS values (1024 Nucleotide Operations Per Second). Although prokaryotes evolved at least 3 billion years before plants and animals, we find that the information content of prokaryotes is similar to plants and animals at the present day. This information-based approach offers a new way to quantify anthropogenic and natural processes in the biosphere and its information diversity over time.

Tumor malignancy is engaged to prokaryotic homolog toolbox.

PubMed

Fernandes, Janaina; Guedes, Patrícia G; Lage, Celso Luiz S; Rodrigues, Juliany Cola F; Lage, Claudia de Alencar S

2012-04-01

Cancer cells display high proliferation rates and survival provided by high glycolysis, chemoresistance and radioresistance, metabolic features that appear to be activated with malignancy, and seemed to have arisen as early in evolution as in unicellular/prokaryotic organisms. Based on these assumptions, we hypothesize that aggressive phenotypes found in malignant cells may be related to acquired unicellular behavior, launched within a tumor when viral and prokaryotic homologs are overexpressed performing likely robust functions. The ensemble of these expressed viral and prokaryotic close homologs in the proteome of a tumor tissue gives them advantage over normal cells. To assess the hypothesis validity, sequences of human proteins involved in apoptosis, energetic metabolism, cell mobility and adhesion, chemo- and radio-resistance were aligned to homologs present in other life forms, excluding all eukaryotes, using PSI-BLAST, with further corroboration from data available in the literature. The analysis revealed that selected sequences of proteins involved in apoptosis and tumor suppression (as p53 and pRB) scored non-significant (E-value>0.001) with prokaryotic homologs; on the other hand, human proteins involved in cellular chemo- and radio-resistance scored highly significant with prokaryotic and viral homologs (as catalase, E-value=zero). We inferred that such upregulated and/or functionally activated proteins in aggressive malignant cells represent a toolbox of modern human homologs evolved from a similar key set that have granted survival of ancient prokaryotes against extremely harsh environments. According to what has been discussed along this analysis, high mutation rates usually hit hotspots in important conserved protein domains, allowing uncontrolled expansion of more resistant, death-evading malignant clones. That is the case of point mutations in key viral proteins affording viruses escape to chemotherapy, and human homologs of such retroviral

Driving Apart and Segregating Genomes in Archaea.

PubMed

Barillà, Daniela

2016-12-01

Genome segregation is a fundamental biological process in organisms from all domains of life. How this stage of the cell cycle unfolds in Eukarya has been clearly defined and considerable progress has been made to unravel chromosome partition in Bacteria. The picture is still elusive in Archaea. The lineages of this domain exhibit different cell-cycle lifestyles and wide-ranging chromosome copy numbers, fluctuating from 1 up to 55. This plurality of patterns suggests that a variety of mechanisms might underpin disentangling and delivery of DNA molecules to daughter cells. Here I describe recent developments in archaeal genome maintenance, including investigations of novel genome segregation machines that point to unforeseen bacterial and eukaryotic connections. Crown Copyright © 2016. Published by Elsevier Ltd. All rights reserved.

Prokaryotic Argonautes - variations on the RNA interference theme.

PubMed

van der Oost, John; Swarts, Daan C; Jore, Matthijs M

2014-04-15

The discovery of RNA interference (RNAi) has been a major scientific breakthrough. This RNA-guided RNA interference system plays a crucial role in a wide range of regulatory and defense mechanisms in eukaryotes. The key enzyme of the RNAi system is Argonaute (Ago), an endo-ribonuclease that uses a small RNA guide molecule to specifically target a complementary RNA transcript. Two functional classes of eukaryotic Ago have been described: catalytically active Ago that cleaves RNA targets complementary to its guide, and inactive Ago that uses its guide to bind target RNA to down-regulate translation efficiency. A recent comparative genomics study has revealed that Argonaute-like proteins are also encoded by prokaryotic genomes. Interestingly, there is a lot of variation among these prokaryotic Argonaute (pAgo) proteins with respect to domain architecture: some resemble the eukaryotic Ago (long pAgo) containing a complete or disrupted catalytic site, while others are truncated versions (short pAgo) that generally contain an incomplete catalytic site. Prokaryotic Agos with an incomplete catalytic site often co-occur with (predicted) nucleases. Based on this diversity, and on the fact that homologs of other RNAi-related protein components (such as Dicer nucleases) have never been identified in prokaryotes, it has been predicted that variations on the eukaryotic RNAi theme may occur in prokaryotes.

Prokaryotic Argonautes - variations on the RNA interference theme

PubMed Central

van der Oost, John; Swarts, Daan C.; Jore, Matthijs M.

2014-01-01

The discovery of RNA interference (RNAi) has been a major scientific breakthrough. This RNA-guided RNA interference system plays a crucial role in a wide range of regulatory and defense mechanisms in eukaryotes. The key enzyme of the RNAi system is Argonaute (Ago), an endo-ribonuclease that uses a small RNA guide molecule to specifically target a complementary RNA transcript. Two functional classes of eukaryotic Ago have been described: catalytically active Ago that cleaves RNA targets complementary to its guide, and inactive Ago that uses its guide to bind target RNA to down-regulate translation efficiency. A recent comparative genomics study has revealed that Argonaute-like proteins are also encoded by prokaryotic genomes. Interestingly, there is a lot of variation among these prokaryotic Argonaute (pAgo) proteins with respect to domain architecture: some resemble the eukaryotic Ago (long pAgo) containing a complete or disrupted catalytic site, while others are truncated versions (short pAgo) that generally contain an incomplete catalytic site. Prokaryotic Agos with an incomplete catalytic site often co-occur with (predicted) nucleases. Based on this diversity, and on the fact that homologs of other RNAi-related protein components (such as Dicer nucleases) have never been identified in prokaryotes, it has been predicted that variations on the eukaryotic RNAi theme may occur in prokaryotes. PMID:28357239

Prokaryotic functional diversity in different biogeochemical depth zones in tidal sediments of the Severn Estuary, UK, revealed by stable-isotope probing.

PubMed

Webster, Gordon; Rinna, Joachim; Roussel, Erwan G; Fry, John C; Weightman, Andrew J; Parkes, R John

2010-05-01

Stable isotope probing of prokaryotic DNA was used to determine active prokaryotes using (13)C-labelled substrates (glucose, acetate, CO(2)) in sediment slurries from different biogeochemical zones of the Severn Estuary, UK. Multiple, low concentrations (5 x 100 microM) of (13)C-substrate additions and short-term incubations (7 days) were used to minimize changes in the prokaryotic community, while achieving significant (13)C-incorporation. Analysis demonstrated clear metabolic activity within all slurries, although neither the net sulphate removal nor CH(4) production occurred in the anaerobic sulphate reduction and methanogenesis zone slurries. Some similarities occurred in the prokaryotic populations that developed in different sediment slurries, particularly in the aerobic and dysaerobic zone slurries with (13)C-glucose, which were dominated by Gammaproteobacteria and Marine Group 1 Archaea, whereas both anaerobic sediment slurries incubated with (13)C-acetate showed incorporation into Epsilonproteobacteria and other bacteria, with the sulphate reduction zone slurry also showing (13)C-acetate utilization by Miscellaneous Crenarchaeotic Group Archaea. The lower potential energy methanogenesis zone slurries were the only conditions where no (13)C-incorporation into Archaea occurred, despite Bacteria being labelled; this was surprising because Archaea have been suggested to be adapted to low-energy conditions. Overall, our results highlight that uncultured prokaryotes play important ecological roles in tidal sediments of the Severn Estuary, providing new metabolic information for novel groups of Archaea and suggesting broader metabolisms for largely uncultivated Bacteria.

Energetics and genetics across the prokaryote-eukaryote divide

PubMed Central

2011-01-01

Background All complex life on Earth is eukaryotic. All eukaryotic cells share a common ancestor that arose just once in four billion years of evolution. Prokaryotes show no tendency to evolve greater morphological complexity, despite their metabolic virtuosity. Here I argue that the eukaryotic cell originated in a unique prokaryotic endosymbiosis, a singular event that transformed the selection pressures acting on both host and endosymbiont. Results The reductive evolution and specialisation of endosymbionts to mitochondria resulted in an extreme genomic asymmetry, in which the residual mitochondrial genomes enabled the expansion of bioenergetic membranes over several orders of magnitude, overcoming the energetic constraints on prokaryotic genome size, and permitting the host cell genome to expand (in principle) over 200,000-fold. This energetic transformation was permissive, not prescriptive; I suggest that the actual increase in early eukaryotic genome size was driven by a heavy early bombardment of genes and introns from the endosymbiont to the host cell, producing a high mutation rate. Unlike prokaryotes, with lower mutation rates and heavy selection pressure to lose genes, early eukaryotes without genome-size limitations could mask mutations by cell fusion and genome duplication, as in allopolyploidy, giving rise to a proto-sexual cell cycle. The side effect was that a large number of shared eukaryotic basal traits accumulated in the same population, a sexual eukaryotic common ancestor, radically different to any known prokaryote. Conclusions The combination of massive bioenergetic expansion, release from genome-size constraints, and high mutation rate favoured a protosexual cell cycle and the accumulation of eukaryotic traits. These factors explain the unique origin of eukaryotes, the absence of true evolutionary intermediates, and the evolution of sex in eukaryotes but not prokaryotes. Reviewers This article was reviewed by: Eugene Koonin, William Martin

Stepping stones towards a new prokaryotic taxonomy.

PubMed

Gevers, Dirk; Dawyndt, Peter; Vandamme, Peter; Willems, Anne; Vancanneyt, Marc; Swings, Jean; De Vos, Paul

2006-11-29

Technological developments provide new insights into prokaryotic evolution and diversity and provoke a continuous need to update taxonomy and revise classification schemes. Our present species concept and definition are being challenged by the growing amount of whole genomic information, which should allow improvements in the natural species definition. The continuous quest for an objective and stable method for sorting strains into coherent homogeneous groups is inherent to prokaryotic systematics and nomenclature. Morphological, biochemical, physiological, phenotypic and chemotaxonomic criteria have been complemented by molecular data and pragmatic, purpose built, species definitions are being replaced by more natural ones based on evolutionary insights. It is imperative to give due consideration to both fundamental and applied aspects of future species concepts and definitions. The present paper discusses the present practice in prokaryotic taxonomy of how this system developed and how it may evolve in the future.

[Prokaryotic expression and immunological activity of human neutrophil gelatinase associated lipocalin].

PubMed

Wu, Jianwei; Cai, Lei; Qian, Wei; Jiao, Liyuan; Li, Jiangfeng; Song, Xiaoli; Wang, Jihua

2015-07-01

To construct a prokaryotic expression vector of human neutrophil gelatinase associated lipocalin (NGAL) and identify the bioactivity of the fusion protein. The cDNA of human NGAL obtained from GenBank was linked to a cloning vector to construct the prokaryotic expression vector pCold-NGAL. Then the vector was transformed into E.coli BL21(DE3) plysS. Under the optimal induction condition, the recombinant NGAL (rNGAL) was expressed and purified by Ni Sepharose 6 Fast Flow affinity chromatography. The purity and activity of the rNGAL were respectively identified by SDS-PAGE and Western blotting combined with NGAL reagent (Latex enhanced immunoturbidimetry). Restriction enzyme digestion and nucleotide sequencing proved that the expression vector pCold-NGAL was successfully constructed. Under the optimal induction condition that we determined, the rNGAL was expressed in soluble form in E.coli BL21(DE3) plysS. The relative molecular mass of the rNGAL was 25 000, and its purity was more than 98.0%. Furthermore, Western blotting and immunoturbidimetry indicated that the rNGAL reacted with NGAL mAb specifically. Human rNGAL of high purity and bioactivity was successfully constructed in E.coli BL21(DE3) plysS using the expression vector pCold-NGAL.

Prokaryotic responses to hydrostatic pressure in the ocean--a review.

PubMed

Tamburini, Christian; Boutrif, Mehdi; Garel, Marc; Colwell, Rita R; Deming, Jody W

2013-05-01

Effects of hydrostatic pressure on pure cultures of prokaryotes have been studied extensively but impacts at the community level in the ocean are less well defined. Here we consider hydrostatic pressure effects on natural communities containing both unadapted (piezosensitive) prokaryotes originating from surface water and adapted (including piezophilic) prokaryotes from the deep sea. Results from experiments mimicking pressure changes experienced by particle-associated prokaryotes during their descent through the water column show that rates of degradation of organic matter (OM) by surface-originating microorganisms decrease with sinking. Analysis of a much larger data set shows that, under stratified conditions, deep-sea communities adapt to in situ conditions of high pressure, low temperature and low OM. Measurements made using decompressed samples and atmospheric pressure thus underestimate in situ activity. Exceptions leading to overestimates can be attributed to deep mixing events, large influxes of surface particles, or provision of excessive OM during experimentation. The sediment-water interface, where sinking particles accumulate, will be populated by a mixture of piezosensitive, piezotolerant and piezophilic prokaryotes, with piezophilic activity prevailing deeper within sediment. A schematic representation of how pressure shapes prokaryotic communities in the ocean is provided, allowing a reasonably accurate interpretation of the available activity measurements. © 2013 Society for Applied Microbiology and Blackwell Publishing Ltd.

"Breaking up is hard to do": the formation and resolution of sister chromatid intertwines.

PubMed

Baxter, Jonathan

2015-02-13

The absolute necessity to resolve every intertwine between the two strands of the DNA double helix provides a massive challenge to the cellular processes that duplicate and segregate chromosomes. Although the overwhelming majority of intertwines between the parental DNA strands are resolved during DNA replication, there are numerous chromosomal contexts where some intertwining is maintained into mitosis. These mitotic sister chromatid intertwines (SCIs) can be found as; short regions of unreplicated DNA, fully replicated and intertwined sister chromatids--commonly referred to as DNA catenation--and as sister chromatid linkages generated by homologous recombination-associated processes. Several overlapping mechanisms, including intra-chromosomal compaction, topoisomerase action and Holliday junction resolvases, ensure that all SCIs are removed before they can prevent normal chromosome segregation. Here, I discuss why some DNA intertwines persist into mitosis and review our current knowledge of the SCI resolution mechanisms that are employed in both prokaryotes and eukaryotes, including how deregulating SCI formation during DNA replication or disrupting the resolution processes may contribute to aneuploidy in cancer. Copyright © 2014 Elsevier Ltd. All rights reserved.

Death by Segregation: Does the Dimension of Racial Segregation Matter?

PubMed

Yang, Tse-Chuan; Matthews, Stephen A

2015-01-01

The county-level geographic mortality differentials have persisted in the past four decades in the United States (US). Though several socioeconomic factors (e.g., inequality) partially explain this phenomenon, the role of race/ethnic segregation, in general, and the different dimensions of segregation, more specifically, has been underexplored. Focusing on all-cause age-sex standardized US county-level mortality (2004-2008), this study has two substantive goals: (1) to understand whether segregation is a determinant of mortality and if yes, how the relationship between segregation and mortality varies by racial/ethnic dyads (e.g., white/black), and (2) to explore whether different dimensions of segregation (i.e., evenness, exposure, concentration, centralization, and clustering) are associated with mortality. A third goal is methodological: to assess whether spatial autocorrelation influences our understanding of the associations between the dimensions of segregation and mortality. Race/ethnic segregation was found to contribute to the geographic mortality disparities. Moreover, the relationship with mortality differed by both race/ethnic group and the dimension of segregation. Specifically, white/black segregation is positively related to mortality, whereas the segregation between whites and non-black minorities is negatively associated with mortality. Among the five dimensions of segregation, evenness and exposure are more strongly related to mortality than other dimensions. Spatial filtering approaches also identified six unique spatial patterns that significantly affect the spatial distribution of mortality. These patterns offer possible insights that help identify omitted variables related to the persistent patterning of mortality in the US.

The Diversity of Prokaryotic DDE Transposases of the Mutator Superfamily, Insertion Specificity, and Association with Conjugation Machineries

PubMed Central

Guérillot, Romain; Siguier, Patricia; Gourbeyre, Edith; Chandler, Michael; Glaser, Philippe

2014-01-01

Transposable elements (TEs) are major components of both prokaryotic and eukaryotic genomes and play a significant role in their evolution. In this study, we have identified new prokaryotic DDE transposase families related to the eukaryotic Mutator-like transposases. These genes were retrieved by cascade PSI-Blast using as initial query the transposase of the streptococcal integrative and conjugative element (ICE) TnGBS2. By combining secondary structure predictions and protein sequence alignments, we predicted the DDE catalytic triad and the DNA-binding domain recognizing the terminal inverted repeats. Furthermore, we systematically characterized the organization and the insertion specificity of the TEs relying on these prokaryotic Mutator-like transposases (p-MULT) for their mobility. Strikingly, two distant TE families target their integration upstream σA dependent promoters. This allowed us to identify a transposase sequence signature associated with this unique insertion specificity and to show that the dissymmetry between the two inverted repeats is responsible for the orientation of the insertion. Surprisingly, while DDE transposases are generally associated with small and simple transposons such as insertion sequences (ISs), p-MULT encoding TEs show an unprecedented diversity with several families of IS, transposons, and ICEs ranging in size from 1.1 to 52 kb. PMID:24418649

The New Segregation.

ERIC Educational Resources Information Center

Laosa, Luis M.

2001-01-01

This issue reviews national demographic trends in school segregation, summarizing research findings. Though the national debate on school segregation emphasizes blacks and whites, present-day school segregation includes segregation by socioeconomic level, ethnicity, and native language. The research study examined features of the ecology of…

Comparison of six methods for the recovery of PCR-compatible microbial DNA from an agricultural biogas plant.

PubMed

Stagnati, L; Soffritti, G; Lanubile, A; Busconi, M

2017-05-01

Six different commercial methods were compared to evaluate their efficiency in recovering high quantity/quality PCR compatible microbial DNA from an agricultural biogas plant. Within the last two decades, biogas plants have been developed to produce energy from organic wastes and from devoted biomass. The complex biotransformations are performed by a diverse consortium of microorganisms that is an important reserve of genes and enzymatic activities with a huge range of applications in various commercial fields. In this respect, the ability to isolate DNA from a complex matrix is of high importance. Important parameters of the recovered DNA are good yield, purity, and quality. The methods examined showed considerable differences about quantity and quality of the recovered DNA and, usually, it was observed that a higher amount was accompanied by more degradation. DNA purity was determined by its PCR amplificability. Only two methods were able to provide DNA pure enough to be directly amplified. For the rest of the methods, a few intermediate steps such as dilution and/or the addition of polyvinylpyrrolidone were necessary to remove the inhibitors present and to amplify the DNA. Real-time PCR analysis evidenced that, as expected, prokaryotic DNA was much more abundant than eukaryotic DNA, but some methods were more suited to recovering prokaryotic or eukaryotic DNA. The digestion analysis of ribosomal DNA amplicons confirmed the influence of the methods on the final output, allowing the recovery of only a fraction of the present species as determined by sequencing a small prokaryotic and eukaryotic ribosomal library.

Response of the rhizosphere prokaryotic community of barley (Hordeum vulgare L.) to elevated atmospheric CO2 concentration in open-top chambers.

PubMed

Szoboszlay, Márton; Näther, Astrid; Mitterbauer, Esther; Bender, Jürgen; Weigel, Hans-Joachim; Tebbe, Christoph C

2017-08-01

The effect of elevated atmospheric CO 2 concentration [CO 2 ] on the diversity and composition of the prokaryotic community inhabiting the rhizosphere of winter barley (Hordeum vulgare L.) was investigated in a field experiment, using open-top chambers. Rhizosphere samples were collected at anthesis (flowering stage) from six chambers with ambient [CO 2 ] (approximately 400 ppm) and six chambers with elevated [CO 2 ] (700 ppm). The V4 region of the 16S rRNA gene was PCR-amplified from the extracted DNA and sequenced on an Illumina MiSeq instrument. Above-ground plant biomass was not affected by elevated [CO 2 ] at anthesis, but plants exposed to elevated [CO 2 ] had significantly higher grain yield. The composition of the rhizosphere prokaryotic communities was very similar under ambient and elevated [CO 2 ]. The dominant taxa were Bacteroidetes, Actinobacteria, Alpha-, Gamma-, and Betaproteobacteria. Elevated [CO 2 ] resulted in lower prokaryotic diversity in the rhizosphere, but did not cause a significant difference in community structure. © 2017 The Authors. MicrobiologyOpen published by John Wiley & Sons Ltd.

Flexible DNA bending in HU–DNA cocrystal structures

PubMed Central

Swinger, Kerren K.; Lemberg, Kathryn M.; Zhang, Ying; Rice, Phoebe A.

2003-01-01

HU and IHF are members of a family of prokaryotic proteins that interact with the DNA minor groove in a sequence-specific (IHF) or non-specific (HU) manner to induce and/or stabilize DNA bending. HU plays architectural roles in replication initiation, transcription regulation and site-specific recombination, and is associated with bacterial nucleoids. Cocrystal structures of Anabaena HU bound to DNA (1P71, 1P78, 1P51) reveal that while underlying proline intercalation and asymmetric charge neutralization mechanisms of DNA bending are similar for IHF and HU, HU stabilizes different DNA bend angles (∼105–140°). The two bend angles within a single HU complex are not coplanar, and the resulting dihedral angle is consistent with negative supercoiling. Comparison of HU–DNA and IHF–DNA structures suggests that sharper bending is correlated with longer DNA binding sites and smaller dihedral angles. An HU-induced bend may be better modeled as a hinge, not a rigid bend. The ability to induce or stabilize varying bend angles is consistent with HU’s role as an architectural cofactor in many different systems that may require differing geometries. PMID:12853489

Viral Regulation of Prokaryotic Carbon Metabolism in a Hypereutrophic Freshwater Reservoir Ecosystem (Villerest, France)

PubMed Central

Pradeep Ram, Angia Sriram; Colombet, Jonathan; Perriere, Fanny; Thouvenot, Antoine; Sime-Ngando, Télesphore

2016-01-01

The current consensus concerning the viral regulation of prokaryotic carbon metabolism is less well-studied, compared to substrate availability. We explored the seasonal and vertical distribution of viruses and its relative influence on prokaryotic carbon metabolism in a hypereutrophic reservoir, Lake Villerest (France). Flow cytometry and transmission electron microscopy (TEM) analyses to determine viral abundance (VA; range = 6.1–63.5 × 107 ml-1) and viral infection rates of prokaryotes (range = 5.3–32%) respectively suggested that both the parameters varied more significantly with depths than with seasons. Prokaryotic growth efficiency (PGE, considered as a proxy of prokaryotic carbon metabolism) calculated from prokaryotic production and respiration measurements (PGE = prokaryotic production/[prokaryotic production + prokaryotic respiration] × 100) varied from 14 to 80% across seasons and depths. Viruses through selective lyses had antagonistic impacts on PGE by regulating key prokaryotic metabolic processes (i.e., production and respiration). Higher viral lysis accompanied by higher respiration rates and lower PGE in the summer (mean = 22.9 ± 10.3%) than other seasons (mean = 59.1 ± 18.6%), led to significant loss of carbon through bacterial-viral loop and shifted the reservoir system to net heterotrophy. Our data therefore suggests that the putative adverse impact of viruses on the growth efficiency of the prokaryotic community can have strong implications on nutrient flux patterns and on the overall ecosystem metabolism in anthropogenic dominated aquatic systems such as Lake Villerest. PMID:26903963

The Role of Residential Segregation in Contemporary School Segregation

ERIC Educational Resources Information Center

Frankenberg, Erica

2013-01-01

Inaction to address housing segregation in metropolitan areas has resulted in persistently high levels of residential segregation. As the Supreme Court has recently limited school districts' voluntary integration efforts, this article considers the role of residential segregation in maintaining racially isolated schools, namely what is known about…

Coupling of soil prokaryotic diversity and plant diversity across latitudinal forest ecosystems

NASA Astrophysics Data System (ADS)

Wang, Jun-Tao; Zheng, Yuan-Ming; Hu, Hang-Wei; Li, Jing; Zhang, Li-Mei; Chen, Bao-Dong; Chen, Wei-Ping; He, Ji-Zheng

2016-01-01

The belowground soil prokaryotic community plays a cardinal role in sustaining the stability and functions of forest ecosystems. Yet, the nature of how soil prokaryotic diversity co-varies with aboveground plant diversity along a latitudinal gradient remains elusive. By establishing three hundred 400-m2 quadrats from tropical rainforest to boreal forest in a large-scale parallel study on both belowground soil prokaryote and aboveground tree and herb communities, we found that soil prokaryotic diversity couples with the diversity of herbs rather than trees. The diversity of prokaryotes and herbs responds similarly to environmental factors along the latitudinal gradient. These findings revealed that herbs provide a good predictor of belowground biodiversity in forest ecosystems, and provide new perspective on the aboveground and belowground interactions in forest ecosystems.

Coupling of soil prokaryotic diversity and plant diversity across latitudinal forest ecosystems.

PubMed

Wang, Jun-Tao; Zheng, Yuan-Ming; Hu, Hang-Wei; Li, Jing; Zhang, Li-Mei; Chen, Bao-Dong; Chen, Wei-Ping; He, Ji-Zheng

2016-01-19

The belowground soil prokaryotic community plays a cardinal role in sustaining the stability and functions of forest ecosystems. Yet, the nature of how soil prokaryotic diversity co-varies with aboveground plant diversity along a latitudinal gradient remains elusive. By establishing three hundred 400-m(2) quadrats from tropical rainforest to boreal forest in a large-scale parallel study on both belowground soil prokaryote and aboveground tree and herb communities, we found that soil prokaryotic diversity couples with the diversity of herbs rather than trees. The diversity of prokaryotes and herbs responds similarly to environmental factors along the latitudinal gradient. These findings revealed that herbs provide a good predictor of belowground biodiversity in forest ecosystems, and provide new perspective on the aboveground and belowground interactions in forest ecosystems.

Variation of mutational burden in healthy human tissues suggests non-random strand segregation and allows measuring somatic mutation rates.

PubMed

Werner, Benjamin; Sottoriva, Andrea

2018-06-01

The immortal strand hypothesis poses that stem cells could produce differentiated progeny while conserving the original template strand, thus avoiding accumulating somatic mutations. However, quantitating the extent of non-random DNA strand segregation in human stem cells remains difficult in vivo. Here we show that the change of the mean and variance of the mutational burden with age in healthy human tissues allows estimating strand segregation probabilities and somatic mutation rates. We analysed deep sequencing data from healthy human colon, small intestine, liver, skin and brain. We found highly effective non-random DNA strand segregation in all adult tissues (mean strand segregation probability: 0.98, standard error bounds (0.97,0.99)). In contrast, non-random strand segregation efficiency is reduced to 0.87 (0.78,0.88) in neural tissue during early development, suggesting stem cell pool expansions due to symmetric self-renewal. Healthy somatic mutation rates differed across tissue types, ranging from 3.5 × 10-9/bp/division in small intestine to 1.6 × 10-7/bp/division in skin.

DNA recognition by an RNA-guided bacterial Argonaute

PubMed Central

Doudna, Jennifer A.

2017-01-01

Argonaute (Ago) proteins are widespread in prokaryotes and eukaryotes and share a four-domain architecture capable of RNA- or DNA-guided nucleic acid recognition. Previous studies identified a prokaryotic Argonaute protein from the eubacterium Marinitoga piezophila (MpAgo), which binds preferentially to 5′-hydroxylated guide RNAs and cleaves single-stranded RNA (ssRNA) and DNA (ssDNA) targets. Here we present a 3.2 Å resolution crystal structure of MpAgo bound to a 21-nucleotide RNA guide and a complementary 21-nucleotide ssDNA substrate. Comparison of this ternary complex to other target-bound Argonaute structures reveals a unique orientation of the N-terminal domain, resulting in a straight helical axis of the entire RNA-DNA heteroduplex through the central cleft of the protein. Additionally, mismatches introduced into the heteroduplex reduce MpAgo cleavage efficiency with a symmetric profile centered around the middle of the helix. This pattern differs from the canonical mismatch tolerance of other Argonautes, which display decreased cleavage efficiency for substrates bearing sequence mismatches to the 5′ region of the guide strand. This structural analysis of MpAgo bound to a hybrid helix advances our understanding of the diversity of target recognition mechanisms by Argonaute proteins. PMID:28520746

Overcoming Triple Segregation

ERIC Educational Resources Information Center

Gandara, Patricia

2010-01-01

Latinos are, after whites, the most segregated student group in the United States, and their segregation is closely tied to poor academic outcomes. Latinos experience a triple segregation: by race/ethnicity, poverty, and language. Racial segregation perpetuates negative stereotypes, reduces the likelihood of a strong teaching staff, and is often…

ISEScan: automated identification of insertion sequence elements in prokaryotic genomes.

PubMed

Xie, Zhiqun; Tang, Haixu

2017-11-01

The insertion sequence (IS) elements are the smallest but most abundant autonomous transposable elements in prokaryotic genomes, which play a key role in prokaryotic genome organization and evolution. With the fast growing genomic data, it is becoming increasingly critical for biology researchers to be able to accurately and automatically annotate ISs in prokaryotic genome sequences. The available automatic IS annotation systems are either providing only incomplete IS annotation or relying on the availability of existing genome annotations. Here, we present a new IS elements annotation pipeline to address these issues. ISEScan is a highly sensitive software pipeline based on profile hidden Markov models constructed from manually curated IS elements. ISEScan performs better than existing IS annotation systems when tested on prokaryotic genomes with curated annotations of IS elements. Applying it to 2784 prokaryotic genomes, we report the global distribution of IS families across taxonomic clades in Archaea and Bacteria. ISEScan is implemented in Python and released as an open source software at https://github.com/xiezhq/ISEScan. hatang@indiana.edu. Supplementary data are available at Bioinformatics online. © The Author 2017. Published by Oxford University Press. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com

Vertical distribution of the prokaryotic cell size in the Mediterranean Sea

NASA Astrophysics Data System (ADS)

La Ferla, R.; Maimone, G.; Azzaro, M.; Conversano, F.; Brunet, C.; Cabral, A. S.; Paranhos, R.

2012-12-01

Distributions of prokaryotic cell size and morphology were studied in different areas of the Mediterranean Sea by using image analysis on samples collected from surface down to bathypelagic layers (max depth 4,900 m) in the Southern Tyrrhenian, Southern Adriatic and Eastern Mediterranean Seas. Distribution of cell size of prokaryotes in marine ecosystem is very often not considered, which makes our study first in the context of prokaryotic ecology. In the deep Mediterranean layers, an usually-not-considered form of carbon sequestration through prokaryotic cells has been highlighted, which is consistent with an increase in cell size with the depth of the water column. A wide range in prokaryotic cell volumes was observed (between 0.045 and 0.566 μm3). Increase in cell size with depth was opposed to cell abundance distribution. Our results from microscopic observations were confirmed by the increasing HNA/LNA ratio (HNA, cells with high nucleic acid content; LNA, cells with low nucleic acid content) along the water column. Implications of our results on the increasing cell size with depth are in the fact that the quantitative estimation of prokaryotic biomass changes along the water column and the amount of carbon sequestered in the deep biota is enhanced.

A novel prokaryotic promoter identified in the genome of some monopartite begomoviruses.

PubMed

Wang, Wei-Chen; Hsu, Yau-Heiu; Lin, Na-Sheng; Wu, Chia-Ying; Lai, Yi-Chin; Hu, Chung-Chi

2013-01-01

Geminiviruses are known to exhibit both prokaryotic and eukaryotic features in their genomes, with the ability to express their genes and even replicate in bacterial cells. We have demonstrated previously the existence of unit-length single-stranded circular DNAs of Ageratum yellow vein virus (AYVV, a species in the genus Begomovirus, family Geminiviridae) in Escherichia coli cells, which prompted our search for unknown prokaryotic functions in the begomovirus genomes. By using a promoter trapping strategy, we identified a novel prokaryotic promoter, designated AV3 promoter, in nts 762-831 of the AYVV genome. Activity assays revealed that the AV3 promoter is strong, unidirectional, and constitutive, with an endogenous downstream ribosome binding site and a translatable short open reading frame of eight amino acids. Sequence analyses suggested that the AV3 promoter might be a remnant of prokaryotic ancestors that could be related to certain promoters of bacteria from marine or freshwater environments. The discovery of the prokaryotic AV3 promoter provided further evidence for the prokaryotic origin in the evolutionary history of geminiviruses.

A Novel Prokaryotic Promoter Identified in the Genome of Some Monopartite Begomoviruses

PubMed Central

Wang, Wei-Chen; Hsu, Yau-Heiu; Lin, Na-Sheng; Wu, Chia-Ying; Lai, Yi-Chin; Hu, Chung-Chi

2013-01-01

Geminiviruses are known to exhibit both prokaryotic and eukaryotic features in their genomes, with the ability to express their genes and even replicate in bacterial cells. We have demonstrated previously the existence of unit-length single-stranded circular DNAs of Ageratum yellow vein virus (AYVV, a species in the genus Begomovirus, family Geminiviridae) in Escherichia coli cells, which prompted our search for unknown prokaryotic functions in the begomovirus genomes. By using a promoter trapping strategy, we identified a novel prokaryotic promoter, designated AV3 promoter, in nts 762-831 of the AYVV genome. Activity assays revealed that the AV3 promoter is strong, unidirectional, and constitutive, with an endogenous downstream ribosome binding site and a translatable short open reading frame of eight amino acids. Sequence analyses suggested that the AV3 promoter might be a remnant of prokaryotic ancestors that could be related to certain promoters of bacteria from marine or freshwater environments. The discovery of the prokaryotic AV3 promoter provided further evidence for the prokaryotic origin in the evolutionary history of geminiviruses. PMID:23936138

Light-controlled motility in prokaryotes and the problem of directional light perception

PubMed Central

Wilde, Annegret

2017-01-01

Abstract The natural light environment is important to many prokaryotes. Most obviously, phototrophic prokaryotes need to acclimate their photosynthetic apparatus to the prevailing light conditions, and such acclimation is frequently complemented by motility to enable cells to relocate in search of more favorable illumination conditions. Non-phototrophic prokaryotes may also seek to avoid light at damaging intensities and wavelengths, and many prokaryotes with diverse lifestyles could potentially exploit light signals as a rich source of information about their surroundings and a cue for acclimation and behavior. Here we discuss our current understanding of the ways in which bacteria can perceive the intensity, wavelength and direction of illumination, and the signal transduction networks that link light perception to the control of motile behavior. We discuss the problems of light perception at the prokaryotic scale, and the challenge of directional light perception in small bacterial cells. We explain the peculiarities and the common features of light-controlled motility systems in prokaryotes as diverse as cyanobacteria, purple photosynthetic bacteria, chemoheterotrophic bacteria and haloarchaea. PMID:29077840

Light-controlled motility in prokaryotes and the problem of directional light perception.

PubMed

Wilde, Annegret; Mullineaux, Conrad W

2017-11-01

The natural light environment is important to many prokaryotes. Most obviously, phototrophic prokaryotes need to acclimate their photosynthetic apparatus to the prevailing light conditions, and such acclimation is frequently complemented by motility to enable cells to relocate in search of more favorable illumination conditions. Non-phototrophic prokaryotes may also seek to avoid light at damaging intensities and wavelengths, and many prokaryotes with diverse lifestyles could potentially exploit light signals as a rich source of information about their surroundings and a cue for acclimation and behavior. Here we discuss our current understanding of the ways in which bacteria can perceive the intensity, wavelength and direction of illumination, and the signal transduction networks that link light perception to the control of motile behavior. We discuss the problems of light perception at the prokaryotic scale, and the challenge of directional light perception in small bacterial cells. We explain the peculiarities and the common features of light-controlled motility systems in prokaryotes as diverse as cyanobacteria, purple photosynthetic bacteria, chemoheterotrophic bacteria and haloarchaea. © FEMS 2017.

The prokaryotic FAD synthetase family: a potential drug target.

PubMed

Serrano, Ana; Ferreira, Patricia; Martínez-Júlvez, Marta; Medina, Milagros

2013-01-01

Disruption of cellular production of the flavin cofactors, flavin adenine mononucleotide (FMN) and flavin adenine dinucleotide(FAD) will prevent the assembly of a large number of flavoproteins and flavoenzymes involved in key metabolic processes in all types of organisms. The enzymes responsible for FMN and FAD production in prokaryotes and eukaryotes exhibit various structural characteristics to catalyze the same chemistry, a fact that converts the prokaryotic FAD synthetase (FADS) in a potential drug target for the development of inhibitors endowed with anti-pathogenic activity. The first step before searching for selective inhibitors of FADS is to understand the structural and functional mechanisms for the riboflavin kinase and FMN adenylyltransferase activities of the prokaryotic enzyme, and particularly to identify their differential functional characteristics with regard to the enzymes performing similar functions in other organisms, particularly humans. In this paper, an overview of the current knowledge of the structure-function relationships in prokaryotic FADS has been presented, as well as of the state of the art in the use of these enzymes as drug targets.

Top-Down Control of Diesel-Degrading Prokaryotic Communities.

PubMed

Sauret, Caroline; Böttjer, Daniela; Talarmin, Agathe; Guigue, Catherine; Conan, Pascal; Pujo-Pay, Mireille; Ghiglione, Jean-François

2015-08-01

Biostimulation through the addition of inorganic nutrients has been the most widely practiced bioremediation strategy in oil-polluted marine waters. However, little attention has so far been paid to the microbial food web and the impact of top-down control that directly or indirectly influences the success of the bioremediation. We designed a mesocosm experiment using pre-filtered (<50 μm) surface seawater from the Bay of Banyuls-sur-Mer (North-Western Mediterranean Sea) and examined the top-down effect exerted by heterotrophic nanoflagellates (HNF) and virus-like particles (VLP) on prokaryotic abundance, activity and diversity in the presence or absence of diesel fuel. Prokaryotes, HNF and VLP abundances showed a predator-prey succession, with a co-development of HNF and VLP. In the polluted system, we observed a stronger impact of viral lysis on prokaryotic abundances than in the control. Analysis of the diversity revealed that a bloom of Vibrio sp. occurred in the polluted mesocosm. That bloom was rapidly followed by a less abundant and more even community of predation-resistant bacteria, including known hydrocarbon degraders such as Oleispira spp. and Methylophaga spp. and opportunistic bacteria such as Percisivirga spp., Roseobacter spp. and Phaeobacter spp. The shift in prokaryotic dominance in response to viral lysis provided clear evidence of the 'killing the winner' model. Nevertheless, despite clear effects on prokaryotic abundance, activity and diversity, the diesel degradation was not impacted by top-down control. The present study investigates for the first time the functioning of a complex microbial network (including VLP) using a nutrient-based biostimulation strategy and highlights some key processes useful for tailoring bioremediation.

A prokaryotic viral sequence is expressed and conserved in mammalian brain.

PubMed

Yeh, Yang-Hui; Gunasekharan, Vignesh; Manuelidis, Laura

2017-07-03

A natural and permanent transfer of prokaryotic viral sequences to mammals has not been reported by others. Circular "SPHINX" DNAs <5 kb were previously isolated from nuclease-protected cytoplasmic particles in rodent neuronal cell lines and brain. Two of these DNAs were sequenced after Φ29 polymerase amplification, and they revealed significant but imperfect homology to segments of commensal Acinetobacter phage viruses. These findings were surprising because the brain is isolated from environmental microorganisms. The 1.76-kb DNA sequence (SPHINX 1.8), with an iteron before its ORF, was evaluated here for its expression in neural cells and brain. A rabbit affinity purified antibody generated against a peptide without homology to mammalian sequences labeled a nonglycosylated ∼41-kDa protein (spx1) on Western blots, and the signal was efficiently blocked by the competing peptide. Spx1 was resistant to limited proteinase K digestion, but was unrelated to the expression of host prion protein or its pathologic amyloid form. Remarkably, spx1 concentrated in selected brain synapses, such as those on anterior motor horn neurons that integrate many complex neural inputs. SPHINX 1.8 appears to be involved in tissue-specific differentiation, including essential functions that preserve its propagation during mammalian evolution, possibly via maternal inheritance. The data here indicate that mammals can share and exchange a larger world of prokaryotic viruses than previously envisioned.

Microbial rRNA: rDNA gene ratios may be unexpectedly low due to extracellular DNA preservation in soils

USDA-ARS?s Scientific Manuscript database

We tested a method of estimating the activity of detectable individual bacterial and archaeal OTUs within a community by calculating ratios of absolute 16S rRNA to rDNA copy numbers. We investigated phylogenetically coherent patterns of activity among soil prokaryotes in non-growing soil communitie...

Transplantation of prokaryotic two-component signaling pathways into mammalian cells.

PubMed

Hansen, Jonathan; Mailand, Erik; Swaminathan, Krishna Kumar; Schreiber, Joerg; Angelici, Bartolomeo; Benenson, Yaakov

2014-11-04

Signaling pathway engineering is a promising route toward synthetic biological circuits. Histidine-aspartate phosphorelays are thought to have evolved in prokaryotes where they form the basis for two-component signaling. Tyrosine-serine-threonine phosphorelays, exemplified by MAP kinase cascades, are predominant in eukaryotes. Recently, a prokaryotic two-component pathway was implemented in a plant species to sense environmental trinitrotoluene. We reasoned that "transplantation" of two-component pathways into mammalian host could provide an orthogonal and diverse toolkit for a variety of signal processing tasks. Here we report that two-component pathways could be partially reconstituted in mammalian cell culture and used for programmable control of gene expression. To enable this reconstitution, coding sequences of histidine kinase (HK) and response regulator (RR) components were codon-optimized for human cells, whereas the RRs were fused with a transactivation domain. Responsive promoters were furnished by fusing DNA binding sites in front of a minimal promoter. We found that coexpression of HKs and their cognate RRs in cultured mammalian cells is necessary and sufficient to strongly induce gene expression even in the absence of pathways' chemical triggers in the medium. Both loss-of-function and constitutive mutants behaved as expected. We further used the two-component signaling pathways to implement two-input logical AND, NOR, and OR gene regulation. Thus, two-component systems can be applied in different capacities in mammalian cells and their components can be used for large-scale synthetic gene circuits.

Spatial changes in the prokaryotic community structure across a soil catena

NASA Astrophysics Data System (ADS)

Semenov, Mikhail; Zhuravleva, Anna; Tkhakakhova, Azida

2017-04-01

Mesorelief is a complex biogeochemical factor regulating hydrothermal regimes of the surface soil layer, the type of plant cover, etc., and, therefore, influences on soil microbial community structure. A natural model of soil sequence across the slope is a soil catena. Soils forming on various mesorelief positions significantly differ in physicochemical and biological properties, leading to the changes in spatial distribution of various bacterial and archaeal taxa across the soil catena. The aim of this study was to determine soil microbial community structure of different ecosystems corresponding to three mesorelief positions within the soil catena. The catena was located at the right bank of the Oka River (Moscow region, Russian Federation). Soil samples were taken at depths of 0-20 cm, 20-40 cm, and 40-60 cm from three sites within the transect of 960 m with elevation of 80 m, corresponding to the autonomous (AU), transitional (TR) (both Luvisols), and accumulative (AC) (Fluvisol Umbric) positions of the landscape. The dominant vegetation of studied sites were rootstock- and loose bunchgrasses of the fallow ecosystem (AU), a secondary small-leaved forest of the forest ecosystem (TR), and a meadow-bog association of the meadow-bog ecosystem (AC). The distances between the sites were 680 m (AU and TR), and 280 m (TR and AC). The soil samples were homogenized, and the total community DNA of three replicates was extracted using the FastDNA® SPIN kit for Soil. All DNA replicates were combined in a pooled sample and the DNA was used for PCR with specific primers for the 16S V3 and V4 regions. The products were purified and submitted to Illumina MiSeq sequencing. Obtained sequence data were evaluated using the MiSeq Reporter Metagenomics Workflow and QIIME. Quantification of the bacterial and archaeal metabolically active cells was quantified by the FISH-method. Verrucomicrobia, Proteobacteria, Firmictutes and Actinobacteria were the major phyla in autonomous site

Meiosis I chromosome segregation is established through regulation of microtubule–kinetochore interactions

PubMed Central

Miller, Matthew P; Ünal, Elçin; Brar, Gloria A; Amon, Angelika

2012-01-01

During meiosis, a single round of DNA replication is followed by two consecutive rounds of nuclear divisions called meiosis I and meiosis II. In meiosis I, homologous chromosomes segregate, while sister chromatids remain together. Determining how this unusual chromosome segregation behavior is established is central to understanding germ cell development. Here we show that preventing microtubule–kinetochore interactions during premeiotic S phase and prophase I is essential for establishing the meiosis I chromosome segregation pattern. Premature interactions of kinetochores with microtubules transform meiosis I into a mitosis-like division by disrupting two key meiosis I events: coorientation of sister kinetochores and protection of centromeric cohesin removal from chromosomes. Furthermore we find that restricting outer kinetochore assembly contributes to preventing premature engagement of microtubules with kinetochores. We propose that inhibition of microtubule–kinetochore interactions during premeiotic S phase and prophase I is central to establishing the unique meiosis I chromosome segregation pattern. DOI: http://dx.doi.org/10.7554/eLife.00117.001 PMID:23275833

Chromosomal instability drives metastasis through a cytosolic DNA response.

PubMed

Bakhoum, Samuel F; Ngo, Bryan; Laughney, Ashley M; Cavallo, Julie-Ann; Murphy, Charles J; Ly, Peter; Shah, Pragya; Sriram, Roshan K; Watkins, Thomas B K; Taunk, Neil K; Duran, Mercedes; Pauli, Chantal; Shaw, Christine; Chadalavada, Kalyani; Rajasekhar, Vinagolu K; Genovese, Giulio; Venkatesan, Subramanian; Birkbak, Nicolai J; McGranahan, Nicholas; Lundquist, Mark; LaPlant, Quincey; Healey, John H; Elemento, Olivier; Chung, Christine H; Lee, Nancy Y; Imielenski, Marcin; Nanjangud, Gouri; Pe'er, Dana; Cleveland, Don W; Powell, Simon N; Lammerding, Jan; Swanton, Charles; Cantley, Lewis C

2018-01-25

Chromosomal instability is a hallmark of cancer that results from ongoing errors in chromosome segregation during mitosis. Although chromosomal instability is a major driver of tumour evolution, its role in metastasis has not been established. Here we show that chromosomal instability promotes metastasis by sustaining a tumour cell-autonomous response to cytosolic DNA. Errors in chromosome segregation create a preponderance of micronuclei whose rupture spills genomic DNA into the cytosol. This leads to the activation of the cGAS-STING (cyclic GMP-AMP synthase-stimulator of interferon genes) cytosolic DNA-sensing pathway and downstream noncanonical NF-κB signalling. Genetic suppression of chromosomal instability markedly delays metastasis even in highly aneuploid tumour models, whereas continuous chromosome segregation errors promote cellular invasion and metastasis in a STING-dependent manner. By subverting lethal epithelial responses to cytosolic DNA, chromosomally unstable tumour cells co-opt chronic activation of innate immune pathways to spread to distant organs.

Differential response of marine flagellate communities to prokaryotic food quality

NASA Astrophysics Data System (ADS)

De Corte, D.; Paredes, G.; Sintes, E.; Herndl, G. J.

2016-02-01

Marine prokaryotes play a major role in the biogeochemical cycles. The main predators of prokaryotes are heterotrophic nanoflagellates (HNF). HNF are thus a major link connecting dissolved organic material through prokaryotic grazing to the higher trophic levels. However, little is known about the grazing specificity of HNF on specific prokaryotic taxa. Bacterial and archaeal microbes may have different nutritive values for the HNF communities, thus affecting growth rates and community composition of HNFs. In this study we investigated the influence of prey food quality on Cafeteria roenbergensis and on a natural HNF community isolated in the northern Adriatic Sea. Two Nitrosopumilus maritimus-related strains isolated from the northern Adriatic Sea (Nitrosopumilus adriaticus, Nitrosopumilus piranensis), two Nitrosococcus strains and two fast growing marine Bacteria (Pseudomonas marina and Marinobacter algicola) were fed to the HNFs. The two fast growing bacterial strains resulted in high growth rates of Cafeteria roenbergensis and the mixed HNF community, while the two Nitrosococcus strains did not. Cafeteria roenbergensis fed on N. adriaticus but it did not graze N. piranensis, suggesting that the subtle metabolic and physiological differences between these two closely related thaumarchaeal strains affect the grazing pressure to which they are exposed. Our study also indicates that prokaryotic community composition influences the composition of the HNF community.

Accessory factors promote AlfA-dependent plasmid segregation by regulating filament nucleation, disassembly, and bundling

PubMed Central

Polka, Jessica K.; Kollman, Justin M.; Mullins, R. Dyche

2014-01-01

In bacteria, some plasmids are partitioned to daughter cells by assembly of actin-like proteins (ALPs). The best understood ALP, ParM, has a core set of biochemical properties that contributes to its function, including dynamic instability, spontaneous nucleation, and bidirectional elongation. AlfA, an ALP that pushes plasmids apart in Bacillus, relies on a different set of underlying properties to segregate DNA. AlfA elongates unidirectionally and is not dynamically unstable; its assembly and disassembly are regulated by a cofactor, AlfB. Free AlfB breaks up AlfA bundles and promotes filament turnover. However, when AlfB is bound to the centromeric DNA sequence, parN, it forms a segrosome complex that nucleates and stabilizes AlfA filaments. When reconstituted in vitro, this system creates polarized, motile comet tails that associate by antiparallel filament bundling to form bipolar, DNA-segregating spindles. PMID:24481252

Diversification of DnaA dependency for DNA replication in cyanobacterial evolution.

PubMed

Ohbayashi, Ryudo; Watanabe, Satoru; Ehira, Shigeki; Kanesaki, Yu; Chibazakura, Taku; Yoshikawa, Hirofumi

2016-05-01

Regulating DNA replication is essential for all living cells. The DNA replication initiation factor DnaA is highly conserved in prokaryotes and is required for accurate initiation of chromosomal replication at oriC. DnaA-independent free-living bacteria have not been identified. The dnaA gene is absent in plastids and some symbiotic bacteria, although it is not known when or how DnaA-independent mechanisms were acquired. Here, we show that the degree of dependency of DNA replication on DnaA varies among cyanobacterial species. Deletion of the dnaA gene in Synechococcus elongatus PCC 7942 shifted DNA replication from oriC to a different site as a result of the integration of an episomal plasmid. Moreover, viability during the stationary phase was higher in dnaA disruptants than in wild-type cells. Deletion of dnaA did not affect DNA replication or cell growth in Synechocystis sp. PCC 6803 or Anabaena sp. PCC 7120, indicating that functional dependency on DnaA was already lost in some nonsymbiotic cyanobacterial lineages during diversification. Therefore, we proposed that cyanobacteria acquired DnaA-independent replication mechanisms before symbiosis and such an ancestral cyanobacterium was the sole primary endosymbiont to form a plastid precursor.

Methylated bases in mycoplasmal DNA.

PubMed Central

Razin, A; Razin, S

1980-01-01

The DNAs of four Mycoplasma and one Acholeplasma species were found to contain methylated bases. All of the five species contained 6-methyladenine (m6Ade), the methylated base characteristic of prokaryotic DNA. The extent of methylation of adenine residues in the mycoplasmal DNA ranged from 0.2% in Mycoplasma capricolum to about 2% in Mycoplasma arginini and Mycoplasma hyorhinis with intermediate methylation values for Mycoplasma orale and Acholeplasma laidlawii DNAs. About 5.8% of the cytosine residues in M. hyorhinis DNA were methylated also. Analysis of cell culture DNA for the presence of m6Ade as a means for detection of contamination by mycoplasmas, and the phylogenetic implications of the finding of methylated bases in mycoplasmal DNAs are discussed. PMID:7433124

Prokaryotic Information Games: How and When to Take up and Secrete DNA.

PubMed

Stingl, Kerstin; Koraimann, Günther

2017-01-01

Besides transduction via bacteriophages natural transformation and bacterial conjugation are the most important mechanisms driving bacterial evolution and horizontal gene spread. Conjugation systems have evolved in eubacteria and archaea. In Gram-positive and Gram-negative bacteria, cell-to-cell DNA transport is typically facilitated by a type IV secretion system (T4SS). T4SSs also mediate uptake of free DNA in Helicobacter pylori, while most transformable bacteria use a type II secretion/type IV pilus system. In this chapter, we focus on how and when bacteria "decide" that such a DNA transport apparatus is to be expressed and assembled in a cell that becomes competent. Development of DNA uptake competence and DNA transfer competence is driven by a variety of stimuli and often involves intricate regulatory networks leading to dramatic changes in gene expression patterns and bacterial physiology. In both cases, genetically homogeneous populations generate a distinct subpopulation that is competent for DNA uptake or DNA transfer or might uniformly switch into competent state. Phenotypic conversion from one state to the other can rely on bistable genetic networks that are activated stochastically with the integration of external signaling molecules. In addition, we discuss principles of DNA uptake processes in naturally transformable bacteria and intend to understand the exceptional use of a T4SS for DNA import in the gastric pathogen H. pylori. Realizing the events that trigger developmental transformation into competence within a bacterial population will eventually help to create novel and effective therapies against the transmission of antibiotic resistances among pathogens.

Global diversity and biogeography of deep-sea pelagic prokaryotes.

PubMed

Salazar, Guillem; Cornejo-Castillo, Francisco M; Benítez-Barrios, Verónica; Fraile-Nuez, Eugenio; Álvarez-Salgado, X Antón; Duarte, Carlos M; Gasol, Josep M; Acinas, Silvia G

2016-03-01

The deep-sea is the largest biome of the biosphere, and contains more than half of the whole ocean's microbes. Uncovering their general patterns of diversity and community structure at a global scale remains a great challenge, as only fragmentary information of deep-sea microbial diversity exists based on regional-scale studies. Here we report the first globally comprehensive survey of the prokaryotic communities inhabiting the bathypelagic ocean using high-throughput sequencing of the 16S rRNA gene. This work identifies the dominant prokaryotes in the pelagic deep ocean and reveals that 50% of the operational taxonomic units (OTUs) belong to previously unknown prokaryotic taxa, most of which are rare and appear in just a few samples. We show that whereas the local richness of communities is comparable to that observed in previous regional studies, the global pool of prokaryotic taxa detected is modest (~3600 OTUs), as a high proportion of OTUs are shared among samples. The water masses appear to act as clear drivers of the geographical distribution of both particle-attached and free-living prokaryotes. In addition, we show that the deep-oceanic basins in which the bathypelagic realm is divided contain different particle-attached (but not free-living) microbial communities. The combination of the aging of the water masses and a lack of complete dispersal are identified as the main drivers for this biogeographical pattern. All together, we identify the potential of the deep ocean as a reservoir of still unknown biological diversity with a higher degree of spatial complexity than hitherto considered.

Global diversity and biogeography of deep-sea pelagic prokaryotes

PubMed Central

Salazar, Guillem; Cornejo-Castillo, Francisco M; Benítez-Barrios, Verónica; Fraile-Nuez, Eugenio; Álvarez-Salgado, X Antón; Duarte, Carlos M; Gasol, Josep M; Acinas, Silvia G

2016-01-01

The deep-sea is the largest biome of the biosphere, and contains more than half of the whole ocean's microbes. Uncovering their general patterns of diversity and community structure at a global scale remains a great challenge, as only fragmentary information of deep-sea microbial diversity exists based on regional-scale studies. Here we report the first globally comprehensive survey of the prokaryotic communities inhabiting the bathypelagic ocean using high-throughput sequencing of the 16S rRNA gene. This work identifies the dominant prokaryotes in the pelagic deep ocean and reveals that 50% of the operational taxonomic units (OTUs) belong to previously unknown prokaryotic taxa, most of which are rare and appear in just a few samples. We show that whereas the local richness of communities is comparable to that observed in previous regional studies, the global pool of prokaryotic taxa detected is modest (~3600 OTUs), as a high proportion of OTUs are shared among samples. The water masses appear to act as clear drivers of the geographical distribution of both particle-attached and free-living prokaryotes. In addition, we show that the deep-oceanic basins in which the bathypelagic realm is divided contain different particle-attached (but not free-living) microbial communities. The combination of the aging of the water masses and a lack of complete dispersal are identified as the main drivers for this biogeographical pattern. All together, we identify the potential of the deep ocean as a reservoir of still unknown biological diversity with a higher degree of spatial complexity than hitherto considered. PMID:26251871

A CO-FISH assay to assess sister chromatid segregation patterns in mitosis of mouse embryonic stem cells.

PubMed

Sauer, Stephan; Burkett, Sandra S; Lewandoski, Mark; Klar, Amar J S

2013-05-01

Sister chromatids contain identical DNA sequence but are chiral with respect to both their helical handedness and their replication history. Emerging evidence from various model organisms suggests that certain stem cells segregate sister chromatids nonrandomly to either maintain genome integrity or to bias cellular differentiation in asymmetric cell divisions. Conventional methods for tracing of old vs. newly synthesized DNA strands generally lack resolution for individual chromosomes and employ halogenated thymidine analogs with profound cytotoxic effects on rapidly dividing cells. Here, we present a modified chromosome orientation fluorescence in situ hybridization (CO-FISH) assay, where identification of individual chromosomes and their replication history is achieved in subsequent hybridization steps with chromosome-specific DNA probes and PNA telomere probes. Importantly, we tackle the issue of BrdU cytotoxicity and show that our method is compatible with normal mouse ES cell biology, unlike a recently published related protocol. Results from our CO-FISH assay show that mitotic segregation of mouse chromosome 7 is random in ES cells, which contrasts previously published results from our laboratory and settles a controversy. Our straightforward protocol represents a useful resource for future studies on chromatid segregation patterns of in vitro-cultured cells from distinct model organisms.

Replication and Transcription of Eukaryotic DNA in Esherichia coli

PubMed Central

Morrow, John F.; Cohen, Stanley N.; Chang, Annie C. Y.; Boyer, Herbert W.; Goodman, Howard M.; Helling, Robert B.

1974-01-01

Fragments of amplified Xenopus laevis DNA, coding for 18S and 28S ribosomal RNA and generated by EcoRI restriction endonuclease, have been linked in vitro to the bacterial plasmid pSC101; and the recombinant molecular species have been introduced into E. coli by transformation. These recombinant plasmids, containing both eukaryotic and prokaryotic DNA, replicate stably in E. coli. RNA isolated from E. coli minicells harboring the plasmids hybridizes to amplified X. laevis rDNA. Images PMID:4600264

NCBI prokaryotic genome annotation pipeline.

PubMed

Tatusova, Tatiana; DiCuccio, Michael; Badretdin, Azat; Chetvernin, Vyacheslav; Nawrocki, Eric P; Zaslavsky, Leonid; Lomsadze, Alexandre; Pruitt, Kim D; Borodovsky, Mark; Ostell, James

2016-08-19

Recent technological advances have opened unprecedented opportunities for large-scale sequencing and analysis of populations of pathogenic species in disease outbreaks, as well as for large-scale diversity studies aimed at expanding our knowledge across the whole domain of prokaryotes. To meet the challenge of timely interpretation of structure, function and meaning of this vast genetic information, a comprehensive approach to automatic genome annotation is critically needed. In collaboration with Georgia Tech, NCBI has developed a new approach to genome annotation that combines alignment based methods with methods of predicting protein-coding and RNA genes and other functional elements directly from sequence. A new gene finding tool, GeneMarkS+, uses the combined evidence of protein and RNA placement by homology as an initial map of annotation to generate and modify ab initio gene predictions across the whole genome. Thus, the new NCBI's Prokaryotic Genome Annotation Pipeline (PGAP) relies more on sequence similarity when confident comparative data are available, while it relies more on statistical predictions in the absence of external evidence. The pipeline provides a framework for generation and analysis of annotation on the full breadth of prokaryotic taxonomy. For additional information on PGAP see https://www.ncbi.nlm.nih.gov/genome/annotation_prok/ and the NCBI Handbook, https://www.ncbi.nlm.nih.gov/books/NBK174280/. Published by Oxford University Press on behalf of Nucleic Acids Research 2016. This work is written by (a) US Government employee(s) and is in the public domain in the US.

Differential impact of lytic viruses on prokaryotic morphopopulations in a tropical estuarine system (Cochin estuary, India).

PubMed

Jasna, Vijayan; Pradeep Ram, Angia Sriram; Parvathi, Ammini; Sime-Ngando, Telesphore

2018-01-01

Our understanding on the importance of viral lysis in the functioning of tropical estuarine ecosystem is limited. This study examines viral infection of prokaryotes and subsequent lysis of cells belonging to different morphotypes across a salinity gradient in monsoon driven estuarine ecosystem (Cochin estuary, India). High standing stock of viruses and prokaryotes accompanied by lytic infection rates in the euryhaline/mesohaline region of the estuary suggests salinity to have an influential role in driving interactions between prokaryotes and viruses. High prokaryotic mortality rates, up to 42% of prokaryote population in the pre-monsoon season is further substantiated by a high virus to prokaryote ratio (VPR), suggesting that maintenance of a high number of viruses is dependent on the most active fraction of bacterioplankton. Although myoviruses were the dominant viral morphotype (mean = 43%) throughout the study period, there was significant variation among prokaryotic morphotypes susceptible to viral infection. Among them, the viral infected short rod prokaryote morphotype with lower burst estimates (mean = 18 viruses prokaryote-1) was dominant (35%) in the dry seasons whereas a substantial increase in cocci forms (30%) infected by viruses with high burst size (mean = 31 viruses prokaryote-1) was evident during the monsoon season. Such preferential infections of prokaryotic morphopopulations with respect to seasons can have a strong and variable impact on the carbon and energy flow in this tropical ecosystem.

Differential impact of lytic viruses on prokaryotic morphopopulations in a tropical estuarine system (Cochin estuary, India)

PubMed Central

Jasna, Vijayan; Pradeep Ram, Angia Sriram; Sime-Ngando, Telesphore

2018-01-01

Our understanding on the importance of viral lysis in the functioning of tropical estuarine ecosystem is limited. This study examines viral infection of prokaryotes and subsequent lysis of cells belonging to different morphotypes across a salinity gradient in monsoon driven estuarine ecosystem (Cochin estuary, India). High standing stock of viruses and prokaryotes accompanied by lytic infection rates in the euryhaline/mesohaline region of the estuary suggests salinity to have an influential role in driving interactions between prokaryotes and viruses. High prokaryotic mortality rates, up to 42% of prokaryote population in the pre-monsoon season is further substantiated by a high virus to prokaryote ratio (VPR), suggesting that maintenance of a high number of viruses is dependent on the most active fraction of bacterioplankton. Although myoviruses were the dominant viral morphotype (mean = 43%) throughout the study period, there was significant variation among prokaryotic morphotypes susceptible to viral infection. Among them, the viral infected short rod prokaryote morphotype with lower burst estimates (mean = 18 viruses prokaryote-1) was dominant (35%) in the dry seasons whereas a substantial increase in cocci forms (30%) infected by viruses with high burst size (mean = 31 viruses prokaryote-1) was evident during the monsoon season. Such preferential infections of prokaryotic morphopopulations with respect to seasons can have a strong and variable impact on the carbon and energy flow in this tropical ecosystem. PMID:29534102

A Wide Range of 3243A>G/tRNALeu(UUR) (MELAS) Mutation Loads May Segregate in Offspring through the Female Germline Bottleneck

PubMed Central

Pallotti, Francesco; Binelli, Giorgio; Fabbri, Raffaella; Valentino, Maria L.; Vicenti, Rossella; Macciocca, Maria; Cevoli, Sabina; Baruzzi, Agostino; DiMauro, Salvatore; Carelli, Valerio

2014-01-01

Segregation of mutant mtDNA in human tissues and through the germline is debated, with no consensus about the nature and size of the bottleneck hypothesized to explain rapid generational shifts in mutant loads. We investigated two maternal lineages with an apparently different inheritance pattern of the same pathogenic mtDNA 3243A>G/tRNALeu(UUR) (MELAS) mutation. We collected blood cells, muscle biopsies, urinary epithelium and hair follicles from 20 individuals, as well as oocytes and an ovarian biopsy from one female mutation carrier, all belonging to the two maternal lineages to assess mutant mtDNA load, and calculated the theoretical germline bottleneck size (number of segregating units). We also evaluated “mother-to-offspring” segregations from the literature, for which heteroplasmy assessment was available in at least three siblings besides the proband. Our results showed that mutation load was prevalent in skeletal muscle and urinary epithelium, whereas in blood cells there was an inverse correlation with age, as previously reported. The histoenzymatic staining of the ovarian biopsy failed to show any cytochrome-c-oxidase defective oocyte. Analysis of four oocytes and one offspring from the same unaffected mother of the first family showed intermediate heteroplasmic mutant loads (10% to 75%), whereas very skewed loads of mutant mtDNA (0% or 81%) were detected in five offspring of another unaffected mother from the second family. Bottleneck size was 89 segregating units for the first mother and 84 for the second. This was remarkably close to 88, the number of “segregating units” in the “mother-to-offspring” segregations retrieved from literature. In conclusion, a wide range of mutant loads may be found in offspring tissues and oocytes, resulting from a similar theoretical bottleneck size. PMID:24805791

Income inequality and income segregation.

PubMed

Reardon, Sean F; Bischoff, Kendra

2011-01-01

This article investigates how the growth in income inequality from 1970 to 2000 affected patterns of income segregation along three dimensions: the spatial segregation of poverty and affluence, race-specific patterns of income segregation, and the geographic scale of income segregation. The evidence reveals a robust relationship between income inequality and income segregation, an effect that is larger for black families than for white families. In addition, income inequality affects income segregation primarily through its effect on the large-scale spatial segregation of affluence rather than by affecting the spatial segregation of poverty or by altering small-scale patterns of income segregation.

Viral and grazer regulation of prokaryotic growth efficiency in temperate freshwater pelagic environments.

PubMed

Pradeep Ram, A S; Colombet, Jonathan; Perriere, Fanny; Thouvenot, Antoine; Sime-Ngando, Telesphore

2015-02-01

In aquatic systems, limited data exists on the impact of mortality forces such as viral lysis and flagellate grazing when seeking to explain factors regulating prokaryotic metabolism. We explored the relative influence of top-down factors (viral lysis and heterotrophic nanoflagellate grazing) on prokaryotic mortality and their subsequent impact on their community metabolism in the euphotic zone of 21 temperate freshwater lakes located in the French Massif Central. Prokaryotic growth efficiency (PGE, index of prokaryotic community metabolism) determined from prokaryotic production and respiration measurements varied from 5 to 74% across the lakes. Viral and potential grazer-induced mortality of prokaryotes had contrasting impact on PGE. Potential flagellate grazing was found to enhance PGE whereas viral lysis had antagonistic impacts on PGE. The average PGE value in the grazing and viral lysis dominated lake water samples was 35.4% (±15.2%) and 17.2% (±8.1%), respectively. Selective viral lysis or flagellate grazing on prokaryotes together with the nature of contrasted substrates released through mortality processes can perhaps explain for the observed variation and differences in PGE among the studied lakes. The influences of such specific top-down processes on PGE can have strong implications on the carbon and nutrient fluxes in freshwater pelagic environments. © FEMS 2015. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

Prokaryotic evolution and the tree of life are two different things

PubMed Central

Bapteste, Eric; O'Malley, Maureen A; Beiko, Robert G; Ereshefsky, Marc; Gogarten, J Peter; Franklin-Hall, Laura; Lapointe, François-Joseph; Dupré, John; Dagan, Tal; Boucher, Yan; Martin, William

2009-01-01

Background The concept of a tree of life is prevalent in the evolutionary literature. It stems from attempting to obtain a grand unified natural system that reflects a recurrent process of species and lineage splittings for all forms of life. Traditionally, the discipline of systematics operates in a similar hierarchy of bifurcating (sometimes multifurcating) categories. The assumption of a universal tree of life hinges upon the process of evolution being tree-like throughout all forms of life and all of biological time. In multicellular eukaryotes, the molecular mechanisms and species-level population genetics of variation do indeed mainly cause a tree-like structure over time. In prokaryotes, they do not. Prokaryotic evolution and the tree of life are two different things, and we need to treat them as such, rather than extrapolating from macroscopic life to prokaryotes. In the following we will consider this circumstance from philosophical, scientific, and epistemological perspectives, surmising that phylogeny opted for a single model as a holdover from the Modern Synthesis of evolution. Results It was far easier to envision and defend the concept of a universal tree of life before we had data from genomes. But the belief that prokaryotes are related by such a tree has now become stronger than the data to support it. The monistic concept of a single universal tree of life appears, in the face of genome data, increasingly obsolete. This traditional model to describe evolution is no longer the most scientifically productive position to hold, because of the plurality of evolutionary patterns and mechanisms involved. Forcing a single bifurcating scheme onto prokaryotic evolution disregards the non-tree-like nature of natural variation among prokaryotes and accounts for only a minority of observations from genomes. Conclusion Prokaryotic evolution and the tree of life are two different things. Hence we will briefly set out alternative models to the tree of life to

Prokaryotic evolution and the tree of life are two different things.

PubMed

Bapteste, Eric; O'Malley, Maureen A; Beiko, Robert G; Ereshefsky, Marc; Gogarten, J Peter; Franklin-Hall, Laura; Lapointe, François-Joseph; Dupré, John; Dagan, Tal; Boucher, Yan; Martin, William

2009-09-29

The concept of a tree of life is prevalent in the evolutionary literature. It stems from attempting to obtain a grand unified natural system that reflects a recurrent process of species and lineage splittings for all forms of life. Traditionally, the discipline of systematics operates in a similar hierarchy of bifurcating (sometimes multifurcating) categories. The assumption of a universal tree of life hinges upon the process of evolution being tree-like throughout all forms of life and all of biological time. In multicellular eukaryotes, the molecular mechanisms and species-level population genetics of variation do indeed mainly cause a tree-like structure over time. In prokaryotes, they do not. Prokaryotic evolution and the tree of life are two different things, and we need to treat them as such, rather than extrapolating from macroscopic life to prokaryotes. In the following we will consider this circumstance from philosophical, scientific, and epistemological perspectives, surmising that phylogeny opted for a single model as a holdover from the Modern Synthesis of evolution. It was far easier to envision and defend the concept of a universal tree of life before we had data from genomes. But the belief that prokaryotes are related by such a tree has now become stronger than the data to support it. The monistic concept of a single universal tree of life appears, in the face of genome data, increasingly obsolete. This traditional model to describe evolution is no longer the most scientifically productive position to hold, because of the plurality of evolutionary patterns and mechanisms involved. Forcing a single bifurcating scheme onto prokaryotic evolution disregards the non-tree-like nature of natural variation among prokaryotes and accounts for only a minority of observations from genomes. Prokaryotic evolution and the tree of life are two different things. Hence we will briefly set out alternative models to the tree of life to study their evolution. Ultimately

Prokaryotic Caspase Homologs: Phylogenetic Patterns and Functional Characteristics Reveal Considerable Diversity

PubMed Central

Asplund-Samuelsson, Johannes; Bergman, Birgitta; Larsson, John

2012-01-01

Caspases accomplish initiation and execution of apoptosis, a programmed cell death process specific to metazoans. The existence of prokaryotic caspase homologs, termed metacaspases, has been known for slightly more than a decade. Despite their potential connection to the evolution of programmed cell death in eukaryotes, the phylogenetic distribution and functions of these prokaryotic metacaspase sequences are largely uncharted, while a few experiments imply involvement in programmed cell death. Aiming at providing a more detailed picture of prokaryotic caspase homologs, we applied a computational approach based on Hidden Markov Model search profiles to identify and functionally characterize putative metacaspases in bacterial and archaeal genomes. Out of the total of 1463 analyzed genomes, merely 267 (18%) were identified to contain putative metacaspases, but their taxonomic distribution included most prokaryotic phyla and a few archaea (Euryarchaeota). Metacaspases were particularly abundant in Alphaproteobacteria, Deltaproteobacteria and Cyanobacteria, which harbor many morphologically and developmentally complex organisms, and a distinct correlation was found between abundance and phenotypic complexity in Cyanobacteria. Notably, Bacillus subtilis and Escherichia coli, known to undergo genetically regulated autolysis, lacked metacaspases. Pfam domain architecture analysis combined with operon identification revealed rich and varied configurations among the metacaspase sequences. These imply roles in programmed cell death, but also e.g. in signaling, various enzymatic activities and protein modification. Together our data show a wide and scattered distribution of caspase homologs in prokaryotes with structurally and functionally diverse sub-groups, and with a potentially intriguing evolutionary role. These features will help delineate future characterizations of death pathways in prokaryotes. PMID:23185476

Viral infections stimulate the metabolism and shape prokaryotic assemblages in submarine mud volcanoes.

PubMed

Corinaldesi, Cinzia; Dell'Anno, Antonio; Danovaro, Roberto

2012-06-01

Mud volcanoes are geological structures in the oceans that have key roles in the functioning of the global ecosystem. Information on the dynamics of benthic viruses and their interactions with prokaryotes in mud volcano ecosystems is still completely lacking. We investigated the impact of viral infection on the mortality and assemblage structure of benthic prokaryotes of five mud volcanoes in the Mediterranean Sea. Mud volcano sediments promote high rates of viral production (1.65-7.89 × 10(9) viruses g(-1) d(-1)), viral-induced prokaryotic mortality (VIPM) (33% cells killed per day) and heterotrophic prokaryotic production (3.0-8.3 μgC g(-1) d(-1)) when compared with sediments outside the mud volcano area. The viral shunt (that is, the microbial biomass converted into dissolved organic matter as a result of viral infection, and thus diverted away from higher trophic levels) provides 49 mgC m(-2) d(-1), thus fuelling the metabolism of uninfected prokaryotes and contributing to the total C budget. Bacteria are the dominant components of prokaryotic assemblages in surface sediments of mud volcanoes, whereas archaea dominate the subsurface sediment layers. Multivariate multiple regression analyses show that prokaryotic assemblage composition is not only dependant on the geochemical features and processes of mud volcano ecosystems but also on synergistic interactions between bottom-up (that is, trophic resources) and top-down (that is, VIPM) controlling factors. Overall, these findings highlight the significant role of the viral shunt in sustaining the metabolism of prokaryotes and shaping their assemblage structure in mud volcano sediments, and they provide new clues for our understanding of the functioning of cold-seep ecosystems.

Control of Smc Coiled Coil Architecture by the ATPase Heads Facilitates Targeting to Chromosomal ParB/parS and Release onto Flanking DNA

PubMed Central

Minnen, Anita; Bürmann, Frank; Wilhelm, Larissa; Anchimiuk, Anna; Diebold-Durand, Marie-Laure; Gruber, Stephan

2016-01-01

Summary Smc/ScpAB promotes chromosome segregation in prokaryotes, presumably by compacting and resolving nascent sister chromosomes. The underlying mechanisms, however, are poorly understood. Here, we investigate the role of the Smc ATPase activity in the recruitment of Smc/ScpAB to the Bacillus subtilis chromosome. We demonstrate that targeting of Smc/ScpAB to ParB/parS loading sites is strictly dependent on engagement of Smc head domains and relies on an open organization of the Smc coiled coils. We find that dimerization of the Smc hinge domain stabilizes closed Smc rods and hinders head engagement as well as chromosomal targeting. Conversely, the ScpAB sub-complex promotes head engagement and Smc rod opening and thereby facilitates recruitment of Smc to parS sites. Upon ATP hydrolysis, Smc/ScpAB is released from loading sites and relocates within the chromosome—presumably through translocation along DNA double helices. Our findings define an intermediate state in the process of chromosome organization by Smc. PMID:26904953

Strand-seq: a unifying tool for studies of chromosome segregation

PubMed Central

Falconer, Ester; Lansdorp, Peter M.

2013-01-01

Non random segregation of sister chromatids has been implicated to help specify daughter cell fate (the Silent Sister Hypothesis [1]) or to protect the genome of long-lived stem cells (the Immortal Strand Hypothesis [2]). The idea that sister chromatids are non-randomly segregated into specific daughter cells is only marginally supported by data in sporadic and often contradictory studies. As a result, the field has moved forward rather slowly. The advent of being able to directly label and differentiate sister chromatids in vivo using fluorescence in situ hybridization [3] was a significant advance for such studies. However, this approach is limited by the need for large tracks of unidirectional repeats on chromosomes and the reliance on quantitative imaging of fluorescent probes and rigorous statistical analysis to discern between the two competing hypotheses. A novel method called Strand-seq which uses next-generation sequencing to assay sister chromatid inheritance patterns independently for each chromosome [4] offers a comprehensive approach to test for non-random segregation. In addition Strand-seq enables studies on the deposition of chromatin marks in relation to DNA replication. This method is expected to help unify the field by testing previous claims of non-random segregation in an unbiased way in many model systems in vitro and in vivo. PMID:23665005

ContEst16S: an algorithm that identifies contaminated prokaryotic genomes using 16S RNA gene sequences.

PubMed

Lee, Imchang; Chalita, Mauricio; Ha, Sung-Min; Na, Seong-In; Yoon, Seok-Hwan; Chun, Jongsik

2017-06-01

Thanks to the recent advancement of DNA sequencing technology, the cost and time of prokaryotic genome sequencing have been dramatically decreased. It has repeatedly been reported that genome sequencing using high-throughput next-generation sequencing is prone to contaminations due to its high depth of sequencing coverage. Although a few bioinformatics tools are available to detect potential contaminations, these have inherited limitations as they only use protein-coding genes. Here we introduce a new algorithm, called ContEst16S, to detect potential contaminations using 16S rRNA genes from genome assemblies. We screened 69 745 prokaryotic genomes from the NCBI Assembly Database using ContEst16S and found that 594 were contaminated by bacteria, human and plants. Of the predicted contaminated genomes, 8 % were not predicted by the existing protein-coding gene-based tool, implying that both methods can be complementary in the detection of contaminations. A web-based service of the algorithm is available at www.ezbiocloud.net/tools/contest16s.

Building a genome engineering toolbox in nonmodel prokaryotic microbes.

PubMed

Freed, Emily; Fenster, Jacob; Smolinski, Sharon L; Walker, Julie; Henard, Calvin A; Gill, Ryan; Eckert, Carrie A

2018-05-11

The realization of a sustainable bioeconomy requires our ability to understand and engineer complex design principles for the development of platform organisms capable of efficient conversion of cheap and sustainable feedstocks (e.g., sunlight, CO 2 , and nonfood biomass) into biofuels and bioproducts at sufficient titers and costs. For model microbes, such as Escherichia coli, advances in DNA reading and writing technologies are driving the adoption of new paradigms for engineering biological systems. Unfortunately, microbes with properties of interest for the utilization of cheap and renewable feedstocks, such as photosynthesis, autotrophic growth, and cellulose degradation, have very few, if any, genetic tools for metabolic engineering. Therefore, it is important to develop "design rules" for building a genetic toolbox for novel microbes. Here, we present an overview of our current understanding of these rules for the genetic manipulation of prokaryotic microbes and the available genetic tools to expand our ability to genetically engineer nonmodel systems. © 2018 Wiley Periodicals, Inc.

Linking host prokaryotic physiology to viral lifestyle dynamics in a temperate freshwater lake (Lake Pavin, France).

PubMed

Palesse, S; Colombet, J; Pradeep Ram, A S; Sime-Ngando, T

2014-11-01

In aquatic ecosystems, fluctuations in environmental conditions and prokaryotic host physiological states can strongly affect the dynamics of viral life strategies. The influence of prokaryote physiology and environmental factors on viral replication cycles (lytic and lysogeny) was investigated from April to September 2011 at three different strata (epi, meta, and hypolimnion) in the mixolimnion of deep volcanic temperate freshwater Lake Pavin (France). Overall, the euphotic region (epi and metalimnion) was more dynamic and showed significant variation in microbial standing stocks, prokaryotic physiological state, and viral life strategies compared to the aphotic hypolimnion which was stable within sampled months. The prokaryotic host physiology as inferred from the nucleic acid content of prokaryotic cells (high or low nucleic acid) was strongly regulated by the chlorophyll concentration. The predominance of the high nucleic acid (HNA) prokaryotes (cells) over low nucleic acid (LNA) prokaryotes (cells) in the spring (HNA/LNA = 1.2) and vice versa in the summer period (HNA/LNA = 0.4) suggest that the natural prokaryotic communities underwent major shifts in their physiological states during investigated time period. The increase in the percentage of inducible lysogenic prokaryotes in the summer period was associated with the switch in the dominance of LNA over HNA cells, which coincided with the periods of strong resource (nutrient) limitation. This supports the idea that lysogeny represents a maintenance strategy for viruses in unproductive or harsh nutrient/host conditions. A negative correlation of percentage of lysogenic prokaryotes with HNA cell abundance and chlorophyll suggest that lysogenic cycle is closely related to prokaryotic cells which are stressed or starved due to unavailability of resources for its growth and activity. Our results provide support to previous findings that changes in prokaryote physiology are critical for the promotion and

Biodegradation of Emiliania huxleyi Aggregates by natural Prokaryotic Communities under Increasing Hydrostatic Pressure.

NASA Astrophysics Data System (ADS)

Riou, V.; Para, J.; Garel, M.; Guigue, C.; Al Ali, B.; Santinelli, C.; Lefèvre, D.; Gattuso, J. P.; Goutx, M.; Panagiotopoulos, C.; Beaufort, L.; Jacquet, S.; Le Moigne, F. A. C.; Tachikawa, K.; Tamburini, C.

2016-02-01

Fluxes of particulate organic carbon (POC) and minerals are positively correlated, suggesting that minerals could enhance the flux of POC into the deep ocean. The so called "ballast effect" posits that minerals could increase sinking particle densities and/or protect the organic matter from heterotrophic degradation. Laboratory controlled experiments on coccolithophorid aggregates under atmospheric pressure show that biogenic calcite both increases particle settling velocities and preserves the organic matter. However, such experiments have yet to include genuine prokaryote rates indicators as well as the effect of increasing pressure. Here, we used the PArticle Sinking Simulator (PASS) to investigate the effect of the increasing pressure on the degradation of Emiliania huxleyi (calcifiers) aggregates. Extra care was taken to obtain culture aggregates with low prokaryotic abundance prior to exposure to natural mesopelagic prokaryotic communities. Particulate organic and inorganic carbon and dissolved organic carbon concentrations were monitored along with the lipid and carbohydrate compositions, as well as prokaryotic community abundance and specific diversity. A control experiment, without natural prokaryotic community addition, indicates that the pressure increase did not have any effect on calcite dissolution observed after ten days. In contrast, the addition of natural prokaryotic community accelerates calcite dissolution under conditions of increasing pressure. Prokaryotic community development and the lipid fraction of E. huxleyi particulate organic carbon are enhanced under increasing pressure. These results suggest that hydrostatic pressure denatures the structural integrity of the carbonate skeleton that protects the cellular organic matter.

The disposition of DNA in Prochloron (Prochlorophyta)

NASA Technical Reports Server (NTRS)

Coleman, A. W.; Lewin, R. A.

1983-01-01

The discovery of both chlorophyll a and b in the prokaryote Prochloron Lewin, a trait otherwise unique to eukaryotic photosynthetic organisms, has stimulated speculation on the possible endosymbiont origins of the plastids of eukaryotic cells. The arrangement of DNA in Prochloron was therefore dyed in situ with the fluorochrome dye DAPI and compared with the plastid DNA of various eukaryote cells. The DNA of Prochloron is found to be clearly different in arrangement and locale from that of blue-green algae. In the great size of its nucleoids and their apparently high DNA content, Prochloron also differs from the plastids of any eukaryotes, with the possible exception of dinoflagellates. Prochloron remains an evolutionary puzzle.

Rapid isolation of novel FK506 binding proteins from multiple organisms using gDNA and cDNA T7 phage display.

PubMed

Piggott, Andrew M; Kriegel, Alison M; Willows, Robert D; Karuso, Peter

2009-10-01

Reverse chemical proteomics using T7 phage display is a powerful technique for identifying cellular receptors of biologically active small molecules. However, to date this method has generally been limited to cDNA libraries constructed from mRNA isolated from eukaryotes. In this paper, we describe the construction of the first prokaryotic T7 phage display libraries from randomly digested Pseudomonas stutzeri and Vibrio fischeri gDNA, as well as a plant cDNA library from Arabidopsis thaliana. We also describe the use of T7 phage display to identify novel proteins from environmental DNA samples using biotinylated FK506 as a model affinity probe.

Centromeric Heterochromatin: The Primordial Segregation Machine

PubMed Central

Bloom, Kerry S.

2014-01-01

Centromeres are specialized domains of heterochromatin that provide the foundation for the kinetochore. Centromeric heterochromatin is characterized by specific histone modifications, a centromere-specific histone H3 variant (CENP-A), and the enrichment of cohesin, condensin, and topo-isomerase II. Centromere DNA varies orders of magnitude in size from 125 bp (budding yeast) to several megabases (human). In metaphase, sister kinetochores on the surface of replicated chromosomes face away from each other, where they establish microtubule attachment and bi-orientation. Despite the disparity in centromere size, the distance between separated sister kinetochores is remarkably conserved (approximately 1 μm) throughout phylogeny. The centromere functions as a molecular spring that resists microtubule-based extensional forces in mitosis. This review explores the physical properties of DNA in order to understand how the molecular spring is built and how it contributes to the fidelity of chromosome segregation. PMID:25251850

RiboDB Database: A Comprehensive Resource for Prokaryotic Systematics.

PubMed

Jauffrit, Frédéric; Penel, Simon; Delmotte, Stéphane; Rey, Carine; de Vienne, Damien M; Gouy, Manolo; Charrier, Jean-Philippe; Flandrois, Jean-Pierre; Brochier-Armanet, Céline

2016-08-01

Ribosomal proteins (r-proteins) are increasingly used as an alternative to ribosomal rRNA for prokaryotic systematics. However, their routine use is difficult because r-proteins are often not or wrongly annotated in complete genome sequences, and there is currently no dedicated exhaustive database of r-proteins. RiboDB aims at fulfilling this gap. This weekly updated comprehensive database allows the fast and easy retrieval of r-protein sequences from publicly available complete prokaryotic genome sequences. The current version of RiboDB contains 90 r-proteins from 3,750 prokaryotic complete genomes encompassing 38 phyla/major classes and 1,759 different species. RiboDB is accessible at http://ribodb.univ-lyon1.fr and through ACNUC interfaces. © The Author 2016. Published by Oxford University Press on behalf of the Society for Molecular Biology and Evolution. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

Viral infections stimulate the metabolism and shape prokaryotic assemblages in submarine mud volcanoes

PubMed Central

Corinaldesi, Cinzia; Dell'Anno, Antonio; Danovaro, Roberto

2012-01-01

Mud volcanoes are geological structures in the oceans that have key roles in the functioning of the global ecosystem. Information on the dynamics of benthic viruses and their interactions with prokaryotes in mud volcano ecosystems is still completely lacking. We investigated the impact of viral infection on the mortality and assemblage structure of benthic prokaryotes of five mud volcanoes in the Mediterranean Sea. Mud volcano sediments promote high rates of viral production (1.65–7.89 × 109 viruses g−1 d−1), viral-induced prokaryotic mortality (VIPM) (33% cells killed per day) and heterotrophic prokaryotic production (3.0–8.3 μgC g−1 d−1) when compared with sediments outside the mud volcano area. The viral shunt (that is, the microbial biomass converted into dissolved organic matter as a result of viral infection, and thus diverted away from higher trophic levels) provides 49 mgC m−2 d−1, thus fuelling the metabolism of uninfected prokaryotes and contributing to the total C budget. Bacteria are the dominant components of prokaryotic assemblages in surface sediments of mud volcanoes, whereas archaea dominate the subsurface sediment layers. Multivariate multiple regression analyses show that prokaryotic assemblage composition is not only dependant on the geochemical features and processes of mud volcano ecosystems but also on synergistic interactions between bottom-up (that is, trophic resources) and top-down (that is, VIPM) controlling factors. Overall, these findings highlight the significant role of the viral shunt in sustaining the metabolism of prokaryotes and shaping their assemblage structure in mud volcano sediments, and they provide new clues for our understanding of the functioning of cold-seep ecosystems. PMID:22170423

DNA recombination protein-dependent mechanism of homoplasmy and its proposed functions.

PubMed

Shibata, Takehiko; Ling, Feng

2007-01-01

Homoplasmy is a basic genetic state of mitochondria, in which all of the hundreds to thousands of mitochondrial (mt)DNA copies within a cell or an individual have the same nucleotide-sequence. It was recently found that "vegetative segregation" to generate homoplasmic cells is an active process under genetic control. In the yeast Saccharomyces cerevisiae, the Mhr1 protein which catalyzes a key reaction in mtDNA homologous recombination, plays a pivotal role in vegetative segregation. Conversely, within the nuclear genome, homologous DNA recombination causes genetic diversity. Considering these contradictory roles of this key reaction in DNA recombination, possible functions of homoplasmy are discussed.

The Prokaryote-Eukaryote Dichotomy: Meanings and Mythology

PubMed Central

Sapp, Jan

2005-01-01

Drawing on documents both published and archival, this paper explains how the prokaryote-eukaryote dichotomy of the 1960s was constructed, the purposes it served, and what it implied in terms of classification and phylogeny. In doing so, I first show how the concept was attributed to Edouard Chatton and the context in which he introduced the terms. Following, I examine the context in which the terms were reintroduced into biology in 1962 by Roger Stanier and C. B. van Niel. I study the discourse over the subsequent decade to understand how the organizational dichotomy took on the form of a natural classification as the kingdom Monera or superkingdom Procaryotae. Stanier and van Niel admitted that, in regard to constructing a natural classification of bacteria, structural characteristics were no more useful than physiological properties. They repeatedly denied that bacterial phylogenetics was possible. I thus examine the great historical irony that the “prokaryote,” in both its organizational and phylogenetic senses, was defined (negatively) on the basis of structure. Finally, we see how phylogenetic research based on 16S rRNA led by Carl Woese and his collaborators confronted the prokaryote concept while moving microbiology to the center of evolutionary biology. PMID:15944457

Effect of hydrostatic pressure on prokaryotic heterotrophic activity in the dark ocean

NASA Astrophysics Data System (ADS)

Amano, C.; Sintes, E.; Utsumi, M.; Herndl, G. J.

2016-02-01

The pioneering work of ZoBell in the 1940s revealed the existence of piezophilic bacteria in the deep ocean, capable of growing only under high-pressure conditions. However, it is still unclear to what extent the bulk prokaryotic community inhabiting the deep ocean is affected by hydrostatic pressure. Essentially, the fractions of the bulk microbial community being piezophilic, piezotolerant and piezosensitive remain unknown. To determine the influence of hydrostatic pressure on the heterotrophic microbial activity, an in situ microbial incubator (ISMI) was deployed in the North Atlantic Ocean at depths down to 3200 m. Natural prokaryotic communities were incubated under both in situ hydrostatic pressure and atmospheric pressure conditions at in situ temperature following the addition of 5 nM 3H-leucine. Bulk leucine incorporation rates and single cell activity assessed by microautoradiography combined with catalyzed reporter deposition fluorescence in situ hybridization (MICRO-CARD-FISH) were determined. Prokaryotic leucine incorporation rates obtained under in situ pressure conditions were generally lower than under atmospheric pressure conditions, suggesting that hydrostatic pressure inhibits overall heterotrophic activity in the deep sea. The ratio of leucine incorporation rates obtained under in situ pressure conditions to atmospheric pressure conditions decreased with depth for the bulk prokaryotic community. Moreover, MICRO-CARD-FISH revealed that specific prokaryotic groups are apparently more affected by hydrostatic pressure than others. Taken together, our results indicate varying sensitivities of prokaryotic groups to hydrostatic pressure.

Endosymbiotic gene transfer from prokaryotic pangenomes: Inherited chimerism in eukaryotes.

PubMed

Ku, Chuan; Nelson-Sathi, Shijulal; Roettger, Mayo; Garg, Sriram; Hazkani-Covo, Einat; Martin, William F

2015-08-18

Endosymbiotic theory in eukaryotic-cell evolution rests upon a foundation of three cornerstone partners--the plastid (a cyanobacterium), the mitochondrion (a proteobacterium), and its host (an archaeon)--and carries a corollary that, over time, the majority of genes once present in the organelle genomes were relinquished to the chromosomes of the host (endosymbiotic gene transfer). However, notwithstanding eukaryote-specific gene inventions, single-gene phylogenies have never traced eukaryotic genes to three single prokaryotic sources, an issue that hinges crucially upon factors influencing phylogenetic inference. In the age of genomes, single-gene trees, once used to test the predictions of endosymbiotic theory, now spawn new theories that stand to eventually replace endosymbiotic theory with descriptive, gene tree-based variants featuring supernumerary symbionts: prokaryotic partners distinct from the cornerstone trio and whose existence is inferred solely from single-gene trees. We reason that the endosymbiotic ancestors of mitochondria and chloroplasts brought into the eukaryotic--and plant and algal--lineage a genome-sized sample of genes from the proteobacterial and cyanobacterial pangenomes of their respective day and that, even if molecular phylogeny were artifact-free, sampling prokaryotic pangenomes through endosymbiotic gene transfer would lead to inherited chimerism. Recombination in prokaryotes (transduction, conjugation, transformation) differs from recombination in eukaryotes (sex). Prokaryotic recombination leads to pangenomes, and eukaryotic recombination leads to vertical inheritance. Viewed from the perspective of endosymbiotic theory, the critical transition at the eukaryote origin that allowed escape from Muller's ratchet--the origin of eukaryotic recombination, or sex--might have required surprisingly little evolutionary innovation.

Advances in algal-prokaryotic wastewater treatment: A review of nitrogen transformations, reactor configurations and molecular tools.

PubMed

Wang, Meng; Keeley, Ryan; Zalivina, Nadezhda; Halfhide, Trina; Scott, Kathleen; Zhang, Qiong; van der Steen, Peter; Ergas, Sarina J

2018-07-01

The synergistic activity of algae and prokaryotic microorganisms can be used to improve the efficiency of biological wastewater treatment, particularly with regards to nitrogen removal. For example, algae can provide oxygen through photosynthesis needed for aerobic degradation of organic carbon and nitrification and harvested algal-prokaryotic biomass can be used to produce high value chemicals or biogas. Algal-prokaryotic consortia have been used to treat wastewater in different types of reactors, including waste stabilization ponds, high rate algal ponds and closed photobioreactors. This review addresses the current literature and identifies research gaps related to the following topics: 1) the complex interactions between algae and prokaryotes in wastewater treatment; 2) advances in bioreactor technologies that can achieve high nitrogen removal efficiencies in small reactor volumes, such as algal-prokaryotic biofilm reactors and enhanced algal-prokaryotic treatment systems (EAPS); 3) molecular tools that have expanded our understanding of the activities of algal and prokaryotic communities in wastewater treatment processes. Copyright © 2018 Elsevier Ltd. All rights reserved.

Abundance and distribution of the highly iterated palindrome 1 (HIP1) among prokaryotes

PubMed Central

Moya, Andrés

2011-01-01

We have studied the abundance and phylogenetic distribution of the Highly Iterated Palindrome 1 (HIP1) among sequenced prokaryotic genomes. We show that an overrepresentation of HIP1 is exclusive of some lineages of cyanobacteria, and that this abundance was gained only once during evolution and was subsequently lost in the lineage leading to marine pico-cyanobacteria. We show that among cyanobacterial protein sequences with annotated Pfam domains, only OpcA (glucose 6-phosphate dehydrogenase assembly protein) has a phylogenetic distribution fully matching HIP1 abundance, suggesting a functional relationship; we also show that DAM methylase (an enzyme that has the four central nucleotides of HIP1 as is site of action) is present in all cyanobacterial genomes (independently of their HIP1 content) with the exception of marine pico-cyanobacteria whom might have lost this enzyme during the process of genome reduction. Our analyses also show that in some prokaryotic lineages (particularly in those species with large genomes), HIP1 is unevenly distributed between coding and non-coding DNA (being more common in coding regions; with the exception of Cyanobacteria Yellowstone B' and Synechococcus elongates where the reverse pattern is true). Finally, we explore the hypothesis that the HIP1 can be used as a molecular “water-mark” to identify horizontally transferred genes from cyanobacteria to other species. PMID:22312590

Abundance and distribution of the highly iterated palindrome 1 (HIP1) among prokaryotes.

PubMed

Delaye, Luis; Moya, Andrés

2011-09-01

We have studied the abundance and phylogenetic distribution of the Highly Iterated Palindrome 1 (HIP1) among sequenced prokaryotic genomes. We show that an overrepresentation of HIP1 is exclusive of some lineages of cyanobacteria, and that this abundance was gained only once during evolution and was subsequently lost in the lineage leading to marine pico-cyanobacteria. We show that among cyanobacterial protein sequences with annotated Pfam domains, only OpcA (glucose 6-phosphate dehydrogenase assembly protein) has a phylogenetic distribution fully matching HIP1 abundance, suggesting a functional relationship; we also show that DAM methylase (an enzyme that has the four central nucleotides of HIP1 as is site of action) is present in all cyanobacterial genomes (independently of their HIP1 content) with the exception of marine pico-cyanobacteria whom might have lost this enzyme during the process of genome reduction. Our analyses also show that in some prokaryotic lineages (particularly in those species with large genomes), HIP1 is unevenly distributed between coding and non-coding DNA (being more common in coding regions; with the exception of Cyanobacteria Yellowstone B' and Synechococcus elongates where the reverse pattern is true). Finally, we explore the hypothesis that the HIP1 can be used as a molecular "water-mark" to identify horizontally transferred genes from cyanobacteria to other species.

Mesoscale eddies: hotspots of prokaryotic activity and differential community structure in the ocean.

PubMed

Baltar, Federico; Arístegui, Javier; Gasol, Josep M; Lekunberri, Itziar; Herndl, Gerhard J

2010-08-01

To investigate the effects of mesoscale eddies on prokaryotic assemblage structure and activity, we sampled two cyclonic eddies (CEs) and two anticyclonic eddies (AEs) in the permanent eddy-field downstream the Canary Islands. The eddy stations were compared with two far-field (FF) stations located also in the Canary Current, but outside the influence of the eddy field. The distribution of prokaryotic abundance (PA), bulk prokaryotic heterotrophic activity (PHA), various indicators of single-cell activity (such as nucleic acid content, proportion of live cells, and fraction of cells actively incorporating leucine), as well as bacterial and archaeal community structure were determined from the surface to 2000 m depth. In the upper epipelagic layer (0-200 m), the effect of eddies on the prokaryotic community was more apparent, as indicated by the higher PA, PHA, fraction of living cells, and percentage of active cells incorporating leucine within eddies than at FF stations. Prokaryotic community composition differed also between eddy and FF stations in the epipelagic layer. In the mesopelagic layer (200-1000 m), there were also significant differences in PA and PHA between eddy and FF stations, although in general, there were no clear differences in community composition or single-cell activity. The effects on prokaryotic activity and community structure were stronger in AE than CE, decreasing with depth in both types of eddies. Overall, both types of eddies show distinct community compositions (as compared with FF in the epipelagic), and represent oceanic 'hotspots' of prokaryotic activity (in the epi- and mesopelagic realms).

Flexibility and symmetry of prokaryotic genome rearrangement reveal lineage-associated core-gene-defined genome organizational frameworks.

PubMed

Kang, Yu; Gu, Chaohao; Yuan, Lina; Wang, Yue; Zhu, Yanmin; Li, Xinna; Luo, Qibin; Xiao, Jingfa; Jiang, Daquan; Qian, Minping; Ahmed Khan, Aftab; Chen, Fei; Zhang, Zhang; Yu, Jun

2014-11-25

The prokaryotic pangenome partitions genes into core and dispensable genes. The order of core genes, albeit assumed to be stable under selection in general, is frequently interrupted by horizontal gene transfer and rearrangement, but how a core-gene-defined genome maintains its stability or flexibility remains to be investigated. Based on data from 30 species, including 425 genomes from six phyla, we grouped core genes into syntenic blocks in the context of a pangenome according to their stability across multiple isolates. A subset of the core genes, often species specific and lineage associated, formed a core-gene-defined genome organizational framework (cGOF). Such cGOFs are either single segmental (one-third of the species analyzed) or multisegmental (the rest). Multisegment cGOFs were further classified into symmetric or asymmetric according to segment orientations toward the origin-terminus axis. The cGOFs in Gram-positive species are exclusively symmetric and often reversible in orientation, as opposed to those of the Gram-negative bacteria, which are all asymmetric and irreversible. Meanwhile, all species showing strong strand-biased gene distribution contain symmetric cGOFs and often specific DnaE (α subunit of DNA polymerase III) isoforms. Furthermore, functional evaluations revealed that cGOF genes are hub associated with regard to cellular activities, and the stability of cGOF provides efficient indexes for scaffold orientation as demonstrated by assembling virtual and empirical genome drafts. cGOFs show species specificity, and the symmetry of multisegmental cGOFs is conserved among taxa and constrained by DNA polymerase-centric strand-biased gene distribution. The definition of species-specific cGOFs provides powerful guidance for genome assembly and other structure-based analysis. Prokaryotic genomes are frequently interrupted by horizontal gene transfer (HGT) and rearrangement. To know whether there is a set of genes not only conserved in position

Prokaryotic Ubiquitin-Like Protein Modification

PubMed Central

Maupin-Furlow, Julie A.

2016-01-01

Prokaryotes form ubiquitin (Ub)-like isopeptide bonds on the lysine residues of proteins by at least two distinct pathways that are reversible and regulated. In mycobacteria, the C-terminal Gln of Pup (prokaryotic ubiquitin-like protein) is deamidated and isopeptide linked to proteins by a mechanism distinct from ubiquitylation in enzymology yet analogous to ubiquitylation in targeting proteins for destruction by proteasomes. Ub-fold proteins of archaea (SAMPs, small archaeal modifier proteins) and Thermus (TtuB, tRNA-two-thiouridine B) that differ from Ub in amino acid sequence, yet share a common β-grasp fold, also form isopeptide bonds by a mechanism that appears streamlined compared with ubiquitylation. SAMPs and TtuB are found to be members of a small group of Ub-fold proteins that function not only in protein modification but also in sulfur-transfer pathways associated with tRNA thiolation and molybdopterin biosynthesis. These multifunctional Ub-fold proteins are thought to be some of the most ancient of Ub-like protein modifiers. PMID:24995873

Emerging spatial patterns in Antarctic prokaryotes

PubMed Central

Chong, Chun-Wie; Pearce, David A.; Convey, Peter

2015-01-01

Recent advances in knowledge of patterns of biogeography in terrestrial eukaryotic organisms have led to a fundamental paradigm shift in understanding of the controls and history of life on land in Antarctica, and its interactions over the long term with the glaciological and geological processes that have shaped the continent. However, while it has long been recognized that the terrestrial ecosystems of Antarctica are dominated by microbes and their processes, knowledge of microbial diversity and distributions has lagged far behind that of the macroscopic eukaryote organisms. Increasing human contact with and activity in the continent is leading to risks of biological contamination and change in a region whose isolation has protected it for millions of years at least; these risks may be particularly acute for microbial communities which have, as yet, received scant recognition and attention. Even a matter apparently as straightforward as Protected Area designation in Antarctica requires robust biodiversity data which, in most parts of the continent, remain almost completely unavailable. A range of important contributing factors mean that it is now timely to reconsider the state of knowledge of Antarctic terrestrial prokaryotes. Rapid advances in molecular biological approaches are increasingly demonstrating that bacterial diversity in Antarctica may be far greater than previously thought, and that there is overlap in the environmental controls affecting both Antarctic prokaryotic and eukaryotic communities. Bacterial dispersal mechanisms and colonization patterns remain largely unaddressed, although evidence for regional evolutionary differentiation is rapidly accruing and, with this, there is increasing appreciation of patterns in regional bacterial biogeography in this large part of the globe. In this review, we set out to describe the state of knowledge of Antarctic prokaryote diversity patterns, drawing analogy with those of eukaryote groups where appropriate

Chromosomal inactivation of Bacillus subtilis exfusants: a prokaryotic model of epigenetic regulation.

PubMed

Grandjean, V; Hauck, Y; Beloin, C; Le Hégarat, F; Hirschbein, L

1998-01-01

Epigenetic mechanisms are not exclusively reserved to eukaryotic organisms. They are also observed in prokaryotes. As described first by Hotchkiss and Gabor, protoplast fusion between strains of Bacillus subtilis produces heterodiploid cells. Heterodiploidy is associated with the inactivation of one of the chromosomes. To study the physical structure of the fusion product and the molecular mechanisms of inactivation, we constructed heterodiploid clones containing two chromosomes labeled by a NotI restriction fragment length polymorphism. In the progeny, we identified haploid recombinant clones that contain a chromosome carrying large regions of inactivated DNA. Studies of both recombinants of the latter kind and heterodiploid cells indicated that chromosomal inactivation was not determined by alteration of the inactivated nucleotide sequence, but was probably due to a modification in the structure of the bacterial chromatin.

Strand-seq: a unifying tool for studies of chromosome segregation.

PubMed

Falconer, Ester; Lansdorp, Peter M

2013-01-01

Non random segregation of sister chromatids has been implicated to help specify daughter cell fate (the Silent Sister Hypothesis [1]) or to protect the genome of long-lived stem cells (the Immortal Strand Hypothesis [2]). The idea that sister chromatids are non-randomly segregated into specific daughter cells is only marginally supported by data in sporadic and often contradictory studies. As a result, the field has moved forward rather slowly. The advent of being able to directly label and differentiate sister chromatids in vivo using fluorescence in situ hybridization [3] was a significant advance for such studies. However, this approach is limited by the need for large tracks of unidirectional repeats on chromosomes and the reliance on quantitative imaging of fluorescent probes and rigorous statistical analysis to discern between the two competing hypotheses. A novel method called Strand-seq which uses next-generation sequencing to assay sister chromatid inheritance patterns independently for each chromosome [4] offers a comprehensive approach to test for non-random segregation. In addition Strand-seq enables studies on the deposition of chromatin marks in relation to DNA replication. This method is expected to help unify the field by testing previous claims of non-random segregation in an unbiased way in many model systems in vitro and in vivo. Crown Copyright © 2013. Published by Elsevier Ltd. All rights reserved.

Evolution and Diversity of the Ras Superfamily of Small GTPases in Prokaryotes

PubMed Central

Wuichet, Kristin; Søgaard-Andersen, Lotte

2015-01-01

The Ras superfamily of small GTPases are single domain nucleotide-dependent molecular switches that act as highly tuned regulators of complex signal transduction pathways. Originally identified in eukaryotes for their roles in fundamental cellular processes including proliferation, motility, polarity, nuclear transport, and vesicle transport, recent studies have revealed that single domain GTPases also control complex functions such as cell polarity, motility, predation, development and antibiotic resistance in bacteria. Here, we used a computational genomics approach to understand the abundance, diversity, and evolution of small GTPases in prokaryotes. We collected 520 small GTPase sequences present in 17% of 1,611 prokaryotic genomes analyzed that cover diverse lineages. We identified two discrete families of small GTPases in prokaryotes that show evidence of three distinct catalytic mechanisms. The MglA family includes MglA homologs, which are typically associated with the MglB GTPase activating protein, whereas members of the Rup (Ras superfamily GTPase of unknown function in prokaryotes) family are not predicted to interact with MglB homologs. System classification and genome context analyses support the involvement of small GTPases in diverse prokaryotic signal transduction pathways including two component systems, laying the foundation for future experimental characterization of these proteins. Phylogenetic analysis of prokaryotic and eukaryotic GTPases supports that the last universal common ancestor contained ancestral MglA and Rup family members. We propose that the MglA family was lost from the ancestral eukaryote and that the Ras superfamily members in extant eukaryotes are the result of vertical and horizontal gene transfer events of ancestral Rup GTPases. PMID:25480683

Links between viruses and prokaryotes throughout the water column along a North Atlantic latitudinal transect

PubMed Central

De Corte, Daniele; Sintes, Eva; Yokokawa, Taichi; Reinthaler, Thomas; Herndl, Gerhard J

2012-01-01

Viruses are an abundant, diverse and dynamic component of marine ecosystems and have a key role in the biogeochemical processes of the ocean by controlling prokaryotic and phytoplankton abundance and diversity. However, most of the studies on virus–prokaryote interactions in marine environments have been performed in nearshore waters. To assess potential variations in the relation between viruses and prokaryotes in different oceanographic provinces, we determined viral and prokaryotic abundance and production throughout the water column along a latitudinal transect in the North Atlantic. Depth-related trends in prokaryotic and viral abundance (both decreasing by one order of magnitude from epi- to abyssopelagic waters), and prokaryotic production (decreasing by three orders of magnitude) were observed along the latitudinal transect. The virus-to-prokaryote ratio (VPR) increased from ∼19 in epipelagic to ∼53 in the bathy- and abyssopelagic waters. Although the lytic viral production decreased significantly with depth, the lysogenic viral production did not vary with depth. In bathypelagic waters, pronounced differences in prokaryotic and viral abundance were found among different oceanic provinces with lower leucine incorporation rates and higher VPRs in the North Atlantic Gyre province than in the provinces further north and south. The percentage of lysogeny increased from subpolar regions toward the more oligotrophic lower latitudes. Based on the observed trends over this latitudinal transect, we conclude that the viral–host interactions significantly change among different oceanic provinces in response to changes in the biotic and abiotic variables. PMID:22258100

Geochemical Interactions and Viral-Prokaryote Relationships in Freshwater Environments

NASA Astrophysics Data System (ADS)

Kyle, J. E.; Ferris, G.

2009-05-01

Viral and prokaryotic abundances were surveyed throughout southern Ontario aquatic habitats to determine relationships with geochemical parameters in the natural environment. Surface water samples were collected from acid mine drainage in summer of 2007 and 2008 and from circum-neutral pH environments in October to November 2008. Site determination was based on collecting samples from various aquatic habitats (acid mine drainage, lakes, rivers, tributaries, wetlands) with differing bedrock geology (limestone and shale dominated vs granitic Canadian Shield) to obtain a range of geochemical conditions. At each site, measurements of temperature, pH, and Eh were conducted. Samples collected for microbial counts and electron imaging were preserved to a final concentration of 2.5 % (v/v) glutaraldehyde. Additional sample were filtered into 60 mL nalgene bottles and amber EPA certified 40 mL glass vials to determine chemical constituents and dissolved organic carbon (DOC), respectively. Water was also collected to determine additional physiochemical parameters (dissolved total iron, ferric iron, nitrate, sulfate, phosphate, alkalinity, and turbidity). All samples were stored at 4 °C until analysis. Viral and prokaryotic abundance was determined by staining samples with SYBR Green I and examining with a epifluorescence microscope under blue excitation. Multiple regression analysis using stepwise backwards regression and general linear models revealed that viral abundance was the most influential predictor of prokaryotic abundance. Additional predictors include pH, sulfate, phosphate, and magnesium. The strength of the model was very strong with 90 % of the variability explained (R2 = 0.90, p < 0.007). This is the first report, to our knowledge, of viruses exhibiting such strong controls over prokaryotic abundance in the natural environment. All relationships are positively correlated with the exception of Mg, which is negatively correlated. Iron was also noted as a

Condensin II Resolves Chromosomal Associations to Enable Anaphase I Segregation in Drosophila Male Meiosis

PubMed Central

Hartl, Tom A.; Sweeney, Sarah J.; Knepler, Peter J.; Bosco, Giovanni

2008-01-01

Several meiotic processes ensure faithful chromosome segregation to create haploid gametes. Errors to any one of these processes can lead to zygotic aneuploidy with the potential for developmental abnormalities. During prophase I of Drosophila male meiosis, each bivalent condenses and becomes sequestered into discrete chromosome territories. Here, we demonstrate that two predicted condensin II subunits, Cap-H2 and Cap-D3, are required to promote territory formation. In mutants of either subunit, territory formation fails and chromatin is dispersed throughout the nucleus. Anaphase I is also abnormal in Cap-H2 mutants as chromatin bridges are found between segregating heterologous and homologous chromosomes. Aneuploid sperm may be generated from these defects as they occur at an elevated frequency and are genotypically consistent with anaphase I segregation defects. We propose that condensin II–mediated prophase I territory formation prevents and/or resolves heterologous chromosomal associations to alleviate their potential interference in anaphase I segregation. Furthermore, condensin II–catalyzed prophase I chromosome condensation may be necessary to resolve associations between paired homologous chromosomes of each bivalent. These persistent chromosome associations likely consist of DNA entanglements, but may be more specific as anaphase I bridging was rescued by mutations in the homolog conjunction factor teflon. We propose that the consequence of condensin II mutations is a failure to resolve heterologous and homologous associations mediated by entangled DNA and/or homolog conjunction factors. Furthermore, persistence of homologous and heterologous interchromosomal associations lead to anaphase I chromatin bridging and the generation of aneuploid gametes. PMID:18927632

Recognizing short coding sequences of prokaryotic genome using a novel iteratively adaptive sparse partial least squares algorithm

PubMed Central

2013-01-01

Background Significant efforts have been made to address the problem of identifying short genes in prokaryotic genomes. However, most known methods are not effective in detecting short genes. Because of the limited information contained in short DNA sequences, it is very difficult to accurately distinguish between protein coding and non-coding sequences in prokaryotic genomes. We have developed a new Iteratively Adaptive Sparse Partial Least Squares (IASPLS) algorithm as the classifier to improve the accuracy of the identification process. Results For testing, we chose the short coding and non-coding sequences from seven prokaryotic organisms. We used seven feature sets (including GC content, Z-curve, etc.) of short genes. In comparison with GeneMarkS, Metagene, Orphelia, and Heuristic Approachs methods, our model achieved the best prediction performance in identification of short prokaryotic genes. Even when we focused on the very short length group ([60–100 nt)), our model provided sensitivity as high as 83.44% and specificity as high as 92.8%. These values are two or three times higher than three of the other methods while Metagene fails to recognize genes in this length range. The experiments also proved that the IASPLS can improve the identification accuracy in comparison with other widely used classifiers, i.e. Logistic, Random Forest (RF) and K nearest neighbors (KNN). The accuracy in using IASPLS was improved 5.90% or more in comparison with the other methods. In addition to the improvements in accuracy, IASPLS required ten times less computer time than using KNN or RF. Conclusions It is conclusive that our method is preferable for application as an automated method of short gene classification. Its linearity and easily optimized parameters make it practicable for predicting short genes of newly-sequenced or under-studied species. Reviewers This article was reviewed by Alexey Kondrashov, Rajeev Azad (nominated by Dr J.Peter Gogarten) and Yuriy Fofanov

Multiple conformational states of DnaA protein regulate its interaction with DnaA boxes in the initiation of DNA replication.

PubMed

Patel, Meera J; Bhatia, Lavesh; Yilmaz, Gulden; Biswas-Fiss, Esther E; Biswas, Subhasis B

2017-09-01

DnaA protein is the initiator of genomic DNA replication in prokaryotes. It binds to specific DNA sequences in the origin of DNA replication and unwinds small AT-rich sequences downstream for the assembly of the replisome. The mechanism of activation of DnaA that enables it to bind and organize the origin DNA and leads to replication initiation remains unclear. In this study, we have developed double-labeled fluorescent DnaA probes to analyze conformational states of DnaA protein upon binding DNA, nucleotide, and Soj sporulation protein using Fluorescence Resonance Energy Transfer (FRET). Our studies demonstrate that DnaA protein undergoes large conformational changes upon binding to substrates and there are multiple distinct conformational states that enable it to initiate DNA replication. DnaA protein adopted a relaxed conformation by expanding ~15Å upon binding ATP and DNA to form the ATP·DnaA·DNA complex. Hydrolysis of bound ATP to ADP led to a contraction of DnaA within the complex. The relaxed conformation of DnaA is likely required for the formation of the multi-protein ATP·DnaA·DNA complex. In the initiation of sporulation, Soj binding to DnaA prevented relaxation of its conformation. Soj·ADP appeared to block the activation of DnaA, suggesting a mechanism for Soj·ADP in switching initiation of DNA replication to sporulation. Our studies demonstrate that multiple conformational states of DnaA protein regulate its binding to DNA in the initiation of DNA replication. Copyright © 2017 Elsevier B.V. All rights reserved.

Partial purification and characterization of DNA topoisomerase II from Plasmodium falciparum.

PubMed

Chavalitshewinkoon, P; Leelaphiwat, S; Wilairat, P

1994-03-01

DNA topoisomerase II from Plasmodium falciparum was partially purified by FPLC using three columns: Econo-Pac Q, heparin-agarose and Mono Q. The enzyme showed ATP- and Mg2 +/- dependent activities in a decatenation assay, with optimum concentrations of 0.5 and 10 mM, respectively. Furthermore, highest activity was detected in the presence of 100 mM KCI. Enzyme decatenation activity was not inhibited by the DNA topoisomerase I inhibitor, camptothecin, but was sensitive to both prokaryotic and eukaryotic DNA topoisomerase II inhibitors.

Building a Genome Engineering Toolbox in Non-Model Prokaryotic Microbes

DOE Office of Scientific and Technical Information (OSTI.GOV)

Eckert, Carrie A; Freed, Emily; Smolinski, Sharon

The realization of a sustainable bioeconomy requires our ability to understand and engineer complex design principles for the development of platform organisms capable of efficient conversion of cheap and sustainable feedstocks (e.g. sunlight, CO2, non-food biomass) to biofuels and bioproducts at sufficient titers and costs. For model microbes such as E. coli, advances in DNA reading and writing technologies are driving adoption of new paradigms for engineering biological systems. Unfortunately, microbes with properties of interest for the utilization of cheap and renewable feedstocks such as photosynthesis, autotrophic growth, and cellulose degradation have very few, if any, genetic tools for metabolicmore » engineering. Therefore, it is important to begin to develop 'design rules' for building a genetic toolbox for novel microbes. Here, we present an overview of our current understanding of these rules for the genetic manipulation of prokaryotic microbes and available genetic tools to expand our ability to genetically engineer non-model systems.« less

Known knowns, known unknowns and unknown unknowns in prokaryotic transposition.

PubMed

Siguier, Patricia; Gourbeyre, Edith; Chandler, Michael

2017-08-01

Although the phenomenon of transposition has been known for over 60 years, its overarching importance in modifying and streamlining genomes took some time to recognize. In spite of a robust understanding of transposition of some TE, there remain a number of important TE groups with potential high genome impact and unknown transposition mechanisms and yet others, only recently identified by bioinformatics, yet to be formally confirmed as mobile. Here, we point to some areas of limited understanding concerning well established important TE groups with DDE Tpases, to address central gaps in our knowledge of characterised Tn with other types of Tpases and finally, to highlight new potentially mobile DNA species. It is not exhaustive. Examples have been chosen to provide encouragement in the continued exploration of the considerable prokaryotic mobilome especially in light of the current threat to public health posed by the spread of multiple Ab R . Copyright © 2017 Elsevier Ltd. All rights reserved.

PSSRdb: a relational database of polymorphic simple sequence repeats extracted from prokaryotic genomes.

PubMed

Kumar, Pankaj; Chaitanya, Pasumarthy S; Nagarajaram, Hampapathalu A

2011-01-01

PSSRdb (Polymorphic Simple Sequence Repeats database) (http://www.cdfd.org.in/PSSRdb/) is a relational database of polymorphic simple sequence repeats (PSSRs) extracted from 85 different species of prokaryotes. Simple sequence repeats (SSRs) are the tandem repeats of nucleotide motifs of the sizes 1-6 bp and are highly polymorphic. SSR mutations in and around coding regions affect transcription and translation of genes. Such changes underpin phase variations and antigenic variations seen in some bacteria. Although SSR-mediated phase variation and antigenic variations have been well-studied in some bacteria there seems a lot of other species of prokaryotes yet to be investigated for SSR mediated adaptive and other evolutionary advantages. As a part of our on-going studies on SSR polymorphism in prokaryotes we compared the genome sequences of various strains and isolates available for 85 different species of prokaryotes and extracted a number of SSRs showing length variations and created a relational database called PSSRdb. This database gives useful information such as location of PSSRs in genomes, length variation across genomes, the regions harboring PSSRs, etc. The information provided in this database is very useful for further research and analysis of SSRs in prokaryotes.

Prokaryotic Contig Annotation Pipeline Server: Web Application for a Prokaryotic Genome Annotation Pipeline Based on the Shiny App Package.

PubMed

Park, Byeonghyeok; Baek, Min-Jeong; Min, Byoungnam; Choi, In-Geol

2017-09-01

Genome annotation is a primary step in genomic research. To establish a light and portable prokaryotic genome annotation pipeline for use in individual laboratories, we developed a Shiny app package designated as "P-CAPS" (Prokaryotic Contig Annotation Pipeline Server). The package is composed of R and Python scripts that integrate publicly available annotation programs into a server application. P-CAPS is not only a browser-based interactive application but also a distributable Shiny app package that can be installed on any personal computer. The final annotation is provided in various standard formats and is summarized in an R markdown document. Annotation can be visualized and examined with a public genome browser. A benchmark test showed that the annotation quality and completeness of P-CAPS were reliable and compatible with those of currently available public pipelines.

Intrinsically bent DNA in replication origins and gene promoters.

PubMed

Gimenes, F; Takeda, K I; Fiorini, A; Gouveia, F S; Fernandez, M A

2008-06-24

Intrinsically bent DNA is an alternative conformation of the DNA molecule caused by the presence of dA/dT tracts, 2 to 6 bp long, in a helical turn phase DNA or with multiple intervals of 10 to 11 bp. Other than flexibility, intrinsic bending sites induce DNA curvature in particular chromosome regions such as replication origins and promoters. Intrinsically bent DNA sites are important in initiating DNA replication, and are sometimes found near to regions associated with the nuclear matrix. Many methods have been developed to localize bent sites, for example, circular permutation, computational analysis, and atomic force microscopy. This review discusses intrinsically bent DNA sites associated with replication origins and gene promoter regions in prokaryote and eukaryote cells. We also describe methods for identifying bent DNA sites for circular permutation and computational analysis.

Prokaryotic cells: structural organisation of the cytoskeleton and organelles.

PubMed

Souza, Wanderley de

2012-05-01

For many years, prokaryotic cells were distinguished from eukaryotic cells based on the simplicity of their cytoplasm, in which the presence of organelles and cytoskeletal structures had not been discovered. Based on current knowledge, this review describes the complex components of the prokaryotic cell cytoskeleton, including (i) tubulin homologues composed of FtsZ, BtuA, BtuB and several associated proteins, which play a fundamental role in cell division, (ii) actin-like homologues, such as MreB and Mb1, which are involved in controlling cell width and cell length, and (iii) intermediate filament homologues, including crescentin and CfpA, which localise on the concave side of a bacterium and along its inner curvature and associate with its membrane. Some prokaryotes exhibit specialised membrane-bound organelles in the cytoplasm, such as magnetosomes and acidocalcisomes, as well as protein complexes, such as carboxysomes. This review also examines recent data on the presence of nanotubes, which are structures that are well characterised in mammalian cells that allow direct contact and communication between cells.

An unsupervised classification scheme for improving predictions of prokaryotic TIS.

PubMed

Tech, Maike; Meinicke, Peter

2006-03-09

Although it is not difficult for state-of-the-art gene finders to identify coding regions in prokaryotic genomes, exact prediction of the corresponding translation initiation sites (TIS) is still a challenging problem. Recently a number of post-processing tools have been proposed for improving the annotation of prokaryotic TIS. However, inherent difficulties of these approaches arise from the considerable variation of TIS characteristics across different species. Therefore prior assumptions about the properties of prokaryotic gene starts may cause suboptimal predictions for newly sequenced genomes with TIS signals differing from those of well-investigated genomes. We introduce a clustering algorithm for completely unsupervised scoring of potential TIS, based on positionally smoothed probability matrices. The algorithm requires an initial gene prediction and the genomic sequence of the organism to perform the reannotation. As compared with other methods for improving predictions of gene starts in bacterial genomes, our approach is not based on any specific assumptions about prokaryotic TIS. Despite the generality of the underlying algorithm, the prediction rate of our method is competitive on experimentally verified test data from E. coli and B. subtilis. Regarding genomes with high G+C content, in contrast to some previously proposed methods, our algorithm also provides good performance on P. aeruginosa, B. pseudomallei and R. solanacearum. On reliable test data we showed that our method provides good results in post-processing the predictions of the widely-used program GLIMMER. The underlying clustering algorithm is robust with respect to variations in the initial TIS annotation and does not require specific assumptions about prokaryotic gene starts. These features are particularly useful on genomes with high G+C content. The algorithm has been implemented in the tool "TICO" (TIs COrrector) which is publicly available from our web site.

Gene duplications in prokaryotes can be associated with environmental adaptation.

PubMed

Bratlie, Marit S; Johansen, Jostein; Sherman, Brad T; Huang, Da Wei; Lempicki, Richard A; Drabløs, Finn

2010-10-20

Gene duplication is a normal evolutionary process. If there is no selective advantage in keeping the duplicated gene, it is usually reduced to a pseudogene and disappears from the genome. However, some paralogs are retained. These gene products are likely to be beneficial to the organism, e.g. in adaptation to new environmental conditions. The aim of our analysis is to investigate the properties of paralog-forming genes in prokaryotes, and to analyse the role of these retained paralogs by relating gene properties to life style of the corresponding prokaryotes. Paralogs were identified in a number of prokaryotes, and these paralogs were compared to singletons of persistent orthologs based on functional classification. This showed that the paralogs were associated with for example energy production, cell motility, ion transport, and defence mechanisms. A statistical overrepresentation analysis of gene and protein annotations was based on paralogs of the 200 prokaryotes with the highest fraction of paralog-forming genes. Biclustering of overrepresented gene ontology terms versus species was used to identify clusters of properties associated with clusters of species. The clusters were classified using similarity scores on properties and species to identify interesting clusters, and a subset of clusters were analysed by comparison to literature data. This analysis showed that paralogs often are associated with properties that are important for survival and proliferation of the specific organisms. This includes processes like ion transport, locomotion, chemotaxis and photosynthesis. However, the analysis also showed that the gene ontology terms sometimes were too general, imprecise or even misleading for automatic analysis. Properties described by gene ontology terms identified in the overrepresentation analysis are often consistent with individual prokaryote lifestyles and are likely to give a competitive advantage to the organism. Paralogs and singletons dominate

Impact of pyrene and cadmium co-contamination on prokaryotic community in coastal sediment microcosms.

PubMed

Ding, Qifang; Huang, Xiaolin; Hu, Hanjing; Hong, Man; Zhang, Demin; Wang, Kai

2017-12-01

Acute ecological impacts of co-contamination of polycyclic aromatic hydrocarbons (PAHs) and heavy metals on diversity and composition of coastal benthic prokaryotes were unclear. We took pyrene (Pyr) and cadmium (Cd) as the representatives and mimicked an eight-week exposure of moderate and high levels of Pyr, Cd and their mixtures. 16S rRNA amplicon sequencing was used to investigate interaction of the contaminants in temporal succession of prokaryotes. Generally, concentrations of Pyr and HCl-extractable Cd in the sediments were stable over time. Effects and interaction of Pyr and Cd on prokaryotic α-diversity were temporally- and dose-dependent with a decreasing trend in richness and Shannon index under various contamination regimes, particularly in the single-Cd contaminated groups at the early stage. Temporal variability and Pyr-induced pattern in prokaryotic composition were observed. However, Pyr and Cd showed a persistent interaction in prokaryotic composition after 7 days, altering successional trajectories of communities. The communities under Pyr contamination regardless of Cd could be at a developing stage for an active PAH-degrading community with appearance of a pioneer Cycloclasticus phylotype, persistently showing a strong correlation with Pyr level. The associations of phylotypes and Cd level were short-lived and weak, corresponding to the overall resistance of prokaryotic composition to Cd. In the high-throughput sequencing era, using microcosm experiment, we renewed the knowledge about how prokaryotes vary in terms of α-diversity, composition and specific taxa in response to co-contamination of model contaminants at a temporal scale. Copyright © 2017 Elsevier Ltd. All rights reserved.

Segregation and Poverty Concentration: The Role of Three Segregations

PubMed Central

Quillian, Lincoln

2014-01-01

A key argument of Massey and Denton’s American Apartheid (1993) is that racial residential segregation and non-white group poverty rates combine interactively to produce spatially concentrated poverty. Despite a compelling theoretical rationale, the empirical tests of this proposition have been negative or mixed. This paper develops a formal decomposition model that expands the Massey model of how segregation, group poverty rates, and other spatial conditions combine to form concentrated poverty. The revised decomposition model allows for income effects on cross-race neighborhood residence and interactive combinations of multiple spatial conditions in the formation of concentrated poverty. Applying the model to data reveals that racial segregation and income segregation within race contribute importantly to poverty concentration, as Massey argued, but that almost equally important for poverty concentration is the disproportionate poverty of the non-group neighbors of blacks and Hispanics. The missing interaction Massey expected in empirical tests can be found with proper accounting for the factors in the expanded model. “Because of racial segregation, a significant share of black America is condemned to experience a social environment where poverty and joblessness are the norm, where a majority of children are born out of wedlock, where most families are on welfare, where educational failure prevails, and where social and physical deterioration abound. Through prolonged exposure to such an environment, black chances for social and economic success are drastically reduced.”--Douglas Massey and Nancy Denton, American Apartheid, p. 2 PMID:24648570

Metabolic profiles of prokaryotic and eukaryotic communities in deep-sea sponge Lamellomorpha sp. indicated by metagenomics

NASA Astrophysics Data System (ADS)

Li, Zhi-Yong; Wang, Yue-Zhu; He, Li-Ming; Zheng, Hua-Jun

2014-01-01

The whole metabolism of a sponge holobiont and the respective contributions of prokaryotic and eukaryotic symbionts and their associations with the sponge host remain largely unclear. Meanwhile, compared with shallow water sponges, deep-sea sponges are rarely understood. Here we report the metagenomic exploration of deep-sea sponge Lamellomorpha sp. at the whole community level. Metagenomic data showed phylogenetically diverse prokaryotes and eukaryotes in Lamellomorpha sp.. MEGAN and gene enrichment analyses indicated different metabolic potentials of prokaryotic symbionts from eukaryotic symbionts, especially in nitrogen and carbon metabolisms, and their molecular interactions with the sponge host. These results supported the hypothesis that prokaryotic and eukaryotic symbionts have different ecological roles and relationships with sponge host. Moreover, vigorous denitrification, and CO2 fixation by chemoautotrophic prokaryotes were suggested for this deep-sea sponge. The study provided novel insights into the respective potentials of prokaryotic and eukaryotic symbionts and their associations with deep-sea sponge Lamellomorpha sp..

Metabolic profiles of prokaryotic and eukaryotic communities in deep-sea sponge Neamphius huxleyi indicated by metagenomics

PubMed Central

Li, Zhi-Yong; Wang, Yue-Zhu; He, Li-Ming; Zheng, Hua-Jun

2014-01-01

The whole metabolism of a sponge holobiont and the respective contributions of prokaryotic and eukaryotic symbionts and their associations with the sponge host remain largely unclear. Meanwhile, compared with shallow water sponges, deep-sea sponges are rarely understood. Here we report the metagenomic exploration of deep-sea sponge Neamphius huxleyi at the whole community level. Metagenomic data showed phylogenetically diverse prokaryotes and eukaryotes in Neamphius huxleyi. MEGAN and gene enrichment analyses indicated different metabolic potentials of prokaryotic symbionts from eukaryotic symbionts, especially in nitrogen and carbon metabolisms, and their molecular interactions with the sponge host. These results supported the hypothesis that prokaryotic and eukaryotic symbionts have different ecological roles and relationships with sponge host. Moreover, vigorous denitrification, and CO2 fixation by chemoautotrophic prokaryotes were suggested for this deep-sea sponge. The study provided novel insights into the respective potentials of prokaryotic and eukaryotic symbionts and their associations with deep-sea sponge Neamphius huxleyi. PMID:24463735

DNA Base Flipping: A General Mechanism for Writing, Reading, and Erasing DNA Modifications

PubMed Central

Cheng, Xiaodong

2017-01-01

The modification of DNA bases is a classic hallmark of epigenetics. Four forms of modified cytosine—5-methylcytosine, 5-hydroxymethylcytosine, 5-formylcytosine, and 5-carboxylcytosine—have been discovered in eukaryotic DNA. In addition to cytosine carbon-5 modifications, cytosine and adenine methylated in the exocyclic amine—N4-methylcytosine and N6-methyladenine—are other modified DNA bases discovered even earlier. Each modified base can be considered a distinct epigenetic signal with broader biological implications beyond simple chemical changes. Since 1994, crystal structures of proteins and enzymes involved in writing, reading, and erasing modified bases have become available. Here, we present a structural synopsis of writers, readers, and erasers of the modified bases from prokaryotes and eukaryotes. Despite significant differences in structures and functions, they are remarkably similar regarding their engagement in flipping a target base/nucleotide within DNA for specific recognitions and/or reactions. We thus highlight base flipping as a common structural framework broadly applied by distinct classes of proteins and enzymes across phyla for epigenetic regulations of DNA. PMID:27826845

Microscopic Identification of Prokaryotes in Modern and Ancient Halite, Saline Valley and Death Valley, California

NASA Astrophysics Data System (ADS)

Schubert, Brian A.; Lowenstein, Tim K.; Timofeeff, Michael N.

2009-06-01

Primary fluid inclusions in halite crystallized in Saline Valley, California, in 1980, 2004-2005, and 2007, contain rod- and coccoid-shaped microparticles the same size and morphology as archaea and bacteria living in modern brines. Primary fluid inclusions from a well-dated (0-100,000 years), 90 m long salt core from Badwater Basin, Death Valley, California, also contain microparticles, here interpreted as halophilic and halotolerant prokaryotes. Prokaryotes are distinguished from crystals on the basis of morphology, optical properties (birefringence), and uniformity of size. Electron micrographs of microparticles from filtered modern brine (Saline Valley), dissolved modern halite crystals (Saline Valley), and dissolved ancient halite crystals (Death Valley) support in situ microscopic observations that prokaryotes are present in fluid inclusions in ancient halite. In the Death Valley salt core, prokaryotes in fluid inclusions occur almost exclusively in halite precipitated in perennial saline lakes 10,000 to 35,000 years ago. This suggests that trapping and preservation of prokaryotes in fluid inclusions is influenced by the surface environment in which the halite originally precipitated. In all cases, prokaryotes in fluid inclusions in halite from the Death Valley salt core are miniaturized (<1 μm diameter cocci, <2.5 μm long, very rare rod shapes), which supports interpretations that the prokaryotes are indigenous to the halite and starvation survival may be the normal response of some prokaryotes to entrapment in fluid inclusions for millennia. These results reinforce the view that fluid inclusions in halite and possibly other evaporites are important repositories of microbial life and should be carefully examined in the search for ancient microorganisms on Earth, Mars, and elsewhere in the Solar System.

Modeling the winter-to-summer transition of prokaryotic and viral abundance in the Arctic Ocean.

PubMed

Winter, Christian; Payet, Jérôme P; Suttle, Curtis A

2012-01-01

One of the challenges in oceanography is to understand the influence of environmental factors on the abundances of prokaryotes and viruses. Generally, conventional statistical methods resolve trends well, but more complex relationships are difficult to explore. In such cases, Artificial Neural Networks (ANNs) offer an alternative way for data analysis. Here, we developed ANN-based models of prokaryotic and viral abundances in the Arctic Ocean. The models were used to identify the best predictors for prokaryotic and viral abundances including cytometrically-distinguishable populations of prokaryotes (high and low nucleic acid cells) and viruses (high- and low-fluorescent viruses) among salinity, temperature, depth, day length, and the concentration of Chlorophyll-a. The best performing ANNs to model the abundances of high and low nucleic acid cells used temperature and Chl-a as input parameters, while the abundances of high- and low-fluorescent viruses used depth, Chl-a, and day length as input parameters. Decreasing viral abundance with increasing depth and decreasing system productivity was captured well by the ANNs. Despite identifying the same predictors for the two populations of prokaryotes and viruses, respectively, the structure of the best performing ANNs differed between high and low nucleic acid cells and between high- and low-fluorescent viruses. Also, the two prokaryotic and viral groups responded differently to changes in the predictor parameters; hence, the cytometric distinction between these populations is ecologically relevant. The models imply that temperature is the main factor explaining most of the variation in the abundances of high nucleic acid cells and total prokaryotes and that the mechanisms governing the reaction to changes in the environment are distinctly different among the prokaryotic and viral populations.

Modeling the Winter–to–Summer Transition of Prokaryotic and Viral Abundance in the Arctic Ocean

PubMed Central

Winter, Christian; Payet, Jérôme P.; Suttle, Curtis A.

2012-01-01

One of the challenges in oceanography is to understand the influence of environmental factors on the abundances of prokaryotes and viruses. Generally, conventional statistical methods resolve trends well, but more complex relationships are difficult to explore. In such cases, Artificial Neural Networks (ANNs) offer an alternative way for data analysis. Here, we developed ANN-based models of prokaryotic and viral abundances in the Arctic Ocean. The models were used to identify the best predictors for prokaryotic and viral abundances including cytometrically-distinguishable populations of prokaryotes (high and low nucleic acid cells) and viruses (high- and low-fluorescent viruses) among salinity, temperature, depth, day length, and the concentration of Chlorophyll-a. The best performing ANNs to model the abundances of high and low nucleic acid cells used temperature and Chl-a as input parameters, while the abundances of high- and low-fluorescent viruses used depth, Chl-a, and day length as input parameters. Decreasing viral abundance with increasing depth and decreasing system productivity was captured well by the ANNs. Despite identifying the same predictors for the two populations of prokaryotes and viruses, respectively, the structure of the best performing ANNs differed between high and low nucleic acid cells and between high- and low-fluorescent viruses. Also, the two prokaryotic and viral groups responded differently to changes in the predictor parameters; hence, the cytometric distinction between these populations is ecologically relevant. The models imply that temperature is the main factor explaining most of the variation in the abundances of high nucleic acid cells and total prokaryotes and that the mechanisms governing the reaction to changes in the environment are distinctly different among the prokaryotic and viral populations. PMID:23285186

[Prokaryote diversity in water environment of land-ocean ecotone of Zhuhai City].

PubMed

Huang, Xiao-Lan; Chen, Jian-Yao; Zhou, Shi-Ning; Xie, Li-Chun; Fu, Cong-Sheng

2010-02-01

By constructing 16S rDNA clone library with PCR-RFLP, the prokaryote diversity in the seawater and groundwater of land-ocean ecotone of Zhuhai City was investigated, and the similarity and cluster analyses were implemented with the database of the sequences in Genbank. In the seawater, Proteobacteria was dominant, followed by Archaeon, Gemmatimonadetes, Candidate division OP3 and OP8, and Planctomycetes, etc.; while in the groundwater, Archaeon was dominant, followed by Proteobacteria, Sphingobacteria, Candidate division OP3, Actinobacterium, and Pseudomonas. The dominant taxa in the groundwater had high similarity to the unculturable groups of marine microorganisms. Large amount of bacteria capable of degrading organic matter and purifying water body existed in the groundwater, suggesting that after long-term evolution, the land-ocean ecotone of Zhuhai City had the characteristics of both land and ocean.

Metabolic Design and Control for Production in Prokaryotes

DOE Office of Scientific and Technical Information (OSTI.GOV)

Chhabra, Swapnil R.; Keasling, J.D.

2010-11-10

Prokaryotic life on earth is manifested by its diversity and omnipresence. These microbes serve as natural sources of a large variety of compounds with the potential to serve the ever growing, medicinal, chemical and transportation needs of the human population. However, commercially viable production of these compounds can be realized only through significant improvement of the native production capacity of natural isolates. The most favorable way to achieve this goal is through the genetic manipulation of metabolic pathways that direct the production of these molecules. While random mutagenesis and screening have dominated the industrial production of such compounds in themore » past our increased understanding of microbial physiology over the last five decades has shifted this trend towards rational approaches for metabolic design. Major drivers of this trend include recombinant DNA technology, high throughput characterization of macromolecular cellular components, quantitative modeling for metabolic engine ring, targeted combinatorial engineering and synthetic biology. In this chapter we track the evolution of microbial engineering technologies from the black box era of random mutagenesis to the science and engineering-driven era of metabolic design.« less

Application of subtracted gDNA microarray-assisted Bulked Segregant Analysis for rapid discovery of molecular markers associated with day-neutrality in strawberry (Fragaria x ananassa)

PubMed Central

Gor, Mian Chee; Mantri, Nitin; Pang, Edwin

2016-01-01

A Fragaria Discovery Panel (FDP; strawberry-specific SDA) containing 287 features was constructed by subtracting the pooled gDNA of nine non-angiosperm species from the pooled gDNA of five strawberry genotypes. This FDP was used for Bulk Segregant Analysis (BSA) to enable identification of molecular markers associated with day-neutrality. Analysis of hybridisation patterns of a short day (SD) DNA bulk and three day-neutral (DN) DNA bulks varying in flowering strength allowed identification of a novel feature, FaP2E11, closely linked to CYTOKININ OXIDASE 1 (CKX1) gene possibly involved in promoting flowering under non-inductive condition. The signal intensities of FaP2E11 feature obtained from the strong DN bulk (DN1) is three fold higher than the short day bulk (SD), indicating that the putative marker may linked to a CKX1 variant allele with lower enzyme activity. We propose a model for flowering regulation based on the hypothesis that flowering strength may be regulated by the copy number of FaP2E11-linked CKX1 alleles. This study demonstrates the feasibility of the SDA-based BSA approach for the identification of molecular markers associated with day-neutrality in strawberry. This innovative strategy is an efficient and cost-effective approach for molecular marker discovery. PMID:27586242

Mitochondrial DNA sequence characteristics modulate the size of the genetic bottleneck.

PubMed

Wilson, Ian J; Carling, Phillipa J; Alston, Charlotte L; Floros, Vasileios I; Pyle, Angela; Hudson, Gavin; Sallevelt, Suzanne C E H; Lamperti, Costanza; Carelli, Valerio; Bindoff, Laurence A; Samuels, David C; Wonnapinij, Passorn; Zeviani, Massimo; Taylor, Robert W; Smeets, Hubert J M; Horvath, Rita; Chinnery, Patrick F

2016-03-01

With a combined carrier frequency of 1:200, heteroplasmic mitochondrial DNA (mtDNA) mutations cause human disease in ∼1:5000 of the population. Rapid shifts in the level of heteroplasmy seen within a single generation contribute to the wide range in the severity of clinical phenotypes seen in families transmitting mtDNA disease, consistent with a genetic bottleneck during transmission. Although preliminary evidence from human pedigrees points towards a random drift process underlying the shifting heteroplasmy, some reports describe differences in segregation pattern between different mtDNA mutations. However, based on limited observations and with no direct comparisons, it is not clear whether these observations simply reflect pedigree ascertainment and publication bias. To address this issue, we studied 577 mother-child pairs transmitting the m.11778G>A, m.3460G>A, m.8344A>G, m.8993T>G/C and m.3243A>G mtDNA mutations. Our analysis controlled for inter-assay differences, inter-laboratory variation and ascertainment bias. We found no evidence of selection during transmission but show that different mtDNA mutations segregate at different rates in human pedigrees. m.8993T>G/C segregated significantly faster than m.11778G>A, m.8344A>G and m.3243A>G, consistent with a tighter mtDNA genetic bottleneck in m.8993T>G/C pedigrees. Our observations support the existence of different genetic bottlenecks primarily determined by the underlying mtDNA mutation, explaining the different inheritance patterns observed in human pedigrees transmitting pathogenic mtDNA mutations. © The Author 2016. Published by Oxford University Press.

"Prokaryotic Pathway" Is Not Prokaryotic: Noncyanobacterial Origin of the Chloroplast Lipid Biosynthetic Pathway Revealed by Comprehensive Phylogenomic Analysis.

PubMed

Sato, Naoki; Awai, Koichiro

2017-11-01

Lipid biosynthesis within the chloroplast, or more generally plastids, was conventionally called "prokaryotic pathway," which produces glycerolipids bearing C18 acids at the sn-1 position and C16 acids at the sn-2 position, as in cyanobacteria such as Anabaena and Synechocystis. This positional specificity is determined during the synthesis of phosphatidate, which is a precursor to diacylglycerol, the acceptor of galactose for the synthesis of galactolipids. The first acylation at sn-1 is catalyzed by glycerol-3-phosphate acyltransferase (GPAT or GPT), whereas the second acylation at sn-2 is performed by lysophosphatidate acyltransferase (LPAAT, AGPAT, or PlsC). Here we present comprehensive phylogenomic analysis of the origins of various acyltransferases involved in the synthesis of phosphatidate, as well as phosphatidate phosphatases in the chloroplasts. The results showed that the enzymes involved in the two steps of acylation in cyanobacteria and chloroplasts are entirely phylogenetically unrelated despite a previous report stating that the chloroplast LPAAT (ATS2) and cyanobacterial PlsC were sister groups. Phosphatidate phosphatases were separated into eukaryotic and prokaryotic clades, and the chloroplast enzymes were not of cyanobacterial origin, in contrast with another previous report. These results indicate that the lipid biosynthetic pathway in the chloroplasts or plastids did not originate from the cyanobacterial endosymbiont and is not "prokaryotic" in the context of endosymbiotic theory of plastid origin. This is another line of evidence for the discontinuity of plastids and cyanobacteria, which has been suggested in the glycolipid biosynthesis. © The Author 2017. Published by Oxford University Press on behalf of the Society for Molecular Biology and Evolution.

Molecular fossils of prokaryotes in ancient authigenic minerals: archives of microbial activity in reefs and mounds?

NASA Astrophysics Data System (ADS)

Heindel, Katrin; Birgel, Daniel; Richoz, Sylvain; Westphal, Hildegard; Peckmann, Jörn

2016-04-01

Molecular fossils (lipid biomarkers) are commonly used as proxies in organic-rich sediments of various sources, including eukaryotes and prokaryotes. Usually, molecular fossils of organisms transferred from the water column to the sediment are studied to monitor environmental changes (e.g., temperature, pH). Apart from these 'allochthonous' molecular fossils, prokaryotes are active in sediments and mats on the seafloor and leave behind 'autochthonous' molecular fossils in situ. In contrast to many phototrophic organisms, most benthic sedimentary prokaryotes are obtaining their energy from oxidation or reduction of organic or inorganic substrates. A peculiarity of some of the sediment-thriving prokaryotes is their ability to trigger in situ mineral precipitation, often but not only due to metabolic activity, resulting in authigenic rocks (microbialites). During that process, prokaryotes are rapidly entombed in the mineral matrix, where the molecular fossils are protected from early (bio)degradation. In contrast to other organic compounds (DNA, proteins etc.), molecular fossils can be preserved over very long time periods (millions of years). Thus, molecular fossils in authigenic mineral phases are perfectly suitable to trace microbial activity back in time. Among the best examples of molecular fossils, which are preserved in authigenic rocks are various microbialites, forming e.g. in phototrophic microbial mats and at cold seeps. Microbialite formation is reported throughout earth history. We here will focus on reefal microbialites form the Early Triassic and the Holocene. After the End-Permian mass extinction, microbialites covered wide areas on the ocean margins. In microbialites from the Griesbachian in Iran and Turkey (both Neotethys), molecular fossils of cyanobacteria, archaea, anoxygenic phototrophs, and sulphate-reducing bacteria indicate the presence of layered microbial mats on the seafloor, in which carbonate precipitation was induced. In association with

The TubR-centromere complex adopts a double-ring segrosome structure in Type III partition systems.

PubMed

Martín-García, Bárbara; Martín-González, Alejandro; Carrasco, Carolina; Hernández-Arriaga, Ana M; Ruíz-Quero, Rubén; Díaz-Orejas, Ramón; Aicart-Ramos, Clara; Moreno-Herrero, Fernando; Oliva, María A

2018-05-14

In prokaryotes, the centromere is a specialized segment of DNA that promotes the assembly of the segrosome upon binding of the Centromere Binding Protein (CBP). The segrosome structure exposes a specific surface for the interaction of the CBP with the motor protein that mediates DNA movement during cell division. Additionally, the CBP usually controls the transcriptional regulation of the segregation system as a cell cycle checkpoint. Correct segrosome functioning is therefore indispensable for accurate DNA segregation. Here, we combine biochemical reconstruction and structural and biophysical analysis to bring light to the architecture of the segrosome complex in Type III partition systems. We present the particular features of the centromere site, tubC, of the model system encoded in Clostridium botulinum prophage c-st. We find that the split centromere site contains two different iterons involved in the binding and spreading of the CBP, TubR. The resulting nucleoprotein complex consists of a novel double-ring structure that covers part of the predicted promoter. Single molecule data provides a mechanism for the formation of the segrosome structure based on DNA bending and unwinding upon TubR binding.

DMINDA: an integrated web server for DNA motif identification and analyses

PubMed Central

Ma, Qin; Zhang, Hanyuan; Mao, Xizeng; Zhou, Chuan; Liu, Bingqiang; Chen, Xin; Xu, Ying

2014-01-01

DMINDA (DNA motif identification and analyses) is an integrated web server for DNA motif identification and analyses, which is accessible at http://csbl.bmb.uga.edu/DMINDA/. This web site is freely available to all users and there is no login requirement. This server provides a suite of cis-regulatory motif analysis functions on DNA sequences, which are important to elucidation of the mechanisms of transcriptional regulation: (i) de novo motif finding for a given set of promoter sequences along with statistical scores for the predicted motifs derived based on information extracted from a control set, (ii) scanning motif instances of a query motif in provided genomic sequences, (iii) motif comparison and clustering of identified motifs, and (iv) co-occurrence analyses of query motifs in given promoter sequences. The server is powered by a backend computer cluster with over 150 computing nodes, and is particularly useful for motif prediction and analyses in prokaryotic genomes. We believe that DMINDA, as a new and comprehensive web server for cis-regulatory motif finding and analyses, will benefit the genomic research community in general and prokaryotic genome researchers in particular. PMID:24753419

Light-dependent segregation of begomoviruses in Asystasia gangetica leaves.

PubMed

Wyant, Patricia; Strohmeier, Stephan; Fischer, Alexander; Schäfer, Benjamin; Briddon, Rob W; Krenz, Björn; Jeske, Holger

2015-01-02

Asystasia gangetica (Acanthaceae) from tropical Africa and Asia is used as source of food and for medical applications. Plants collected in West Africa in the 1980s with typical geminivirus symptoms showed an unusual symptom segregation that included vein yellowing, curling and mosaic, which were present simultaneously or separately on different leaves of the same plant or on different plants propagated as cuttings from a single plant. Rolling-circle amplification in combination with restriction fragment length polymorphism analysis followed by deep sequencing of the RCA products identified two geminiviruses in these plants. One with a bipartite genome, Asystasia begomovirus 1, and the other with a monopartite genome together with its defective DNA, Asystasia begomovirus 2. The relationship between leaf symptoms and virus distribution under different light regimes was investigated, and showed for the first time an unusual segregation of symptoms and viruses, either within a single plant, or even within a leaf. Copyright © 2014 Elsevier B.V. All rights reserved.

Metabolic Activity and Functional Diversity Changes in Sediment Prokaryotic Communities Organically Enriched with Mussel Biodeposits

PubMed Central

Pollet, Thomas; Cloutier, Olivier; Nozais, Christian; McKindsey, Christopher W.; Archambault, Philippe

2015-01-01

This experimental microcosm study reports the influence of organic enrichments by mussel biodeposits on the metabolic activity and functional diversity of benthic prokaryotic communities. The different biodeposit enrichment regimes created, which mimicked the quantity of faeces and pseudo-faeces potentially deposited below mussel farms, show a clear stimulatory effect of this organic enrichment on prokaryotic metabolic activity. This effect was detected once a certain level of biodeposition was attained with a tipping point estimated between 3.25 and 10 g day-1 m-2. Prokaryotic communities recovered their initial metabolic activity by 11 days after the cessation of biodeposit additions. However, their functional diversity remained greater than prior to the disturbance suggesting that mussel biodeposit enrichment may disturb the functioning and perhaps the role of prokaryotic communities in benthic ecosystems. This manipulative approach provided new information on the influence of mussel biodeposition on benthic prokaryotic communities and dose-response relationships and may support the development of carrying capacity models for bivalve culture. PMID:25923715

Metabolic activity and functional diversity changes in sediment prokaryotic communities organically enriched with mussel biodeposits.

PubMed

Pollet, Thomas; Cloutier, Olivier; Nozais, Christian; McKindsey, Christopher W; Archambault, Philippe

2015-01-01

This experimental microcosm study reports the influence of organic enrichments by mussel biodeposits on the metabolic activity and functional diversity of benthic prokaryotic communities. The different biodeposit enrichment regimes created, which mimicked the quantity of faeces and pseudo-faeces potentially deposited below mussel farms, show a clear stimulatory effect of this organic enrichment on prokaryotic metabolic activity. This effect was detected once a certain level of biodeposition was attained with a tipping point estimated between 3.25 and 10 g day-1 m-2. Prokaryotic communities recovered their initial metabolic activity by 11 days after the cessation of biodeposit additions. However, their functional diversity remained greater than prior to the disturbance suggesting that mussel biodeposit enrichment may disturb the functioning and perhaps the role of prokaryotic communities in benthic ecosystems. This manipulative approach provided new information on the influence of mussel biodeposition on benthic prokaryotic communities and dose-response relationships and may support the development of carrying capacity models for bivalve culture.

Do prokaryotes contain microtubules?

NASA Technical Reports Server (NTRS)

Bermudes, D.; Hinkle, G.; Margulis, L.

1994-01-01

In eukaryotic cells, microtubules are 24-nm-diameter tubular structures composed of a class of conserved proteins called tubulin. They are involved in numerous cell functions including ciliary motility, nerve cell elongation, pigment migration, centrosome formation, and chromosome movement. Although cytoplasmic tubules and fibers have been observed in bacteria, some with diameters similar to those of eukaryotes, no homologies to eukaryotic microtubules have been established. Certain groups of bacteria including azotobacters, cyanobacteria, enteric bacteria, and spirochetes have been frequently observed to possess microtubule-like structures, and others, including archaebacteria, have been shown to be sensitive to drugs that inhibit the polymerization of microtubules. Although little biochemical or molecular biological information is available, the differences observed among these prokaryotic structures suggest that their composition generally differs among themselves as well as from that of eukaryotes. We review the distribution of cytoplasmic tubules in prokaryotes, even though, in all cases, their functions remain unknown. At least some tend to occur in cells that are large, elongate, and motile, suggesting that they may be involved in cytoskeletal functions, intracellular motility, or transport activities comparable to those performed by eukaryotic microtubules. In Escherichia coli, the FtsZ protein is associated with the formation of a ring in the division zone between the newly forming offspring cells. Like tubulin, FtsZ is a GTPase and shares with tubulin a 7-amino-acid motif, making it a promising candidate in which to seek the origin of tubulins.

Prokaryotic diversity and dynamics in a full-scale municipal solid waste anaerobic reactor from start-up to steady-state conditions.

PubMed

Cardinali-Rezende, Juliana; Colturato, Luís F D B; Colturato, Thiago D B; Chartone-Souza, Edmar; Nascimento, Andréa M A; Sanz, José L

2012-09-01

The prokaryotic diversity of an anaerobic reactor for the treatment of municipal solid waste was investigated over the course of 2 years with the use of 16S rDNA-targeted molecular approaches. The fermentative Bacteroidetes and Firmicutes predominated, and Proteobacteria, Actinobacteria, Tenericutes and the candidate division WWE1 were also identified. Methane production was dominated by the hydrogenotrophic Methanomicrobiales (Methanoculleus sp.) and their syntrophic association with acetate-utilizing and propionate-oxidizing bacteria. qPCR demonstrated the predominance of the hydrogenotrophic over aceticlastic Methanosarcinaceae (Methanosarcina sp. and Methanimicrococcus sp.), and Methanosaetaceae (Methanosaeta sp.) were measured in low numbers in the reactor. According to the FISH and CARD-FISH analyses, Bacteria and Archaea accounted for 85% and 15% of the cells, respectively. Different cell counts for these domains were obtained by qPCR versus FISH analyses. The use of several molecular tools increases our knowledge of the prokaryotic community dynamics from start-up to steady-state conditions in a full-scale MSW reactor. Copyright © 2012 Elsevier Ltd. All rights reserved.

Proteolytic enzymes in seawater: contribution of prokaryotes and protists

NASA Astrophysics Data System (ADS)

Obayashi, Y.; Suzuki, S.

2016-02-01

Proteolytic enzyme is one of the major catalysts of microbial processing of organic matter in biogeochemical cycle. Here we summarize some of our studies about proteases in seawater, including 1) distribution of protease activities in coastal and oceanic seawater, 2) responses of microbial community and protease activities in seawater to organic matter amending, and 3) possible contribution of heterotrophic protists besides prokaryotes to proteases in seawater, to clarify cleared facts and remaining questions. Activities of aminopeptidases, trypsin-type and chymotrypsin-type proteases were detected from both coastal and oceanic seawater by using MCA-substrate assay. Significant activities were detected from not only particulate (cell-associated) fraction but also dissolved fraction of seawater, especially for trypsin-type and chymotrypsin-type proteases. Hydrolytic enzymes in seawater have been commonly thought to be mainly derived from heterotrophic prokaryotes; however, it was difficult to determine actual source organisms of dissolved enzymes in natural seawater. Our experiment with addition of dissolved protein to subtropical oligotrophic Pacific water showed drastically enhancement of the protease activities especially aminopeptidases in seawater, and the prokaryotic community structure simultaneously changed to be dominant of Bacteroidetes, indicating that heterotrophic bacteria were actually one of the sources of proteases in seawater. Another microcosm experiment with free-living marine heterotrophic ciliate Paranophrys marina together with an associated bacterium showed that extracellular trypsin-type activity was mainly attributed to the ciliate. The protist seemed to work in organic matter digestion in addition to be a grazer. From the results, we propose a system of organic matter digestion by prokaryotes and protists in aquatic environments, although their actual contribution in natural environments should be estimated in future studies.

Metabolic profiles of prokaryotic and eukaryotic communities in deep-sea sponge Neamphius huxleyi [corrected]. indicated by metagenomics.

PubMed

Li, Zhi-Yong; Wang, Yue-Zhu; He, Li-Ming; Zheng, Hua-Jun

2014-01-27

The whole metabolism of a sponge holobiont and the respective contributions of prokaryotic and eukaryotic symbionts and their associations with the sponge host remain largely unclear. Meanwhile, compared with shallow water sponges, deep-sea sponges are rarely understood. Here we report the metagenomic exploration of deep-sea sponge Neamphius huxleyi [corrected] . at the whole community level. Metagenomic data showed phylogenetically diverse prokaryotes and eukaryotes in Neamphius huxleyi [corrected]. MEGAN and gene enrichment analyses indicated different metabolic potentials of prokaryotic symbionts from eukaryotic symbionts, especially in nitrogen and carbon metabolisms, and their molecular interactions with the sponge host. These results supported the hypothesis that prokaryotic and eukaryotic symbionts have different ecological roles and relationships with sponge host. Moreover, vigorous denitrification, and CO2 fixation by chemoautotrophic prokaryotes were suggested for this deep-sea sponge. The study provided novel insights into the respective potentials of prokaryotic and eukaryotic symbionts and their associations with deep-sea sponge Neamphius huxleyi [corrected].

Emerging facets of prokaryotic glycosylation

PubMed Central

Schäffer, Christina; Messner, Paul

2017-01-01

Glycosylation of proteins is one of the most prevalent post-translational modifications occurring in nature, with a wide repertoire of biological implications. Pathways for the main types of this modification, the N- and O-glycosylation, can be found in all three domains of life—the Eukarya, Bacteria and Archaea—thereby following common principles, which are valid also for lipopolysaccharides, lipooligosaccharides and glycopolymers. Thus, studies on any glycoconjugate can unravel novel facets of the still incompletely understood fundamentals of protein N- and O-glycosylation. While it is estimated that more than two-thirds of all eukaryotic proteins would be glycosylated, no such estimate is available for prokaryotic glycoproteins, whose understanding is lagging behind, mainly due to the enormous variability of their glycan structures and variations in the underlying glycosylation processes. Combining glycan structural information with bioinformatic, genetic, biochemical and enzymatic data has opened up an avenue for in-depth analyses of glycosylation processes as a basis for glycoengineering endeavours. Here, the common themes of glycosylation are conceptualised for the major classes of prokaryotic (i.e. bacterial and archaeal) glycoconjugates, with a special focus on glycosylated cell-surface proteins. We describe the current knowledge of biosynthesis and importance of these glycoconjugates in selected pathogenic and beneficial microbes. PMID:27566466

Superstatistical model of bacterial DNA architecture

NASA Astrophysics Data System (ADS)

Bogachev, Mikhail I.; Markelov, Oleg A.; Kayumov, Airat R.; Bunde, Armin

2017-02-01

Understanding the physical principles that govern the complex DNA structural organization as well as its mechanical and thermodynamical properties is essential for the advancement in both life sciences and genetic engineering. Recently we have discovered that the complex DNA organization is explicitly reflected in the arrangement of nucleotides depicted by the universal power law tailed internucleotide interval distribution that is valid for complete genomes of various prokaryotic and eukaryotic organisms. Here we suggest a superstatistical model that represents a long DNA molecule by a series of consecutive ~150 bp DNA segments with the alternation of the local nucleotide composition between segments exhibiting long-range correlations. We show that the superstatistical model and the corresponding DNA generation algorithm explicitly reproduce the laws governing the empirical nucleotide arrangement properties of the DNA sequences for various global GC contents and optimal living temperatures. Finally, we discuss the relevance of our model in terms of the DNA mechanical properties. As an outlook, we focus on finding the DNA sequences that encode a given protein while simultaneously reproducing the nucleotide arrangement laws observed from empirical genomes, that may be of interest in the optimization of genetic engineering of long DNA molecules.

DNA Knots: Theory and Experiments

NASA Astrophysics Data System (ADS)

Sumners, D. W.

Cellular DNA is a long, thread-like molecule with remarkably complex topology. Enzymes that manipulate the geometry and topology of cellular DNA perform many vital cellular processes (including segregation of daughter chromosomes, gene regulation, DNA repair, and generation of antibody diversity). Some enzymes pass DNA through itself via enzyme-bridged transient breaks in the DNA; other enzymes break the DNA apart and reconnect it to different ends. In the topological approach to enzymology, circular DNA is incubated with an enzyme, producing an enzyme signature in the form of DNA knots and links. By observing the changes in DNA geometry (supercoiling) and topology (knotting and linking) due to enzyme action, the enzyme binding and mechanism can often be characterized. This paper will discuss some personal research history, and the tangle model for the analysis of site-specific recombination experiments on circular DNA.

Source tracking of prokaryotic communities in fermented grain of Chinese strong-flavor liquor.

PubMed

Wang, Xueshan; Du, Hai; Xu, Yan

2017-03-06

The fermentation process of Chinese strong-flavor liquor involves numerous microbes originating from Daqu and pit mud. Daqu is the starter of fermentation, and pit mud acts as another source of inoculum of microbes in the liquor-making process. However, the contribution of microbes in pit mud and Daqu to fermented grain, and the sources of microbes in fermented grain are still waiting to be defined clearly. In this study, prokaryotic communities in fermented grain, pit mud and Daqu were identified via next generation sequencing of the V4 region of 16S rRNA gene. Principal-coordinate analysis indicated that Daqu had stronger influence on the prokaryotic communities in fermented grain at the prophase of fermentation, but pit mud influenced the fermented grain continuously during the whole fermentation process. Totally, 299 genera were detected in all fermented grain, pit mud and Daqu samples. Among them, 204 genera were detected in 3days' fermented grain. Ten genera (Lactobacillus, Leuconostoc, Staphylococcus, Gluconobacter, Acetobacter, Petrimonas, Clostridium, Ruminococcus, Methanobacterium and Methanobrevibacter) were dominant, and accounted for 84.31%-87.13% relative abundance of the total prokaryotic community in fermented grain. Venn analysis indicated Daqu was the main source of strict aerobes and facultative aerobes, which took up over 74% of prokaryotic communities in fermented grain. Conversely, pit mud was the sustained-release source of anaerobes, which accounted for over 14% of prokaryotic communities in fermented grain. In addition, part of anaerobes originated from both Daqu and pit mud. This study could help track the source of prokaryotic communities in fermented grain, and improve the quality and controllability in liquor production. Copyright © 2017 Elsevier B.V. All rights reserved.

Assessment of DNA Contamination in RNA Samples Based on Ribosomal DNA

PubMed Central

Hashemipetroudi, Seyyed Hamidreza; Nematzadeh, Ghorbanali; Ahmadian, Gholamreza; Yamchi, Ahad; Kuhlmann, Markus

2018-01-01

One method extensively used for the quantification of gene expression changes and transcript abundances is reverse-transcription quantitative real-time PCR (RT-qPCR). It provides accurate, sensitive, reliable, and reproducible results. Several factors can affect the sensitivity and specificity of RT-qPCR. Residual genomic DNA (gDNA) contaminating RNA samples is one of them. In gene expression analysis, non-specific amplification due to gDNA contamination will overestimate the abundance of transcript levels and can affect the RT-qPCR results. Generally, gDNA is detected by qRT-PCR using primer pairs annealing to intergenic regions or an intron of the gene of interest. Unfortunately, intron/exon annotations are not yet known for all genes from vertebrate, bacteria, protist, fungi, plant, and invertebrate metazoan species. Here we present a protocol for detection of gDNA contamination in RNA samples by using ribosomal DNA (rDNA)-based primers. The method is based on the unique features of rDNA: their multigene nature, highly conserved sequences, and high frequency in the genome. Also as a case study, a unique set of primers were designed based on the conserved region of ribosomal DNA (rDNA) in the Poaceae family. The universality of these primer pairs was tested by melt curve analysis and agarose gel electrophoresis. Although our method explains how rDNA-based primers can be applied for the gDNA contamination assay in the Poaceae family, it could be easily used to other prokaryote and eukaryote species PMID:29443017

Gene duplications in prokaryotes can be associated with environmental adaptation

PubMed Central

2010-01-01

Background Gene duplication is a normal evolutionary process. If there is no selective advantage in keeping the duplicated gene, it is usually reduced to a pseudogene and disappears from the genome. However, some paralogs are retained. These gene products are likely to be beneficial to the organism, e.g. in adaptation to new environmental conditions. The aim of our analysis is to investigate the properties of paralog-forming genes in prokaryotes, and to analyse the role of these retained paralogs by relating gene properties to life style of the corresponding prokaryotes. Results Paralogs were identified in a number of prokaryotes, and these paralogs were compared to singletons of persistent orthologs based on functional classification. This showed that the paralogs were associated with for example energy production, cell motility, ion transport, and defence mechanisms. A statistical overrepresentation analysis of gene and protein annotations was based on paralogs of the 200 prokaryotes with the highest fraction of paralog-forming genes. Biclustering of overrepresented gene ontology terms versus species was used to identify clusters of properties associated with clusters of species. The clusters were classified using similarity scores on properties and species to identify interesting clusters, and a subset of clusters were analysed by comparison to literature data. This analysis showed that paralogs often are associated with properties that are important for survival and proliferation of the specific organisms. This includes processes like ion transport, locomotion, chemotaxis and photosynthesis. However, the analysis also showed that the gene ontology terms sometimes were too general, imprecise or even misleading for automatic analysis. Conclusions Properties described by gene ontology terms identified in the overrepresentation analysis are often consistent with individual prokaryote lifestyles and are likely to give a competitive advantage to the organism

Segregation and manifestations of the mtDNA tRNA[sup Lys] A[r arrow]G[sup (8344)] mutation of myoclonus epilepsy and ragged-red fibers (MERRF) syndrome

DOE Office of Scientific and Technical Information (OSTI.GOV)

Larsson, N.G.; Tulinius, M.H.; Holme, E.

1992-12-01

The authors have studied the segregation and manifestations of the tRNA[sup Lys] A[r arrow]G[sup (8344)] mutation of mtDNA. Three unrelated patients with myoclonus epilepsy and ragged-red fibers (MERRF) syndrome were investigated, along with 30 of their maternal relatives. Mutated mtDNA was not always found in the offspring of women carrying the tRNA[sup Lys] mutation. Four women had 10%-33% of mutated mtDNA in lymphocytes, and no mutated mtDNA was found in 7 of their 14 investigated children. The presence of mutated mtDNA was excluded at a level of 3:1,000. Five women had a proportion of 43%-73% mutated mtDNA in lymphocytes, andmore » mutated mtDNA was found in all their 12 investigated children. This suggests that the risk for transmission of mutated mtDNA to the offspring increases if high levels are present in the mother and that, above a threshold level of 35%-40%, it is very likely that transmission will occur to all children. The three patients with MERRF syndrone had, in muscle, both 94%-96% mutated mtDNA and biochemical and histochemical evidence of a respiratory-chain dysfunction. Four relatives had a proportion of 61%-92% mutated mtDNA in muscle, and biochemical measurements showed a normal respiratory-chain function in muscle in all cases. These findings suggest that >92% of mtDNA with the tRNA[sup Lys] mutation in muscle is required to cause a respiratory-chain dysfunction that can be detected by biochemical methods. There was a positive correlation between the levels of mtDNA with the tRNA[sup Lys] mutation in lymphocytes and the levels in muscle, in all nine investigated cases. The levels of mutated mtDNA were higher in muscle than in lymphocytes in all cases. 30 refs., 3 figs., 5 tabs.« less

Residential Segregation and School Integration.

ERIC Educational Resources Information Center

Rivkin, Steven G.

1994-01-01

Asserts that school districts' efforts to integrate schools have failed to ameliorate the racial isolation of black students. Finds that schools remain segregated primarily because of continued residential segregation and that school integration efforts have had little long-term effect on residential segregation. (CFR)

Understanding Segregation Processes

NASA Astrophysics Data System (ADS)

Bruch, Elizabeth

There is growing consensus that living in neighborhoods of concentrated poverty increases the likelihood of social problems such as teenage parenthood, drug and alcohol use, crime victimization, and chronic unemployment. Neighborhood inequality is also implicated in studies of enduring race/ethnic health disparities, and there are recent moves to broaden the definition of health care policy to policies targeting social inequality (Mechanic 2007). Residential segregation affects health outcomes in several different ways. First, income, education, and occupation are all strongly related to health (Adler and Newman 2002). Segregation is a key mechanism through which socioeconomic inequality is perpetuated and reinforced, as it hinders the upward mobility of disadvantaged groups by limiting their educational and employment opportunities. Second, segregation increases minority exposure to unhealthy neighborhood environments. Residential segregation creates areas with concentrated poverty and unemployment, both of which are key factors that predict violence and create racial differences in homicide (Samson and Wilson 1995). Neighborhood characteristics, such as exposure to environmental hazards, fear of violence, and access to grocery stores, affect health risks and health behaviors (Cheadle et al. 1991). Tobacco and alcohol industries also advertise their products disproportionately in poor, minority areas (Moore, Williams, and Qualls 1996). Finally, residential segregation leads to inequalitie in health care resources, which contributes to disparities in quality of treatment (Smedley, Stith, and Nelson 2002).

Generative Models of Segregation: Investigating Model-Generated Patterns of Residential Segregation by Ethnicity and Socioeconomic Status

PubMed Central

Fossett, Mark

2011-01-01

This paper considers the potential for using agent models to explore theories of residential segregation in urban areas. Results of generative experiments conducted using an agent-based simulation of segregation dynamics document that varying a small number of model parameters representing constructs from urban-ecological theories of segregation can generate a wide range of qualitatively distinct and substantively interesting segregation patterns. The results suggest how complex, macro-level patterns of residential segregation can arise from a small set of simple micro-level social dynamics operating within particular urban-demographic contexts. The promise and current limitations of agent simulation studies are noted and optimism is expressed regarding the potential for such studies to engage and contribute to the broader research literature on residential segregation. PMID:21379372

Crystal structure of a suicidal DNA repair protein: the Ada O6-methylguanine-DNA methyltransferase from E. coli.

PubMed

Moore, M H; Gulbis, J M; Dodson, E J; Demple, B; Moody, P C

1994-04-01

The mutagenic and carcinogenic effects of simple alkylating agents are mainly due to methylation at the O6 position of guanine in DNA. O6-methylguanine directs the incorporation of either thymine or cytosine without blocking DNA replication, resulting in GC to AT transition mutations. In prokaryotic and eukaryotic cells antimutagenic repair is effected by direct reversal of this DNA damage. A suicidal methyltransferase repair protein removes the methyl group from DNA to one of its own cysteine residues. The resulting self-methylation of the active site cysteine renders the protein inactive. Here we report the X-ray structure of the 19 kDa C-terminal domain of the Escherichia coli ada gene product, the prototype of these suicidal methyltransferases. In the crystal structure the active site cysteine is buried. We propose a model for the significant conformational change that the protein must undergo in order to bind DNA and effect methyl transfer.

DNA N(6)-methyladenine: a new epigenetic mark in eukaryotes?

PubMed

Luo, Guan-Zheng; Blanco, Mario Andres; Greer, Eric Lieberman; He, Chuan; Shi, Yang

2015-12-01

DNA N(6)-adenine methylation (N(6)-methyladenine; 6mA) in prokaryotes functions primarily in the host defence system. The prevalence and significance of this modification in eukaryotes had been unclear until recently. Here, we discuss recent publications documenting the presence of 6mA in Chlamydomonas reinhardtii, Drosophila melanogaster and Caenorhabditis elegans; consider possible roles for this DNA modification in regulating transcription, the activity of transposable elements and transgenerational epigenetic inheritance; and propose 6mA as a new epigenetic mark in eukaryotes.

Single-molecule study of DNA unlinking by eukaryotic and prokaryotic type-II topoisomerases

PubMed Central

Charvin, G.; Bensimon, D.; Croquette, V.

2003-01-01

Type-II topoisomerases are responsible for untangling DNA during replication by removing supercoiled and interlinked DNA structures. Using a single-molecule micromanipulation setup, we follow the real-time decatenation of two mechanically braided DNA molecules by Drosophila melanogaster topoisomerase (Topo) II and Escherichia coli Topo IV. Although Topo II relaxes left-handed (L) and right-handed (R-) braids similarly at a rate of ≈2.9 s–1, Topo IV has a marked preference for L-braids, which it relaxes completely and processively at a rate of ≈2.4 s–1. However, Topo IV can unlink R-braids at about half that rate when they supercoil to form L-plectonemes. These results imply that the preferred substrate for unlinking by Topo IV has the symmetry of an L-crossing and shed new light on the decatenation of daughter strands during DNA replication, which are usually assumed to be linked in an R-braid. PMID:12902541

Discovering non-random segregation of sister chromatids: the naïve treatment of a premature discovery

PubMed Central

Lark, Karl G.

2013-01-01

The discovery of non-random chromosome segregation (Figure 1) is discussed from the perspective of what was known in 1965 and 1966. The distinction between daughter, parent, or grandparent strands of DNA was developed in a bacterial system and led to the discovery that multiple copies of DNA elements of bacteria are not distributed randomly with respect to the age of the template strand. Experiments with higher eukaryotic cells demonstrated that during mitosis Mendel’s laws were violated; and the initial serendipitous choice of eukaryotic cell system led to the striking example of non-random segregation of parent and grandparent DNA template strands in primary cultures of cells derived from mouse embryos. Attempts to extrapolate these findings to established tissue culture lines demonstrated that the property could be lost. Experiments using plant root tips demonstrated that the phenomenon exists in plants and that it was, at some level, under genetic control. Despite publication in major journals and symposia (Lark et al., 1966, 1967; Lark, 1967, 1969a,b,c) the potential implications of these findings were ignored for several decades. Here we explore possible reasons for the pre-maturity (Stent, 1972) of this discovery. PMID:23378946

Nanoscale Spatial Organization of Prokaryotic Cells Studied by Super-Resolution Optical Microscopy

NASA Astrophysics Data System (ADS)

McEvoy, Andrea Lynn

All cells spatially organize their interiors, and this arrangement is necessary for cell viability. Until recently, it was believed that only eukaryotic cells spatially segregate their components. However, it is becoming increasingly clear that bacteria also assemble their proteins into complex patterns. In eukaryotic cells, spatial organization arises from membrane bound organelles as well as motor transport proteins which can move cargos within the cell. To date, there are no known motor transport proteins in bacteria and most microbes lack membrane bound organelles, so it remains a mystery how bacterial spatial organization emerges. In hind-sight it is not surprising that bacteria also exhibit complex spatial organization considering much of what we have learned about the basic processes that take place in all cells, such as transcription and translation was first discovered in prokaryotic cells. Perhaps the fundamental principles that govern spatial organization in prokaryotic cells may be applicable in eukaryotic cells as well. In addition, bacteria are attractive model organism for spatial organization studies because they are genetically tractable, grow quickly and much biochemical and structural data is known about them. A powerful tool for observing spatial organization in cells is the fluorescence microscope. By specifically tagging a protein of interest with a fluorescent probe, it is possible to examine how proteins organize and dynamically assemble inside cells. A significant disadvantage of this technology is its spatial resolution (approximately 250 nm laterally and 500 nm axially). This limitation on resolution causes closely spaced proteins to look blurred making it difficult to observe the fine structure within the complexes. This resolution limit is especially problematic within small cells such as bacteria. With the recent invention of new optical microscopies, we now can surpass the existing limits of fluorescence imaging. In some cases, we can

Effects of viruses and predators on prokaryotic community composition.

PubMed

Jardillier, Ludwig; Bettarel, Yvan; Richardot, Mathilde; Bardot, Corinne; Amblard, Christian; Sime-Ngando, Télesphore; Debroas, Didier

2005-11-01

Dialysis bags were used to examine the impact of predation and viral lysis on prokaryotic community composition (PCC) over a 5-day experiment in the oligomesotrophic Lake Pavin (France). The impact of the different predator communities (protists and metazoans) of prokaryotes was estimated by water fractionation (<5 microm: treatment filtered on 5 microm, without ciliates and metazoans; UNF: unfiltered treatment with all planktonic communities). Enrichments of natural viruses (<1.2 microm: with a natural virus concentration; <1.2 mum V and VV: with enrichment leading to a double or triple concentration of viruses, respectively) were used to indirectly assess the control of virioplankton. Viral activity was estimated from the frequency of visibly infected cells (FVIC). PCC was determined by fluorescence in situ hybridization (FISH) and terminal restriction fragment length polymorphism (T-RFLP). In this study, PCC was affected by the eukaryote communities (especially flagellates), and viruses to a lesser extent. Cyanobacteria declined significantly during the experiment and were highly correlated with the FVIC. In addition, the 503-bp terminal restriction fragment (T-RF) disappeared in treatments with virus enrichments, suggesting possible viral-associated mortality processes, whereas the 506-bp T-RF was not affected in these treatments. On one hand, these results suggest a control of the PCC: first, by viral lysis of some dominant phylotypes and second, by interspecific competition between resistant strains for the uptake of substrates released by this lysis. The increase of Archaea may suggest that these cells benefit such resources. On the other hand, the disappearance and the stable proportion of some dominant phylotypes suggested a selection pressure due to the predatory activity on prokaryotes. In conclusion, prokaryotic abundance appears to be mainly controlled by flagellate protists, which also affected PCC, whereas viruses seemed to be essentially responsible

Eukaryotic ribosome display with in situ DNA recovery.

PubMed

He, Mingyue; Edwards, Bryan M; Kastelic, Damjana; Taussig, Michael J

2012-01-01

Ribosome display is a cell-free display technology for in vitro selection and optimisation of proteins from large diversified libraries. It operates through the formation of stable protein-ribosome-mRNA (PRM) complexes and selection of ligand-binding proteins, followed by DNA recovery from the selected genetic information. Both prokaryotic and eukaryotic ribosome display systems have been developed. In this chapter, we describe the eukaryotic rabbit reticulocyte method in which a distinct in situ single-primer RT-PCR procedure is used to recover DNA from the selected PRM complexes without the need for prior disruption of the ribosome.

Oligonucleotide Microarray for 16S rRNA Gene-Based Detection of All Recognized Lineages of Sulfate-Reducing Prokaryotes in the Environment

PubMed Central

Loy, Alexander; Lehner, Angelika; Lee, Natuschka; Adamczyk, Justyna; Meier, Harald; Ernst, Jens; Schleifer, Karl-Heinz; Wagner, Michael

2002-01-01

For cultivation-independent detection of sulfate-reducing prokaryotes (SRPs) an oligonucleotide microarray consisting of 132 16S rRNA gene-targeted oligonucleotide probes (18-mers) having hierarchical and parallel (identical) specificity for the detection of all known lineages of sulfate-reducing prokaryotes (SRP-PhyloChip) was designed and subsequently evaluated with 41 suitable pure cultures of SRPs. The applicability of SRP-PhyloChip for diversity screening of SRPs in environmental and clinical samples was tested by using samples from periodontal tooth pockets and from the chemocline of a hypersaline cyanobacterial mat from Solar Lake (Sinai, Egypt). Consistent with previous studies, SRP-PhyloChip indicated the occurrence of Desulfomicrobium spp. in the tooth pockets and the presence of Desulfonema- and Desulfomonile-like SRPs (together with other SRPs) in the chemocline of the mat. The SRP-PhyloChip results were confirmed by several DNA microarray-independent techniques, including specific PCR amplification, cloning, and sequencing of SRP 16S rRNA genes and the genes encoding the dissimilatory (bi)sulfite reductase (dsrAB). PMID:12324358

Distinct distribution patterns of prokaryotes between sediment and water in the Yellow River estuary.

PubMed

Wei, Guangshan; Li, Mingcong; Li, Fenge; Li, Han; Gao, Zheng

2016-11-01

There are close exchanges between sediment and water in estuaries; however, the patterns of prokaryotic community assembly in these two habitat types are still unclear. This study investigated the bacterial and archaeal abundance, diversity, and community composition in the sediment and the overlying water of the Yellow River estuary. Notably higher prokaryotic abundance and diversity were detected in the sediment than in the water, and bacterial abundance and diversity were remarkably higher than those of archaea. Furthermore, the ratio of bacterial to archaeal 16S rRNA gene abundance was significantly lower in the sediment than in the water. Bacterial communities at different taxonomic levels were apparently distinct between the sediment and water, but archaeal communities were not. The most dominant bacteria were affiliated with Deltaproteobacteria and Gammaproteobacteria in sediment and with Alphaproteobacteria and Betaproteobacteria in water. Euryarchaeota and Thaumarchaeota were the most abundant archaea in both habitats. Although distinct prokaryotic distribution patterns were observed, most of the dominant bacteria and archaea present were related to carbon, nitrogen, and sulfur cycling processes, such as methanogenesis, ammonia oxidation, and sulfate reduction. Unexpectedly, prokaryotes from the water showed a higher sensitivity to environmental factors, while only a few factors affected sediment communities. Additionally, some potential co-occurrence relationships between prokaryotes were also found in this study. These results suggested distinct distribution patterns of bacterial and archaeal communities between sediment and overlying water in this important temperate estuary, which may serve as a useful community model for the further ecological and evolutionary study of prokaryotes in estuarine ecosystems.

Autonomous generation and loading of DNA guides by bacterial Argonaute

PubMed Central

Chandradoss, Stanley D.; Zhu, Yifan; Timmers, Elizabeth M.; Zhang, Yong; Zhao, Hongtu; Lou, Jizhong; Wang, Yanli; Joo, Chirlmin; van der Oost, John

2018-01-01

Summary Several prokaryotic Argonaute proteins (pAgos) utilize small DNA guides to mediate host defense by targeting invading DNA complementary to the DNA guide. It is unknown how these DNA guides are being generated and loaded onto pAgo. Here we demonstrate that guide-free Argonaute from Thermus thermophilus (TtAgo) can degrade dsDNA, thereby generating small dsDNA fragments that subsequently are loaded onto TtAgo. Combining single-molecule fluorescence, molecular dynamic simulations and structural studies, we show that TtAgo loads dsDNA molecules with a preference towards a deoxyguanosine on the passenger strand at the position opposite to the 5’-end of the guide strand. This explains why in vivo TtAgo is preferentially loaded with guides with a 5’-end deoxycytidine. Our data demonstrate that TtAgo can independently generate and selectively load functional DNA guides. PMID:28262506

P2RP: a Web-based framework for the identification and analysis of regulatory proteins in prokaryotic genomes.

PubMed

Barakat, Mohamed; Ortet, Philippe; Whitworth, David E

2013-04-20

Regulatory proteins (RPs) such as transcription factors (TFs) and two-component system (TCS) proteins control how prokaryotic cells respond to changes in their external and/or internal state. Identification and annotation of TFs and TCSs is non-trivial, and between-genome comparisons are often confounded by different standards in annotation. There is a need for user-friendly, fast and convenient tools to allow researchers to overcome the inherent variability in annotation between genome sequences. We have developed the web-server P2RP (Predicted Prokaryotic Regulatory Proteins), which enables users to identify and annotate TFs and TCS proteins within their sequences of interest. Users can input amino acid or genomic DNA sequences, and predicted proteins therein are scanned for the possession of DNA-binding domains and/or TCS domains. RPs identified in this manner are categorised into families, unambiguously annotated, and a detailed description of their features generated, using an integrated software pipeline. P2RP results can then be outputted in user-specified formats. Biologists have an increasing need for fast and intuitively usable tools, which is why P2RP has been developed as an interactive system. As well as assisting experimental biologists to interrogate novel sequence data, it is hoped that P2RP will be built into genome annotation pipelines and re-annotation processes, to increase the consistency of RP annotation in public genomic sequences. P2RP is the first publicly available tool for predicting and analysing RP proteins in users' sequences. The server is freely available and can be accessed along with documentation at http://www.p2rp.org.

Segregation as Splitting, Segregation as Joining: Schools, Housing, and the Many Modes of Jim Crow

ERIC Educational Resources Information Center

Highsmith, Andrew R.; Erickson, Ansley T.

2015-01-01

Popular understandings of segregation often emphasize the Jim Crow South before the 1954 "Brown" decision and, in many instances, explain continued segregation in schooling as the result of segregated housing patterns. The case of Flint, Michigan, complicates these views, at once illustrating the depth of governmental commitment to…

Employing 454 amplicon pyrosequencing to reveal intragenomic divergence in the internal transcribed spacer rDNA region in fungi

Treesearch

Daniel L. Lindner; Tor Carlsen; Henrik Nilsson; Marie Davey; Trond Schumacher; Havard. Kauserud

2013-01-01

The rDNA internal transcribed spacer (ITS) region has been accepted as a DNA barcoding marker for fungi and is widely used in phylogenetic studies; however, intragenomic ITS variability has been observed in a broad range of taxa, including prokaryotes, plants, animals, and fungi, and this variability has the potential to inflate species richness estimates in molecular...

Segregation in Post-Civil Rights America: Stalled Integration or End of the Segregated Century?

PubMed Central

Massey, Douglas S.; Rugh, Jacob S.

2016-01-01

In this paper we adjudicate between competing claims of persisting segregation and rapid integration by analyzing trends in residential dissimilarity and spatial isolation for African Americans, Hispanics, and Asians living in 287 consistently defined metropolitan areas from 1970 to 2010. On average, black segregation and isolation have fallen steadily but still remain very high in many areas, particularly those areas historically characterized by hypersegregation. In contrast, Hispanic segregation has increased slightly but Hispanic isolation has risen substantially owing to rapid population growth. Asian segregation has changed little and remains moderate, and although Asian isolation has increased it remains at low levels compared with other groups. Multivariate analyses reveal that segregation and isolation are being actively produced in some areas by restrictive density zoning regimes, large and/or rising minority percentages, lagging minority socioeconomic status, and active expressions of anti-black and anti-Latino sentiment, especially in large metropolitan areas. Areas displaying these characteristics are either integrating very slowly (in the case of blacks) or becoming more segregated (in the case of Hispanics), whereas those lacking these attributes are clearly moving toward integration, often quite rapidly. PMID:26966459

Diversity and Distribution of Prokaryotes within a Shallow-Water Pockmark Field.

PubMed

Giovannelli, Donato; d'Errico, Giuseppe; Fiorentino, Federica; Fattorini, Daniele; Regoli, Francesco; Angeletti, Lorenzo; Bakran-Petricioli, Tatjana; Vetriani, Costantino; Yücel, Mustafa; Taviani, Marco; Manini, Elena

2016-01-01

Pockmarks are crater-like depression on the seafloor associated with hydrocarbon ascent through muddy sediments in continental shelves around the world. In this study, we examine the diversity and distribution of benthic microbial communities at shallow-water pockmarks adjacent to the Middle Adriatic Ridge. We integrate microbial diversity data with characterization of local hydrocarbons concentrations and sediment geochemistry. Our results suggest these pockmarks are enriched in sedimentary hydrocarbons, and host a microbial community dominated by Bacteria, even in deeper sediment layers. Pockmark sediments showed higher prokaryotic abundance and biomass than surrounding sediments, potentially due to the increased availability of organic matter and higher concentrations of hydrocarbons linked to pockmark activity. Prokaryotic diversity analyses showed that the microbial communities of these shallow-water pockmarks are unique, and comprised phylotypes associated with the cycling of sulfur and nitrate compounds, as well as numerous know hydrocarbon degraders. Altogether, this study suggests that shallow-water pockmark habitats enhance the diversity of the benthic prokaryotic biosphere by providing specialized environmental niches.

Thiol biochemistry of prokaryotes

NASA Technical Reports Server (NTRS)

Fahey, Robert C.

1986-01-01

The present studies have shown that GSH metabolism arose in the purple bacteria and cyanobacteria where it functions to protect against oxygen toxicity. Evidence was obtained indicating that GSH metabolism was incorporated into eucaryotes via the endosymbiosis giving rise to mitochrondria and chloroplasts. Aerobic bacteria lacking GSH utilize other thiols for apparently similar functions, the thiol being coenzyme A in Gram positive bacteria and chi-glutamylcysteine in the halobacteria. The thiol biochemistry of prokaryotes is thus seen to be much more highly diversified than that of eucaryotes and much remains to be learned about this subject.

Partial pedigree analysis of the segregation of yeast mitochondrial genes during vegetative reproduction.

PubMed

Waxman, M F; Birk, C W

1982-08-01

A three-factor cross of Saccharomyces cerevisiae involving the cap1, ery1, and oli1 loci was done, with partial pedigree analyses of 117 zygotes. First, second, and third buds were removed and the genotypes of their diploid progeny determined, along with those of the residual zygote mother cell. Results were analyzed in terms of frequencies of individual alleles and of recombinant genotypes in the dividing cells. There is a gradual increase in the frequency of homoplasmic cells and in gene frequency variance during these three generations, as would result from stochastic partitioning of mtDNA molecules between mother and bud, probably coupled with random drift of gene frequencies in interphase cells. These phenomena are more pronounced for buds than for mothers, suggesting that buds receive a smaller sample of molecules. End buds are more likely to be homoplasmic and have a lower frequency of recombinant genotypes than do central buds; an end bud is particularly enriched in alleles contributed by the parent that formed that end of the zygote. Zygotes with first central buds produce clones with a higher recombination frequency than do those with first end buds. These results confirm previous studies and suggest that mixing of parental genotypes occurs first in the center of the zygote. If segregation were strictly random, the number of segregating units would have to be much smaller than the number of mtDNA molecules in the zygote. On the other hand, there is no evidence for a region of the molecule ("attachment point") which segregates deterministically.

A computational genomics pipeline for prokaryotic sequencing projects.

PubMed

Kislyuk, Andrey O; Katz, Lee S; Agrawal, Sonia; Hagen, Matthew S; Conley, Andrew B; Jayaraman, Pushkala; Nelakuditi, Viswateja; Humphrey, Jay C; Sammons, Scott A; Govil, Dhwani; Mair, Raydel D; Tatti, Kathleen M; Tondella, Maria L; Harcourt, Brian H; Mayer, Leonard W; Jordan, I King

2010-08-01

New sequencing technologies have accelerated research on prokaryotic genomes and have made genome sequencing operations outside major genome sequencing centers routine. However, no off-the-shelf solution exists for the combined assembly, gene prediction, genome annotation and data presentation necessary to interpret sequencing data. The resulting requirement to invest significant resources into custom informatics support for genome sequencing projects remains a major impediment to the accessibility of high-throughput sequence data. We present a self-contained, automated high-throughput open source genome sequencing and computational genomics pipeline suitable for prokaryotic sequencing projects. The pipeline has been used at the Georgia Institute of Technology and the Centers for Disease Control and Prevention for the analysis of Neisseria meningitidis and Bordetella bronchiseptica genomes. The pipeline is capable of enhanced or manually assisted reference-based assembly using multiple assemblers and modes; gene predictor combining; and functional annotation of genes and gene products. Because every component of the pipeline is executed on a local machine with no need to access resources over the Internet, the pipeline is suitable for projects of a sensitive nature. Annotation of virulence-related features makes the pipeline particularly useful for projects working with pathogenic prokaryotes. The pipeline is licensed under the open-source GNU General Public License and available at the Georgia Tech Neisseria Base (http://nbase.biology.gatech.edu/). The pipeline is implemented with a combination of Perl, Bourne Shell and MySQL and is compatible with Linux and other Unix systems.

DMINDA: an integrated web server for DNA motif identification and analyses.

PubMed

Ma, Qin; Zhang, Hanyuan; Mao, Xizeng; Zhou, Chuan; Liu, Bingqiang; Chen, Xin; Xu, Ying

2014-07-01

DMINDA (DNA motif identification and analyses) is an integrated web server for DNA motif identification and analyses, which is accessible at http://csbl.bmb.uga.edu/DMINDA/. This web site is freely available to all users and there is no login requirement. This server provides a suite of cis-regulatory motif analysis functions on DNA sequences, which are important to elucidation of the mechanisms of transcriptional regulation: (i) de novo motif finding for a given set of promoter sequences along with statistical scores for the predicted motifs derived based on information extracted from a control set, (ii) scanning motif instances of a query motif in provided genomic sequences, (iii) motif comparison and clustering of identified motifs, and (iv) co-occurrence analyses of query motifs in given promoter sequences. The server is powered by a backend computer cluster with over 150 computing nodes, and is particularly useful for motif prediction and analyses in prokaryotic genomes. We believe that DMINDA, as a new and comprehensive web server for cis-regulatory motif finding and analyses, will benefit the genomic research community in general and prokaryotic genome researchers in particular. © The Author(s) 2014. Published by Oxford University Press on behalf of Nucleic Acids Research.

DNA asymmetry in stem cells - immortal or mortal?

PubMed

Yadlapalli, Swathi; Yamashita, Yukiko M

2013-09-15

The immortal strand hypothesis proposes that stem cells retain a template copy of genomic DNA (i.e. an 'immortal strand') to avoid replication-induced mutations. An alternative hypothesis suggests that certain cells segregate sister chromatids non-randomly to transmit distinct epigenetic information. However, this area of research has been highly controversial, with conflicting data even from the same cell types. Moreover, historically, the same term of 'non-random sister chromatid segregation' or 'biased sister chromatid segregation' has been used to indicate distinct biological processes, generating a confusion in the biological significance and potential mechanism of each phenomenon. Here, we discuss the models of non-random sister chromatid segregation, and we explore the strengths and limitations of the various techniques and experimental model systems used to study this question. We also describe our recent study on Drosophila male germline stem cells, where sister chromatids of X and Y chromosomes are segregated non-randomly during cell division. We aim to integrate the existing evidence to speculate on the underlying mechanisms and biological relevance of this long-standing observation on non-random sister chromatid segregation.

DNA–DNA kissing complexes as a new tool for the assembly of DNA nanostructures

PubMed Central

Barth, Anna; Kobbe, Daniela; Focke, Manfred

2016-01-01

Kissing-loop annealing of nucleic acids occurs in nature in several viruses and in prokaryotic replication, among other circumstances. Nucleobases of two nucleic acid strands (loops) interact with each other, although the two strands cannot wrap around each other completely because of the adjacent double-stranded regions (stems). In this study, we exploited DNA kissing-loop interaction for nanotechnological application. We functionalized the vertices of DNA tetrahedrons with DNA stem-loop sequences. The complementary loop sequence design allowed the hybridization of different tetrahedrons via kissing-loop interaction, which might be further exploited for nanotechnology applications like cargo transport and logical elements. Importantly, we were able to manipulate the stability of those kissing-loop complexes based on the choice and concentration of cations, the temperature and the number of complementary loops per tetrahedron either at the same or at different vertices. Moreover, variations in loop sequences allowed the characterization of necessary sequences within the loop as well as additional stability control of the kissing complexes. Therefore, the properties of the presented nanostructures make them an important tool for DNA nanotechnology. PMID:26773051

Mhr1p-dependent concatemeric mitochondrial DNA formation for generating yeast mitochondrial homoplasmic cells.

PubMed

Ling, Feng; Shibata, Takehiko

2004-01-01

Mitochondria carry many copies of mitochondrial DNA (mtDNA), but mt-alleles quickly segregate during mitotic growth through unknown mechanisms. Consequently, all mtDNA copies are often genetically homogeneous within each individual ("homoplasmic"). Our previous study suggested that tandem multimers ("concatemers") formed mainly by the Mhr1p (a yeast nuclear gene-encoded mtDNA-recombination protein)-dependent pathway are required for mtDNA partitioning into buds with concomitant monomerization. The transmission of a few randomly selected clones (as concatemers) of mtDNA into buds is a possible mechanism to establish homoplasmy. The current study provides evidence for this hypothesis as follows: the overexpression of MHR1 accelerates mt-allele-segregation in growing heteroplasmic zygotes, and mhr1-1 (recombination-deficient) causes its delay. The mt-allele-segregation rate correlates with the abundance of concatemers, which depends on Mhr1p. In G1-arrested cells, concatemeric mtDNA was labeled by [14C]thymidine at a much higher density than monomers, indicating concatemers as the immediate products of mtDNA replication, most likely in a rolling circle mode. After releasing the G1 arrest in the absence of [14C]thymidine, the monomers as the major species in growing buds of dividing cells bear a similar density of 14C as the concatemers in the mother cells, indicating that the concatemers in mother cells are the precursors of the monomers in buds.

Effects of antitopoisomerase drugs on chromosome recombinations and segregation in grasshopper

DOE Office of Scientific and Technical Information (OSTI.GOV)

Palitti, F.; Motta, S.; Grazioso, C.

1993-12-31

The role of different cellular functions which are required for the production of euploid cells can be studied through the use of mutants that are defective in the control of both the meiotic and mitotic cell cycle or through the use of compounds which interface with the various cellular targets which have a role in the segregation of chromosomes. The role of the achromatic part of the mitotic apparatus in the production of aneuploidy is well recognized. Substantial progress has been made in understanding the role of the chromatic part, for example, there are observations that disturbances in the normalmore » {open_quotes}metabolism{close_quotes} of the chromosomes (i.e. chromosome condensation, defective DNA repair or recombination) can affect chromosome segregation. Between the processes of both meiosis and mitosis that lead to nuclear division there are, however, important differences.« less

Residential segregation and racial disparities in self-rated health: How do dimensions of residential segregation matter?1

PubMed Central

Yang, Tse-Chuan; Zhao, Yunhan; Song, Qian

2016-01-01

Previous research on segregation and health has been criticized for overlooking the fact that segregation is a multi-dimensional concept (i.e., evenness, exposure, concentration, centralization, and clustering) and recent evidence drawn from non-black minorities challenges the conventional belief that residential segregation widens racial health disparities. Combining a survey data (n=18,752) from Philadelphia with the 2010 Census tract (n=925) data, we examine two theoretical frameworks to understand why the association of segregation with health may differ by race/ethnicity. Specifically, we investigate how each dimension of segregation contributed to racial disparities in self-rated health. We found (1) high levels of white/ black concentration could exacerbate the white/black health disparities up to 25 percent, (2) the white/Hispanic health disparities was narrowed by increasing the level of white/Hispanic centralization, and (3) no single dimension of segregation statistically outperforms others. Our findings supported that segregation is bad for blacks but may be beneficial for Hispanics. PMID:27886735

Viral Impact on Prokaryotic and Microalgal Activities in the Microphytobenthic Biofilm of an Intertidal Mudflat (French Atlantic Coast)

PubMed Central

Montanié, Hélène; De Crignis, Margot G.; Lavaud, Johann

2015-01-01

This is the first report on viriobenthos activity within the microbial biofilm located at the top-surface of the intertidal mudflat during emersion in Marennes-Oléron Bay (France). By combining in situ and ex situ approaches, the viral production (VP) was linked to the dynamics of prokaryotes and microphytobenthos (MPB). VP averaged 2–4 × 108 viruses ml−1 h−1. VP correlated positively with the Virus to Prokaryote Ratio, and both were correlated negatively with the water content. The virus-induced mortality of prokaryotes was lower in winter than in summer (6.8 vs. 39.7% of the production) and the C-shunting may supply 2–12% of their Carbon Demand, respectively. VP accounted for 79% of loss in Prokaryotes but the response was delayed compared to the increase in VP suggesting a simultaneous release of viruses of MPB origin. This hypothesis is supported by capsid-sizing of virions by transmission electronic microscopy and bioassays. Harvesting and ex situ maintenance of top-surface sediments was carried out to monitor the dynamics of viruses, prokaryotes and MPB after inoculation with benthic or planktonic viruses. Benthic viruses modified the prokaryotic and MPB dynamics and decreased the photosynthesis efficiency in contrast to planktonic viruses that impacted MPB but not the prokaryotes. PMID:26617575

[Experimental interaction of halophilic prokaryotes and opportunistic bacteria in brine].

PubMed

Selivanova, E A; Nemtseva, N V

2013-01-01

Study the effect of extremely halophilic archaea and moderately halophilic bacteria on preservation of opportunistic bacteria in brine. 17 strains of moderately halophilic bacteria and 2 strains of extremely halophilic archaea were isolated from continental hypersaline lake Razval of Sol-Iletsk area of Orenburg Region. Identification of pure cultures of prokaryotes was carried out taking into account their phenotype properties and based on determination of 16S RNA gene sequence. The effect of halophilic prokaryote on elimination of Escherichia coli from brine was evaluated during co-cultivation. Antagonistic activity of cell extracts of the studied microorganisms was evaluated by photometric method. A more prolonged preservation of an E. coli strain in brine in the presence of live cells of extremely halophilic archaea Halorubrum tebenquichense and moderately halophilic bacteria Marinococcus halophilus was established. Extracts of cells of extremely halophilic archaea and moderately halophilic bacteria on the contrary displayed antagonistic activity. The protective effect of live cells of halophilic prokaryotes and antagonistic activity of their cell extracts change the period of conservation of opportunistic bacteria in brine that regulates inter-microbial interactions and changes the period of self-purification that reflects the sanitary condition of a hypersaline water body.

Pangenomic Definition of Prokaryotic Species and the Phylogenetic Structure of Prochlorococcus spp.

PubMed

Moldovan, Mikhail A; Gelfand, Mikhail S

2018-01-01

The pangenome is the collection of all groups of orthologous genes (OGGs) from a set of genomes. We apply the pangenome analysis to propose a definition of prokaryotic species based on identification of lineage-specific gene sets. While being similar to the classical biological definition based on allele flow, it does not rely on DNA similarity levels and does not require analysis of homologous recombination. Hence this definition is relatively objective and independent of arbitrary thresholds. A systematic analysis of 110 accepted species with the largest numbers of sequenced strains yields results largely consistent with the existing nomenclature. However, it has revealed that abundant marine cyanobacteria Prochlorococcus marinus should be divided into two species. As a control we have confirmed the paraphyletic origin of Yersinia pseudotuberculosis (with embedded, monophyletic Y. pestis ) and Burkholderia pseudomallei (with B. mallei ). We also demonstrate that by our definition and in accordance with recent studies Escherichia coli and Shigella spp. are one species.

ConSpeciFix: Classifying prokaryotic species based on gene flow.

PubMed

Bobay, Louis-Marie; Ellis, Brian Shin-Hua; Ochman, Howard

2018-05-16

Classification of prokaryotic species is usually based on sequence similarity thresholds, which are easy to apply but lack a biologically-relevant foundation. Here, we present ConSpeciFix, a program that classifies prokaryotes into species using criteria set forth by the Biological Species Concept, thereby unifying species definition in all domains of life. ConSpeciFix's webserver is freely available at www.conspecifix.com. The local version of the program can be freely downloaded from https://github.com/Bobay-Ochman/ConSpeciFix. ConSpeciFix is written in Python 2.7 and requires the following dependencies: Usearch, MCL, MAFFT and RAxML. ljbobay@uncg.edu.

Macro and Microelements Drive Diversity and Composition of Prokaryotic and Fungal Communities in Hypersaline Sediments and Saline-Alkaline Soils.

PubMed

Liu, Kaihui; Ding, Xiaowei; Tang, Xiaofei; Wang, Jianjun; Li, Wenjun; Yan, Qingyun; Liu, Zhenghua

2018-01-01

Understanding the effects of environmental factors on microbial communities is critical for microbial ecology, but it remains challenging. In this study, we examined the diversity (alpha diversity) and community compositions (beta diversity) of prokaryotes and fungi in hypersaline sediments and salinized soils from northern China. Environmental variables were highly correlated, but they differed significantly between the sediments and saline soils. The compositions of prokaryotic and fungal communities in the hypersaline sediments were different from those in adjacent saline-alkaline soils, indicating a habitat-specific microbial distribution pattern. The macroelements (S, P, K, Mg, and Fe) and Ca were, respectively, correlated closely with the alpha diversity of prokaryotes and fungi, while the macronutrients (e.g., Na, S, P, and Ca) were correlated with the prokaryotic and fungal beta-diversity ( P ≤ 0.05). And, the nine microelements (e.g., Al, Ba, Co, Hg, and Mn) and micronutrients (Ba, Cd, and Sr) individually shaped the alpha diversity of prokaryotes and fungi, while the six microelements (e.g., As, Ba, Cr, and Ge) and only the trace elements (Cr and Cu), respectively, influenced the beta diversity of prokaryotes and fungi ( P < 0.05). Variation-partitioning analysis (VPA) showed that environmental variables jointly explained 55.49% and 32.27% of the total variation for the prokaryotic and fungal communities, respectively. Together, our findings demonstrate that the diversity and community composition of the prokaryotes and fungi were driven by different macro and microelements in saline habitats, and that geochemical elements could more widely regulate the diversity and community composition of prokaryotes than these of fungi.

Macro and Microelements Drive Diversity and Composition of Prokaryotic and Fungal Communities in Hypersaline Sediments and Saline–Alkaline Soils

PubMed Central

Liu, Kaihui; Ding, Xiaowei; Tang, Xiaofei; Wang, Jianjun; Li, Wenjun; Yan, Qingyun; Liu, Zhenghua

2018-01-01

Understanding the effects of environmental factors on microbial communities is critical for microbial ecology, but it remains challenging. In this study, we examined the diversity (alpha diversity) and community compositions (beta diversity) of prokaryotes and fungi in hypersaline sediments and salinized soils from northern China. Environmental variables were highly correlated, but they differed significantly between the sediments and saline soils. The compositions of prokaryotic and fungal communities in the hypersaline sediments were different from those in adjacent saline–alkaline soils, indicating a habitat-specific microbial distribution pattern. The macroelements (S, P, K, Mg, and Fe) and Ca were, respectively, correlated closely with the alpha diversity of prokaryotes and fungi, while the macronutrients (e.g., Na, S, P, and Ca) were correlated with the prokaryotic and fungal beta-diversity (P ≤ 0.05). And, the nine microelements (e.g., Al, Ba, Co, Hg, and Mn) and micronutrients (Ba, Cd, and Sr) individually shaped the alpha diversity of prokaryotes and fungi, while the six microelements (e.g., As, Ba, Cr, and Ge) and only the trace elements (Cr and Cu), respectively, influenced the beta diversity of prokaryotes and fungi (P < 0.05). Variation-partitioning analysis (VPA) showed that environmental variables jointly explained 55.49% and 32.27% of the total variation for the prokaryotic and fungal communities, respectively. Together, our findings demonstrate that the diversity and community composition of the prokaryotes and fungi were driven by different macro and microelements in saline habitats, and that geochemical elements could more widely regulate the diversity and community composition of prokaryotes than these of fungi. PMID:29535703

Are We Segregated and Satisfied? Segregation and Inequality in Southern California Schools

ERIC Educational Resources Information Center

Kucsera, John V.; Siegel-Hawley, Genevieve; Orfield, Gary

2015-01-01

Southern California is facing a demographic transformation that will become characteristic of the nation as a whole in coming decades. In this research, we present a historical review of the region's attempt to address school inequity, recent enrollment and segregation trends, and an investigation of whether segregation still matters. Our results…

GAPP: A Proteogenomic Software for Genome Annotation and Global Profiling of Post-translational Modifications in Prokaryotes.

PubMed

Zhang, Jia; Yang, Ming-Kun; Zeng, Honghui; Ge, Feng

2016-11-01

Although the number of sequenced prokaryotic genomes is growing rapidly, experimentally verified annotation of prokaryotic genome remains patchy and challenging. To facilitate genome annotation efforts for prokaryotes, we developed an open source software called GAPP for genome annotation and global profiling of post-translational modifications (PTMs) in prokaryotes. With a single command, it provides a standard workflow to validate and refine predicted genetic models and discover diverse PTM events. We demonstrated the utility of GAPP using proteomic data from Helicobacter pylori, one of the major human pathogens that is responsible for many gastric diseases. Our results confirmed 84.9% of the existing predicted H. pylori proteins, identified 20 novel protein coding genes, and corrected four existing gene models with regard to translation initiation sites. In particular, GAPP revealed a large repertoire of PTMs using the same proteomic data and provided a rich resource that can be used to examine the functions of reversible modifications in this human pathogen. This software is a powerful tool for genome annotation and global discovery of PTMs and is applicable to any sequenced prokaryotic organism; we expect that it will become an integral part of ongoing genome annotation efforts for prokaryotes. GAPP is freely available at https://sourceforge.net/projects/gappproteogenomic/. © 2016 by The American Society for Biochemistry and Molecular Biology, Inc.

Bacterial identification and subtyping using DNA microarray and DNA sequencing.

PubMed

Al-Khaldi, Sufian F; Mossoba, Magdi M; Allard, Marc M; Lienau, E Kurt; Brown, Eric D

2012-01-01

The era of fast and accurate discovery of biological sequence motifs in prokaryotic and eukaryotic cells is here. The co-evolution of direct genome sequencing and DNA microarray strategies not only will identify, isotype, and serotype pathogenic bacteria, but also it will aid in the discovery of new gene functions by detecting gene expressions in different diseases and environmental conditions. Microarray bacterial identification has made great advances in working with pure and mixed bacterial samples. The technological advances have moved beyond bacterial gene expression to include bacterial identification and isotyping. Application of new tools such as mid-infrared chemical imaging improves detection of hybridization in DNA microarrays. The research in this field is promising and future work will reveal the potential of infrared technology in bacterial identification. On the other hand, DNA sequencing by using 454 pyrosequencing is so cost effective that the promise of $1,000 per bacterial genome sequence is becoming a reality. Pyrosequencing technology is a simple to use technique that can produce accurate and quantitative analysis of DNA sequences with a great speed. The deposition of massive amounts of bacterial genomic information in databanks is creating fingerprint phylogenetic analysis that will ultimately replace several technologies such as Pulsed Field Gel Electrophoresis. In this chapter, we will review (1) the use of DNA microarray using fluorescence and infrared imaging detection for identification of pathogenic bacteria, and (2) use of pyrosequencing in DNA cluster analysis to fingerprint bacterial phylogenetic trees.

Segregation effects during solidification in weightless melts

NASA Technical Reports Server (NTRS)

Li, C.

1973-01-01

Two types of melt segregation effects were studied: (1) evaporative segregation, or segregation due to surface evaporation; and (2) freezing segregation, or segregation due to liquid-solid phase transformation. These segregation effects are closely related. In fact, evaporative segregation always precedes freezing segregation to some degree and must often be studied prior to performing meaningful solidification experiments. This is particularly true since evaporation may cause the melt composition, at least at the critical surface regions or layers to be affected manyfold within seconds so that the surface region or layer melting point and other thermophysical properties, nucleation characteristics, base for undercooling, and critical velocity to avoid constitutional supercooling, may be completely unexpected. An important objective was, therefore, to develop the necessary normal evaporation equations for predicting the compositional changes within specified times at temperature and to correlate these equations with actual experimental data collected from the literature.

Connectivity between surface and deep waters determines prokaryotic diversity in the North Atlantic Deep Water

PubMed Central

Frank, Alexander H.; Garcia, Juan A. L.; Herndl, Gerhard J.

2016-01-01

Summary To decipher the influence of depth stratification and surface provincialism on the dark ocean prokaryotic community composition, we sampled the major deep‐water masses in the eastern North Atlantic covering three biogeographic provinces. Their diversity was evaluated using ordination and canonical analysis of 454 pyrotag sequences. Variance partitioning suggested that 16% of the variation in the bacterial community composition was based on depth stratification while 9% of the variation was due to geographic location. General linear mixed effect models showed that the community of the subsurface waters was connected to the dark ocean prokaryotic communities in different biogeographic provinces. Cluster analysis indicated that some prokaryotic taxa are specific to distinct regions in bathypelagic water masses. Taken together, our data suggest that the dark ocean prokaryotic community composition of the eastern North Atlantic is primed by the formation and the horizontal transport of water masses. PMID:26914787

Dynamics of natural prokaryotes, viruses, and heterotrophic nanoflagellates in alpine karstic groundwater

PubMed Central

Wilhartitz, Inés C; Kirschner, Alexander K T; Brussaard, Corina P D; Fischer, Ulrike R; Wieltschnig, Claudia; Stadler, Hermann; Farnleitner, Andreas H

2013-01-01

Abstract Seasonal dynamics of naturally occurring prokaryotes, viruses, and heterotrophic nanoflagellates in two hydro-geologically contrasting alpine karst springs were monitored over three annual cycles. To our knowledge, this study is the first to shed light on the occurrence and possible interrelationships between these three groups in karstic groundwater. Hydrological and microbiological standard indicators were recovered simultaneously in order to estimate surface influence, especially during rainfall events. Data revealed a strong dependence of the microbial communities on the prevailing hydrological situation. Prokaryotic numbers averaged 5.1 × 107 and 1.3 × 107 cells L−1, and heterotrophic nanoflagellate abundance averaged 1.1 × 104 and 3 × 103 cells L−1 in the limestone spring type (LKAS2) and the dolomitic spring type (DKAS1), respectively. Viral abundance in LKAS2 and DKAS1 averaged 9.4 × 108 and 1.1 × 108 viruses L−1. Unlike in DKAS1, the dynamic spring type LKAS2 revealed a clear difference between base flow and high discharge conditions. The virus-to-prokaryotes ratio was generally lower by a factor of 2–3, at higher average water residence times. Furthermore, the high prokaryotes-to-heterotrophic nanoflagellate ratios, namely about 4700 and 5400 for LKAS2 and DKAS1, respectively, pointed toward an uncoupling of these two groups in the planktonic fraction of alpine karstic aquifers. Seasonal dynamics of naturally occurring prokaryotes, viruses and heterotrophic nanoflagellates in two hydro-geologically contrasting alpine karst springs were monitored over three annual cycles. Data revealed a strong dependence of the microbial communities on the prevailing hydrological situation. PMID:23828838

Predatory prokaryotes: predation and primary consumption evolved in bacteria

NASA Technical Reports Server (NTRS)

Guerrero, R.; Pedros-Alio, C.; Esteve, I.; Mas, J.; Chase, D.; Margulis, L.

1986-01-01

Two kinds of predatory bacteria have been observed and characterized by light and electron microscopy in samples from freshwater sulfurous lakes in northeastern Spain. The first bacterium, named Vampirococcus, is Gram-negative and ovoidal (0.6 micrometer wide). An anaerobic epibiont, it adheres to the surface of phototrophic bacteria (Chromatium spp.) by specific attachment structures and, as it grows and divides by fission, destroys its prey. An important in situ predatory role can be inferred for Vampirococcus from direct counts in natural samples. The second bacterium, named Daptobacter, is a Gram-negative, facultatively anaerobic straight rod (0.5 x 1.5 micrometers) with a single polar flagellum, which collides, penetrates, and grows inside the cytoplasm of its prey (several genera of Chromatiaceae). Considering also the well-known case of Bdellovibrio, a Gram-negative, aerobic curved rod that penetrates and divides in the periplasmic space of many chemotrophic Gram-negative bacteria, there are three types of predatory prokaryotes presently known (epibiotic, cytoplasmic, and periplasmic). Thus, we conclude that antagonistic relationships such as primary consumption, predation, and scavenging had already evolved in microbial ecosystems prior to the appearance of eukaryotes. Furthermore, because they represent methods by which prokaryotes can penetrate other prokaryotes in the absence of phagocytosis, these associations can be considered preadaptation for the origin of intracellular organelles.

The use of an optical method to evaluate prokaryotic oxygen consumption under high pressure condition

NASA Astrophysics Data System (ADS)

Garel, M.; Martini, S.; Lefèvre, D.; Tamburini, C.

2016-02-01

The heterotrophic prokaryotes are the main contributor to organic matter degradation in the ocean and particularly in the deep ocean. Nowadays, a classical way to evaluate the prokaryotic carbon demand (PCD) needs the estimation of both prokaryotic heterotrophic production (PHP) and prokaryotic respiration (PR). PHP measurements in deep-sea waters are relatively well documented and the importance of maintaining the in situ conditions (pressure and temperature) to avoid bias of the real deep-sea activities has been highlighted. However, no accurate methodology is available to measure directly, under in situ conditions (pressure and temperature) PR in the dark ocean. This study is presenting PR measurements under in situ conditions. High-pressure bottles have been adapted with a non-invasive sensor to measure prokaryotic oxygen consumption. The methodology is based on fluorescence quenching where molecular oxygen quenches the luminescence of planar-optode-oxygen sensor widely used in oceanography. Firstly, accuracy, detection limit, precision and response time of oxygen concentration measurements have been investigated in relation to an increase of hydrostatic pressure. Secondly, we will present experiments performed on natural prokaryotic consortium mixed with freshly collected particles to assess the O2 consumption in relation with increasing hydrostatic pressure (150 m depth per day). Finally, first results of coupled PHP and PR measurements at in situ conditions (temperature and pressure) from mesopelagic and bathypelagic samples of the Atlantic Ocean (PAP site), will be discussed. Finally, we will discuss first results of coupled PHP and PR measurements at in situ conditions (temperature and pressure) from Atlantic Ocean mesopelagic and bathypelagic samples (PAP site).

Quantification of Viral and Prokaryotic Production Rates in Benthic Ecosystems: A Methods Comparison

PubMed Central

Rastelli, Eugenio; Dell’Anno, Antonio; Corinaldesi, Cinzia; Middelboe, Mathias; Noble, Rachel T.; Danovaro, Roberto

2016-01-01

Viruses profoundly influence benthic marine ecosystems by infecting and subsequently killing their prokaryotic hosts, thereby impacting the cycling of carbon and nutrients. Previously conducted studies, based on different methodologies, have provided widely differing estimates of the relevance of viruses on benthic prokaryotes. There has been no attempt so far to compare these independent approaches, including contextual comparisons among different approaches for sample manipulation (i.e., dilution or not of the sediments during incubations), between methods based on epifluorescence microscopy (EFM) or radiotracers, and between the use of different radiotracers. Therefore, it has been difficult to identify the most suitable methodologies and protocols to be used as standard approaches for the quantification of viral infections of prokaryotes. Here, we compared for the first time different methods for determining viral and prokaryotic production rates in marine sediments collected at two benthic sites, differing in depth and environmental conditions. We used a highly replicated experimental design, testing the potential biases associated to the incubation of sediments as diluted or undiluted. In parallel, we also compared EFM counts with the 3H-thymidine incubations for the determination of viral production rates, and the use of 3H-thymidine versus 3H-leucine radiotracers for the determination of prokaryotic production. We show here that, independent from sediment dilution, EFM-based values of viral production ranged from 1.4 to 4.6 × 107 viruses g-1 h-1, and were similar but overall less variable compared to those obtained by the 3H-thymidine method (0.3 to 9.0 × 107 viruses g-1h-1). In addition, the prokaryotic production rates were not affected by sediment dilution, and the use of different radiotracers provided very consistent estimates (10.3–35.1 and 9.3–34.6 ngC g-1h-1 using the 3H-thymidine or 3H-leucine method, respectively). These results indicated

Pyrosequencing assessment of prokaryotic and eukaryotic diversity in biofilm communities from a French river.

PubMed

Bricheux, Geneviève; Morin, Loïc; Le Moal, Gwenaël; Coffe, Gérard; Balestrino, Damien; Charbonnel, Nicolas; Bohatier, Jacques; Forestier, Christiane

2013-06-01

Despite the recent and significant increase in the study of aquatic microbial communities, little is known about the microbial diversity of complex ecosystems such as running waters. This study investigated the biodiversity of biofilm communities formed in a river with 454 Sequencing™. This river has the particularity of integrating both organic and microbiological pollution, as receiver of agricultural pollution in its upstream catchment area and urban pollution through discharges of the wastewater treatment plant of the town of Billom. Different regions of the small subunit (SSU) ribosomal RNA gene were targeted using nine pairs of primers, either universal or specific for bacteria, eukarya, or archaea. Our aim was to characterize the widest range of rDNA sequences using different sets of polymerase chain reaction (PCR) primers. A first look at reads abundance revealed that a large majority (47-48%) were rare sequences (<5 copies). Prokaryotic phyla represented the species richness, and eukaryotic phyla accounted for a small part. Among the prokaryotic phyla, Proteobacteria (beta and alpha) predominated, followed by Bacteroidetes together with a large number of nonaffiliated bacterial sequences. Bacillariophyta plastids were abundant. The remaining bacterial phyla, Verrucomicrobia and Cyanobacteria, made up the rest of the bulk biodiversity. The most abundant eukaryotic phyla were annelid worms, followed by Diatoms, and Chlorophytes. These latter phyla attest to the abundance of plastids and the importance of photosynthetic activity for the biofilm. These findings highlight the existence and plasticity of multiple trophic levels within these complex biological systems. © 2013 The Authors. Microbiology Open published by John Wiley & Sons Ltd.

Pyrosequencing assessment of prokaryotic and eukaryotic diversity in biofilm communities from a French river

PubMed Central

Bricheux, Geneviève; Morin, Loïc; Le Moal, Gwenaël; Coffe, Gérard; Balestrino, Damien; Charbonnel, Nicolas; Bohatier, Jacques; Forestier, Christiane

2013-01-01

Despite the recent and significant increase in the study of aquatic microbial communities, little is known about the microbial diversity of complex ecosystems such as running waters. This study investigated the biodiversity of biofilm communities formed in a river with 454 Sequencing™. This river has the particularity of integrating both organic and microbiological pollution, as receiver of agricultural pollution in its upstream catchment area and urban pollution through discharges of the wastewater treatment plant of the town of Billom. Different regions of the small subunit (SSU) ribosomal RNA gene were targeted using nine pairs of primers, either universal or specific for bacteria, eukarya, or archaea. Our aim was to characterize the widest range of rDNA sequences using different sets of polymerase chain reaction (PCR) primers. A first look at reads abundance revealed that a large majority (47–48%) were rare sequences (<5 copies). Prokaryotic phyla represented the species richness, and eukaryotic phyla accounted for a small part. Among the prokaryotic phyla, Proteobacteria (beta and alpha) predominated, followed by Bacteroidetes together with a large number of nonaffiliated bacterial sequences. Bacillariophyta plastids were abundant. The remaining bacterial phyla, Verrucomicrobia and Cyanobacteria, made up the rest of the bulk biodiversity. The most abundant eukaryotic phyla were annelid worms, followed by Diatoms, and Chlorophytes. These latter phyla attest to the abundance of plastids and the importance of photosynthetic activity for the biofilm. These findings highlight the existence and plasticity of multiple trophic levels within these complex biological systems. PMID:23520129

OperomeDB: A Database of Condition-Specific Transcription Units in Prokaryotic Genomes.

PubMed

Chetal, Kashish; Janga, Sarath Chandra

2015-01-01

Background. In prokaryotic organisms, a substantial fraction of adjacent genes are organized into operons-codirectionally organized genes in prokaryotic genomes with the presence of a common promoter and terminator. Although several available operon databases provide information with varying levels of reliability, very few resources provide experimentally supported results. Therefore, we believe that the biological community could benefit from having a new operon prediction database with operons predicted using next-generation RNA-seq datasets. Description. We present operomeDB, a database which provides an ensemble of all the predicted operons for bacterial genomes using available RNA-sequencing datasets across a wide range of experimental conditions. Although several studies have recently confirmed that prokaryotic operon structure is dynamic with significant alterations across environmental and experimental conditions, there are no comprehensive databases for studying such variations across prokaryotic transcriptomes. Currently our database contains nine bacterial organisms and 168 transcriptomes for which we predicted operons. User interface is simple and easy to use, in terms of visualization, downloading, and querying of data. In addition, because of its ability to load custom datasets, users can also compare their datasets with publicly available transcriptomic data of an organism. Conclusion. OperomeDB as a database should not only aid experimental groups working on transcriptome analysis of specific organisms but also enable studies related to computational and comparative operomics.

The Universally Conserved Prokaryotic GTPases

PubMed Central

Verstraeten, Natalie; Fauvart, Maarten; Versées, Wim; Michiels, Jan

2011-01-01

Summary: Members of the large superclass of P-loop GTPases share a core domain with a conserved three-dimensional structure. In eukaryotes, these proteins are implicated in various crucial cellular processes, including translation, membrane trafficking, cell cycle progression, and membrane signaling. As targets of mutation and toxins, GTPases are involved in the pathogenesis of cancer and infectious diseases. In prokaryotes also, it is hard to overestimate the importance of GTPases in cell physiology. Numerous papers have shed new light on the role of bacterial GTPases in cell cycle regulation, ribosome assembly, the stress response, and other cellular processes. Moreover, bacterial GTPases have been identified as high-potential drug targets. A key paper published over 2 decades ago stated that, “It may never again be possible to capture [GTPases] in a family portrait” (H. R. Bourne, D. A. Sanders, and F. McCormick, Nature 348:125-132, 1990) and indeed, the last 20 years have seen a tremendous increase in publications on the subject. Sequence analysis identified 13 bacterial GTPases that are conserved in at least 75% of all bacterial species. We here provide an overview of these 13 protein subfamilies, covering their cellular functions as well as cellular localization and expression levels, three-dimensional structures, biochemical properties, and gene organization. Conserved roles in eukaryotic homologs will be discussed as well. A comprehensive overview summarizing current knowledge on prokaryotic GTPases will aid in further elucidating the function of these important proteins. PMID:21885683

Surveying the expanding prokaryotic Rubisco multiverse.

PubMed

Liu, Di; Ramya, Ramaswamy Chettiyan Seetharaman; Mueller-Cajar, Oliver

2017-09-01

The universal, but catalytically modest, CO2-fixing enzyme Rubisco is currently experiencing intense interest by researchers aiming to enhance crop photosynthesis. These efforts are mostly focused on the highly conserved hexadecameric enzyme found in land plants. In comparison, prokaryotic organisms harbor a far greater diversity in Rubisco forms. Recent work towards improving our appreciation of microbial Rubisco properties and harnessing their potential is surveyed. New structural models are providing informative glimpses into catalytic subtleties and diverse oligomeric states. Ongoing characterization is informing us about the conservation of constraints, such as sugar phosphate inhibition and the associated dependence on Rubisco activase helper proteins. Prokaryotic Rubiscos operate under a far wider range of metabolic contexts than the photosynthetic function of higher plant enzymes. Relaxed selection pressures may have resulted in the exploration of a larger volume of sequence space than permitted in organisms performing oxygenic photosynthesis. To tap into the potential of microbial Rubiscos, in vivo selection systems are being used to discover functional metagenomic Rubiscos. Various directed evolution systems to optimize their function have been developed. It is anticipated that this approach will provide access to biotechnologically valuable enzymes that cannot be encountered in the higher plant Rubisco space. © FEMS 2017. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

Macroevolutionary trends of atomic composition and related functional group proportion in eukaryotic and prokaryotic proteins.

PubMed

Zhang, Yu-Juan; Yang, Chun-Lin; Hao, You-Jin; Li, Ying; Chen, Bin; Wen, Jian-Fan

2014-01-25

To fully explore the trends of atomic composition during the macroevolution from prokaryote to eukaryote, five atoms (oxygen, sulfur, nitrogen, carbon, hydrogen) and related functional groups in prokaryotic and eukaryotic proteins were surveyed and compared. Genome-wide analysis showed that eukaryotic proteins have more oxygen, sulfur and nitrogen atoms than prokaryotes do. Clusters of Orthologous Groups (COG) analysis revealed that oxygen, sulfur, carbon and hydrogen frequencies are higher in eukaryotic proteins than in their prokaryotic orthologs. Furthermore, functional group analysis demonstrated that eukaryotic proteins tend to have higher proportions of sulfhydryl, hydroxyl and acylamino, but lower of sulfide and carboxyl. Taken together, an apparent trend of increase was observed for oxygen and sulfur atoms in the macroevolution; the variation of oxygen and sulfur compositions and their related functional groups in macroevolution made eukaryotic proteins carry more useful functional groups. These results will be helpful for better understanding the functional significances of atomic composition evolution. Copyright © 2013 Elsevier B.V. All rights reserved.

Connectivity between surface and deep waters determines prokaryotic diversity in the North Atlantic Deep Water.

PubMed

Frank, Alexander H; Garcia, Juan A L; Herndl, Gerhard J; Reinthaler, Thomas

2016-06-01

To decipher the influence of depth stratification and surface provincialism on the dark ocean prokaryotic community composition, we sampled the major deep-water masses in the eastern North Atlantic covering three biogeographic provinces. Their diversity was evaluated using ordination and canonical analysis of 454 pyrotag sequences. Variance partitioning suggested that 16% of the variation in the bacterial community composition was based on depth stratification while 9% of the variation was due to geographic location. General linear mixed effect models showed that the community of the subsurface waters was connected to the dark ocean prokaryotic communities in different biogeographic provinces. Cluster analysis indicated that some prokaryotic taxa are specific to distinct regions in bathypelagic water masses. Taken together, our data suggest that the dark ocean prokaryotic community composition of the eastern North Atlantic is primed by the formation and the horizontal transport of water masses. © 2016 Society for Applied Microbiology and John Wiley & Sons Ltd.

18 CFR 401.113 - Segregable materials.

Code of Federal Regulations, 2010 CFR

2010-04-01

... 18 Conservation of Power and Water Resources 2 2010-04-01 2010-04-01 false Segregable materials. 401.113 Section 401.113 Conservation of Power and Water Resources DELAWARE RIVER BASIN COMMISSION... Segregable materials. Any reasonably segregable portion of a record shall be provided to any person...

18 CFR 401.113 - Segregable materials.

Code of Federal Regulations, 2013 CFR

2013-04-01

... 18 Conservation of Power and Water Resources 2 2013-04-01 2012-04-01 true Segregable materials. 401.113 Section 401.113 Conservation of Power and Water Resources DELAWARE RIVER BASIN COMMISSION... Segregable materials. Any reasonably segregable portion of a record shall be provided to any person...

18 CFR 401.113 - Segregable materials.

Code of Federal Regulations, 2014 CFR

2014-04-01

... 18 Conservation of Power and Water Resources 2 2014-04-01 2014-04-01 false Segregable materials. 401.113 Section 401.113 Conservation of Power and Water Resources DELAWARE RIVER BASIN COMMISSION... Segregable materials. Any reasonably segregable portion of a record shall be provided to any person...

18 CFR 401.113 - Segregable materials.

Code of Federal Regulations, 2012 CFR

2012-04-01

... 18 Conservation of Power and Water Resources 2 2012-04-01 2012-04-01 false Segregable materials. 401.113 Section 401.113 Conservation of Power and Water Resources DELAWARE RIVER BASIN COMMISSION... Segregable materials. Any reasonably segregable portion of a record shall be provided to any person...

49 CFR 176.708 - Segregation distances.

Code of Federal Regulations, 2011 CFR

2011-10-01

... 49 Transportation 2 2011-10-01 2011-10-01 false Segregation distances. 176.708 Section 176.708... Requirements for Radioactive Materials § 176.708 Segregation distances. (a) Table IV lists minimum separation... into account any relocation of cargo during the voyage. (e) Any departure from the segregation...

Characterization of Ancient DNA Supports Long-Term Survival of Haloarchaea

PubMed Central

Lowenstein, Tim K.; Timofeeff, Michael N.; Schubert, Brian A.; Lum, J. Koji

2014-01-01

Abstract Bacteria and archaea isolated from crystals of halite 104 to 108 years old suggest long-term survival of halophilic microorganisms, but the results are controversial. Independent verification of the authenticity of reputed living prokaryotes in ancient salt is required because of the high potential for environmental and laboratory contamination. Low success rates of prokaryote cultivation from ancient halite, however, hamper direct replication experiments. In such cases, culture-independent approaches that use the polymerase chain reaction (PCR) and sequencing of 16S ribosomal DNA are a robust alternative. Here, we use amplification, cloning, and sequencing of 16S ribosomal DNA to investigate the authenticity of halophilic archaea cultured from subsurface halite, Death Valley, California, 22,000 to 34,000 years old. We recovered 16S ribosomal DNA sequences that are identical, or nearly so (>99%), to two strains, Natronomonas DV462A and Halorubrum DV427, which were previously isolated from the same halite interval. These results provide the best independent support to date for the long-term survival of halophilic archaea in ancient halite. PCR-based approaches are sensitive to small amounts of DNA and could allow investigation of even older halites, 106 to 108 years old, from which microbial cultures have been reported. Such studies of microbial life in ancient salt are particularly important as we search for microbial signatures in similar deposits on Mars and elsewhere in the Solar System. Key Words: Ancient DNA—Halite—Haloarchaea—Long-term survival. Astrobiology 14, 553–560. PMID:24977469

Deep sequencing approaches for the analysis of prokaryotic transcriptional boundaries and dynamics.

PubMed

James, Katherine; Cockell, Simon J; Zenkin, Nikolay

2017-05-01

The identification of the protein-coding regions of a genome is straightforward due to the universality of start and stop codons. However, the boundaries of the transcribed regions, conditional operon structures, non-coding RNAs and the dynamics of transcription, such as pausing of elongation, are non-trivial to identify, even in the comparatively simple genomes of prokaryotes. Traditional methods for the study of these areas, such as tiling arrays, are noisy, labour-intensive and lack the resolution required for densely-packed bacterial genomes. Recently, deep sequencing has become increasingly popular for the study of the transcriptome due to its lower costs, higher accuracy and single nucleotide resolution. These methods have revolutionised our understanding of prokaryotic transcriptional dynamics. Here, we review the deep sequencing and data analysis techniques that are available for the study of transcription in prokaryotes, and discuss the bioinformatic considerations of these analyses. Copyright © 2017 Elsevier Inc. All rights reserved.

[Prokaryotic expression, purification and biological activity analysis of recombinant β-Lactamase protein].

PubMed

Zhou, Xiao-liang; Shi, Pei-ji; Wang, Hao

2011-01-01

To prepare RGD4CβL fusion protein using prokaryotic expression system and evaluate the biological activity of the RGD4CβL. RGD4CβL gene was cloned into pColdII to contruct β-Lactamase prokaryotic expression vector. After transformation, the recombinant vector was induced to express recombinant protein RGD4CβL by IPTG in E.coli BL(DE3). The recombinant protein was purified by Ni-NTA resin under denaturing condition and then dialyzed to renature. The tumor cell targeting ability of the recombinant protein was analyzed by flow cytometric analysis. After cleavage and purification, β-Lactamase moiety showed the expected size of 42 000 on Tricine-SDS-PAGE, and was further confirmed by Western blotting. Based on flow cytometric analysis, the purified protein specially targeted breast cancer cell line MCF-7. This research successfully estiblished a method for prokaryotic expression and purification of β-lactamase. These results suggest the potential use of the protein as an agent for ADEPT.

Variable viral and grazer control of prokaryotic growth efficiency in temperate freshwater lakes (French Massif Central).

PubMed

Ram, A S Pradeep; Palesse, S; Colombet, J; Sabart, M; Perriere, F; Sime-Ngando, T

2013-11-01

The effects of viral lysis and heterotrophic nanoflagellate grazing (top down forces) on prokaryotic mortality and their subsequent impact on their metabolism were estimated in the upper euphotic and deeper aphotic depth of 11 freshwater lakes located in the French Massif Central. The standing stocks of viruses (VA) and heterotrophic nanoflagellate (HNF) varied significantly (p < 0.05) with sampled depth. VA was substantially (twofold on an average) and significantly higher (p < 0.03) at the aphotic compared to euphotic depth, whereas the reverse was true (p < 0.02) for HNF. Among the prokaryote subgroup, high nucleic acid content prokaryotes explained for significant variability in the total VA and served as principle host target for viral proliferation. Like standing stocks, flagellate grazing and viral infection rates also followed similar patterns. In the investigated lakes, the mechanism for regulating prokaryotic production varied with sampled depth from grazing control in the euphotic to control due to viral lysis in the aphotic. We also tested the hypothesis of top down control on prokaryotic growth efficiency (PGE, which we used as an index of prokaryotic physiological and energetic status at the community level) at both depths. Overall, among the studied lakes, PGE varied widely (4-51 %) with significantly (p < 0.05) lower values in the aphotic (mean = 18 ± 4 %) than euphotic depth (mean = 32 ± 9 %). Contrasting observations on the top down control of PGE between sampled depths were observed. The presence of grazers was found to stimulate PGE at the euphotic, whereas viruses through their lytic infection had a strong negative impact on PGE at the aphotic depth. Such observed differences in PGE and the mechanism controlling prokaryotic production with depth could eventually have strong implication on carbon and nutrient flux patterns in the studied lakes.

Income Segregation between Schools and School Districts

ERIC Educational Resources Information Center

Owens, Ann; Reardon, Sean F.; Jencks, Christopher

2016-01-01

Although trends in the racial segregation of schools are well documented, less is known about trends in income segregation. We use multiple data sources to document trends in income segregation between schools and school districts. Between-district income segregation of families with children enrolled in public school increased by over 15% from…

Linear and Nonlinear Statistical Characterization of DNA

NASA Astrophysics Data System (ADS)

Norio Oiwa, Nestor; Goldman, Carla; Glazier, James

2002-03-01

We find spatial order in the distribution of protein-coding (including RNAs) and control segments of GenBank genomic sequences, irrespective of ATCG content. This is achieved by correlations, histograms, fractal dimensions and singularity spectra. Estimates of these quantities in complete nuclear genome indicate that coding sequences are long-range correlated and their disposition are self-similar (multifractal) for eukaryotes. These characteristics are absent in prokaryotes, where there are few noncoding sequences, suggesting the `junk' DNA play a relevant role to the genome structure and function. Concerning the genetic message of ATCG sequences, we build a random walk (Levy flight), using DNA symmetry arguments, where we associate A, T, C and G as left, right, down and up steps, respectively. Nonlinear analysis of mitochondrial DNA walks reveal multifractal pattern based on palindromic sequences, which fold in hairpins and loops.

Mgm101p Is a Novel Component of the Mitochondrial Nucleoid That Binds DNA and Is Required for the Repair of Oxidatively Damaged Mitochondrial DNA

PubMed Central

Meeusen, Shelly; Tieu, Quinton; Wong, Edith; Weiss, Eric; Schieltz, David; Yates, John R.; Nunnari, Jodi

1999-01-01

Maintenance of mitochondrial DNA (mtDNA) during cell division is required for progeny to be respiratory competent. Maintenance involves the replication, repair, assembly, segregation, and partitioning of the mitochondrial nucleoid. MGM101 has been identified as a gene essential for mtDNA maintenance in S. cerevisiae, but its role is unknown. Using liquid chromatography coupled with tandem mass spectrometry, we identified Mgm101p as a component of highly enriched nucleoids, suggesting that it plays a nucleoid-specific role in maintenance. Subcellular fractionation, indirect immunofluorescence and GFP tagging show that Mgm101p is exclusively associated with the mitochondrial nucleoid structure in cells. Furthermore, DNA affinity chromatography of nucleoid extracts indicates that Mgm101p binds to DNA, suggesting that its nucleoid localization is in part due to this activity. Phenotypic analysis of cells containing a temperature sensitive mgm101 allele suggests that Mgm101p is not involved in mtDNA packaging, segregation, partitioning or required for ongoing mtDNA replication. We examined Mgm101p's role in mtDNA repair. As compared with wild-type cells, mgm101 cells were more sensitive to mtDNA damage induced by UV irradiation and were hypersensitive to mtDNA damage induced by gamma rays and H2O2 treatment. Thus, we propose that Mgm101p performs an essential function in the repair of oxidatively damaged mtDNA that is required for the maintenance of the mitochondrial genome. PMID:10209025

Proposal to include the rank of phylum in the International Code of Nomenclature of Prokaryotes.

PubMed

Oren, Aharon; da Costa, Milton S; Garrity, George M; Rainey, Fred A; Rosselló-Móra, Ramon; Schink, Bernhard; Sutcliffe, Iain; Trujillo, Martha E; Whitman, William B

2015-11-01

The International Code of Nomenclature of Prokaryotes covers the nomenclature of prokaryotes up to the rank of class. We propose here modifying the Code to include the rank of phylum so that names of phyla that fulfil the rules of the Code will obtain standing in the nomenclature.

Computing Prediction and Functional Analysis of Prokaryotic Propionylation.

PubMed

Wang, Li-Na; Shi, Shao-Ping; Wen, Ping-Ping; Zhou, Zhi-You; Qiu, Jian-Ding

2017-11-27

Identification and systematic analysis of candidates for protein propionylation are crucial steps for understanding its molecular mechanisms and biological functions. Although several proteome-scale methods have been performed to delineate potential propionylated proteins, the majority of lysine-propionylated substrates and their role in pathological physiology still remain largely unknown. By gathering various databases and literatures, experimental prokaryotic propionylation data were collated to be trained in a support vector machine with various features via a three-step feature selection method. A novel online tool for seeking potential lysine-propionylated sites (PropSeek) ( http://bioinfo.ncu.edu.cn/PropSeek.aspx ) was built. Independent test results of leave-one-out and n-fold cross-validation were similar to each other, showing that PropSeek is a stable and robust predictor with satisfying performance. Meanwhile, analyses of Gene Ontology, Kyoto Encyclopedia of Genes and Genomes pathways, and protein-protein interactions implied a potential role of prokaryotic propionylation in protein synthesis and metabolism.

Rule Mining Techniques to Predict Prokaryotic Metabolic Pathways.

PubMed

Saidi, Rabie; Boudellioua, Imane; Martin, Maria J; Solovyev, Victor

2017-01-01

It is becoming more evident that computational methods are needed for the identification and the mapping of pathways in new genomes. We introduce an automatic annotation system (ARBA4Path Association Rule-Based Annotator for Pathways) that utilizes rule mining techniques to predict metabolic pathways across wide range of prokaryotes. It was demonstrated that specific combinations of protein domains (recorded in our rules) strongly determine pathways in which proteins are involved and thus provide information that let us very accurately assign pathway membership (with precision of 0.999 and recall of 0.966) to proteins of a given prokaryotic taxon. Our system can be used to enhance the quality of automatically generated annotations as well as annotating proteins with unknown function. The prediction models are represented in the form of human-readable rules, and they can be used effectively to add absent pathway information to many proteins in UniProtKB/TrEMBL database.

A Revaluation of Indexes of Residential Segregation

ERIC Educational Resources Information Center

Winship, Christopher

1977-01-01

Shows that there are at least two different perspectives from which residential segregation can be examined. Segregation can be measured as it deviates from a situation of complete desegregation or in terms of a situation in which there is random segregation in the city. New criteria for indexes of residential segregation are developed. (Author/JM)

Genomics of bacteria and archaea: the emerging dynamic view of the prokaryotic world

PubMed Central

Koonin, Eugene V.; Wolf, Yuri I.

2008-01-01

The first bacterial genome was sequenced in 1995, and the first archaeal genome in 1996. Soon after these breakthroughs, an exponential rate of genome sequencing was established, with a doubling time of approximately 20 months for bacteria and approximately 34 months for archaea. Comparative analysis of the hundreds of sequenced bacterial and dozens of archaeal genomes leads to several generalizations on the principles of genome organization and evolution. A crucial finding that enables functional characterization of the sequenced genomes and evolutionary reconstruction is that the majority of archaeal and bacterial genes have conserved orthologs in other, often, distant organisms. However, comparative genomics also shows that horizontal gene transfer (HGT) is a dominant force of prokaryotic evolution, along with the loss of genetic material resulting in genome contraction. A crucial component of the prokaryotic world is the mobilome, the enormous collection of viruses, plasmids and other selfish elements, which are in constant exchange with more stable chromosomes and serve as HGT vehicles. Thus, the prokaryotic genome space is a tightly connected, although compartmentalized, network, a novel notion that undermines the ‘Tree of Life’ model of evolution and requires a new conceptual framework and tools for the study of prokaryotic evolution. PMID:18948295

Segregation and Civic Virtue

ERIC Educational Resources Information Center

Merry, Michael S.

2012-01-01

In this essay Michael Merry defends the following prima facie argument: that civic virtue is not dependent on integration and in fact may be best fostered under conditions of segregation. He demonstrates that civic virtue can and does take place under conditions of involuntary segregation, but that voluntary separation--as a response to…

36 CFR 254.6 - Segregative effect.

Code of Federal Regulations, 2010 CFR

2010-07-01

... 36 Parks, Forests, and Public Property 2 2010-07-01 2010-07-01 false Segregative effect. 254.6... ADJUSTMENTS Land Exchanges § 254.6 Segregative effect. (a) If a proposal is made to exchange Federal lands... segregative effect terminates as follows: (1) Automatically, upon issuance of a patent or other document of...

CAR: contig assembly of prokaryotic draft genomes using rearrangements.

PubMed

Lu, Chin Lung; Chen, Kun-Tze; Huang, Shih-Yuan; Chiu, Hsien-Tai

2014-11-28

Next generation sequencing technology has allowed efficient production of draft genomes for many organisms of interest. However, most draft genomes are just collections of independent contigs, whose relative positions and orientations along the genome being sequenced are unknown. Although several tools have been developed to order and orient the contigs of draft genomes, more accurate tools are still needed. In this study, we present a novel reference-based contig assembly (or scaffolding) tool, named as CAR, that can efficiently and more accurately order and orient the contigs of a prokaryotic draft genome based on a reference genome of a related organism. Given a set of contigs in multi-FASTA format and a reference genome in FASTA format, CAR can output a list of scaffolds, each of which is a set of ordered and oriented contigs. For validation, we have tested CAR on a real dataset composed of several prokaryotic genomes and also compared its performance with several other reference-based contig assembly tools. Consequently, our experimental results have shown that CAR indeed performs better than all these other reference-based contig assembly tools in terms of sensitivity, precision and genome coverage. CAR serves as an efficient tool that can more accurately order and orient the contigs of a prokaryotic draft genome based on a reference genome. The web server of CAR is freely available at http://genome.cs.nthu.edu.tw/CAR/ and its stand-alone program can also be downloaded from the same website.

Chromosome segregation drives division site selection in Streptococcus pneumoniae.

PubMed

van Raaphorst, Renske; Kjos, Morten; Veening, Jan-Willem

2017-07-18

Accurate spatial and temporal positioning of the tubulin-like protein FtsZ is key for proper bacterial cell division. Streptococcus pneumoniae (pneumococcus) is an oval-shaped, symmetrically dividing opportunistic human pathogen lacking the canonical systems for division site control (nucleoid occlusion and the Min-system). Recently, the early division protein MapZ was identified and implicated in pneumococcal division site selection. We show that MapZ is important for proper division plane selection; thus, the question remains as to what drives pneumococcal division site selection. By mapping the cell cycle in detail, we show that directly after replication both chromosomal origin regions localize to the future cell division sites, before FtsZ. Interestingly, Z-ring formation occurs coincidently with initiation of DNA replication. Perturbing the longitudinal chromosomal organization by mutating the condensin SMC, by CRISPR/Cas9-mediated chromosome cutting, or by poisoning DNA decatenation resulted in mistiming of MapZ and FtsZ positioning and subsequent cell elongation. Together, we demonstrate an intimate relationship between DNA replication, chromosome segregation, and division site selection in the pneumococcus, providing a simple way to ensure equally sized daughter cells.

A computational genomics pipeline for prokaryotic sequencing projects

PubMed Central

Kislyuk, Andrey O.; Katz, Lee S.; Agrawal, Sonia; Hagen, Matthew S.; Conley, Andrew B.; Jayaraman, Pushkala; Nelakuditi, Viswateja; Humphrey, Jay C.; Sammons, Scott A.; Govil, Dhwani; Mair, Raydel D.; Tatti, Kathleen M.; Tondella, Maria L.; Harcourt, Brian H.; Mayer, Leonard W.; Jordan, I. King

2010-01-01

Motivation: New sequencing technologies have accelerated research on prokaryotic genomes and have made genome sequencing operations outside major genome sequencing centers routine. However, no off-the-shelf solution exists for the combined assembly, gene prediction, genome annotation and data presentation necessary to interpret sequencing data. The resulting requirement to invest significant resources into custom informatics support for genome sequencing projects remains a major impediment to the accessibility of high-throughput sequence data. Results: We present a self-contained, automated high-throughput open source genome sequencing and computational genomics pipeline suitable for prokaryotic sequencing projects. The pipeline has been used at the Georgia Institute of Technology and the Centers for Disease Control and Prevention for the analysis of Neisseria meningitidis and Bordetella bronchiseptica genomes. The pipeline is capable of enhanced or manually assisted reference-based assembly using multiple assemblers and modes; gene predictor combining; and functional annotation of genes and gene products. Because every component of the pipeline is executed on a local machine with no need to access resources over the Internet, the pipeline is suitable for projects of a sensitive nature. Annotation of virulence-related features makes the pipeline particularly useful for projects working with pathogenic prokaryotes. Availability and implementation: The pipeline is licensed under the open-source GNU General Public License and available at the Georgia Tech Neisseria Base (http://nbase.biology.gatech.edu/). The pipeline is implemented with a combination of Perl, Bourne Shell and MySQL and is compatible with Linux and other Unix systems. Contact: king.jordan@biology.gatech.edu Supplementary information: Supplementary data are available at Bioinformatics online. PMID:20519285

Prokaryotic selectivity and LPS-neutralizing activity of short antimicrobial peptides designed from the human antimicrobial peptide LL-37.

PubMed

Nan, Yong Hai; Bang, Jeong-Kyu; Jacob, Binu; Park, Il-Seon; Shin, Song Yub

2012-06-01

To develop novel antimicrobial peptides (AMPs) with shorter lengths, improved prokaryotic selectivity and retained lipolysaccharide (LPS)-neutralizing activity compared to human cathelicidin AMP, LL-37, a series of amino acid-substituted analogs based on IG-19 (residues 13-31 of LL-37) were synthesized. Among the IG-19 analogs, the analog a4 showed the highest prokaryotic selectivity, but much lower LPS-neutralizing activity compared to parental LL-37. The analogs, a5, a6, a7 and a8 with higher hydrophobicity displayed LPS-neutralizing activity comparable to that of LL-37, but much lesser prokaryotic selectivity. These results indicate that the proper hydrophobicity of the peptides is crucial to exert the amalgamated property of LPS-neutralizing activity and prokaryotic selectivity. Furthermore, to increase LPS-neutralizing activity of the analog a4 without a remarkable decrease in prokaryotic selectivity, we synthesized Trp-substituted analogs (a4-W1 and a4-W2), in which Phe(5) or Phe(15) of a4 is replaced by Trp. Despite their same prokaryotic selectivity, a4-W2 displayed much higher LPS-neutralizing activity compared to a4-W1. When compared with parental LL-37, a4-W2 showed retained LPS-neutralizing activity and 2.8-fold enhanced prokaryotic selectivity. These results suggest that the effective site for Trp-substitution when designing novel AMPs with higher LPS-neutralizing activity, without a remarkable reduction in prokaryotic selectivity, is the amphipathic interface between the end of the hydrophilic side and the start of the hydrophobic side rather than the central position of the hydrophobic side in their α-helical wheel projection. Taken together, the analog a4-W2 can serve as a promising template for the development of therapeutic agents for the treatment of endotoxic shock and bacterial infection. Copyright © 2012. Published by Elsevier Inc.

43 CFR 2091.2-1 - Segregation.

Code of Federal Regulations, 2011 CFR

2011-10-01

... 43 Public Lands: Interior 2 2011-10-01 2011-10-01 false Segregation. 2091.2-1 Section 2091.2-1..., DEPARTMENT OF THE INTERIOR LAND RESOURCE MANAGEMENT (2000) SPECIAL LAWS AND RULES Segregation and Opening of Lands § 2091.2-1 Segregation. The publication of a Notice of Realty Action in the Federal Register...

Illegitimate recombination mediated by calf thymus DNA topoisomerase II in vitro.

PubMed Central

Bae, Y S; Kawasaki, I; Ikeda, H; Liu, L F

1988-01-01

We have found that purified calf thymus DNA topoisomerase II mediates recombination between two phage lambda DNA molecules in an in vitro system. The enzyme mainly produced a linear monomer recombinant DNA that can be packaged in vitro. Novobiocin and anti-calf thymus DNA topoisomerase II antibody inhibit this ATP-dependent recombination. The recombinant molecules contain duplications or deletions, and most crossovers take place between nonhomologous sequences of lambda DNA, as judged by the sequences of recombination junctions. Therefore, the recombination mediated by the calf thymus DNA topoisomerase II is an illegitimate recombination that is similar to recombination mediated by Escherichia coli DNA gyrase or phage T4 DNA topoisomerase. The subunit exchange model, which has been suggested for the DNA gyrase-mediated recombination, is now generalized as follows: DNA topoisomerase II molecules bind to DNAs, associate with each other, and lead to the exchange of DNA strands through the exchange of topoisomerase II subunits. Illegitimate recombination might be carried out by a general mechanism in organisms ranging from prokaryotes to higher eukaryotes. Images PMID:2832845

Next-generation systematics: An innovative approach to resolve the structure of complex prokaryotic taxa

NASA Astrophysics Data System (ADS)

Sangal, Vartul; Goodfellow, Michael; Jones, Amanda L.; Schwalbe, Edward C.; Blom, Jochen; Hoskisson, Paul A.; Sutcliffe, Iain C.

2016-12-01

Prokaryotic systematics provides the fundamental framework for microbiological research but remains a discipline that relies on a labour- and time-intensive polyphasic taxonomic approach, including DNA-DNA hybridization, variation in 16S rRNA gene sequence and phenotypic characteristics. These techniques suffer from poor resolution in distinguishing between closely related species and often result in misclassification and misidentification of strains. Moreover, guidelines are unclear for the delineation of bacterial genera. Here, we have applied an innovative phylogenetic and taxogenomic approach to a heterogeneous actinobacterial taxon, Rhodococcus, to identify boundaries for intrageneric and supraspecific classification. Seven species-groups were identified within the genus Rhodococcus that are as distantly related to one another as they are to representatives of other mycolic acid containing actinobacteria and can thus be equated with the rank of genus. It was also evident that strains assigned to rhodococcal species-groups are underspeciated with many misclassified using conventional taxonomic criteria. The phylogenetic and taxogenomic methods used in this study provide data of theoretical value for the circumscription of generic and species boundaries and are also of practical significance as they provide a robust basis for the classification and identification of rhodococci of agricultural, industrial and medical/veterinary significance.

DNA Damage Responses in Prokaryotes: Regulating Gene Expression, Modulating Growth Patterns, and Manipulating Replication Forks

PubMed Central

Kreuzer, Kenneth N.

2013-01-01

Recent advances in the area of bacterial DNA damage responses are reviewed here. The SOS pathway is still the major paradigm of bacterial DNA damage response, and recent studies have clarified the mechanisms of SOS induction and key physiological roles of SOS including a very major role in genetic exchange and variation. When considering diverse bacteria, it is clear that SOS is not a uniform pathway with one purpose, but rather a platform that has evolved for differing functions in different bacteria. Relating in part to the SOS response, the field has uncovered multiple apparent cell-cycle checkpoints that assist cell survival after DNA damage and remarkable pathways that induce programmed cell death in bacteria. Bacterial DNA damage responses are also much broader than SOS, and several important examples of LexA-independent regulation will be reviewed. Finally, some recent advances that relate to the replication and repair of damaged DNA will be summarized. PMID:24097899

Benthic prokaryotic community dynamics along the Ardencaple Canyon, Western Greenland Sea

NASA Astrophysics Data System (ADS)

Quéric, Nadia-Valérie; Soltwedel, Thomas

2012-07-01

The Ardencaple Canyon, emanating from the Eastern Greenland continental rise over a distance of about 200 km towards the Greenland Basin, was investigated to determine the effect of enhanced down-slope transport mechanisms on deep-sea benthic prokaryotic communities. The concentration of viable bacterial cells (Live/Dead®BacLight) and prokaryotic incorporation rates (3H-thymidine, 14C-leucine) increased with increasing distance from the continental shelf. Multidimensional scaling (MDS) results from terminal restriction fragment length polymorphism (T-RFLP) analysis indicated a spatial coherence between the benthic bacterial community structure, prokaryotic incorporation rates, water content, protein concentration and the total organic matter in the sediments. The community complexity in sediments at 4-5 cm depth was lower in the central parts of the channel compared with the northern and the southern levees, while richness in surface sediments of all stations was similar. Lacking any clear indications for a recent mass sediment transport or funneled shelf drainage flows, high similarities between bacterial assemblages in sediments along the canyon course may thus be governed by a combination of an ice-edge induced particle flux, episodic down-slope and canyon-guided transport mechanisms.

In-cell NMR of intrinsically disordered proteins in prokaryotic cells.

PubMed

Ito, Yutaka; Mikawa, Tsutomu; Smith, Brian O

2012-01-01

In-cell NMR, i.e., the acquisition of heteronuclear multidimensional NMR of biomacromolecules inside living cells, is, to our knowledge, the only method for investigating the three-dimensional structure and dynamics of proteins at atomic detail in the intracellular environment. Since the inception of the method, intrinsically disordered proteins have been regarded as particular targets for in-cell NMR, due to their expected sensitivity to the molecular crowding in the intracellular environment. While both prokaryotic and eukaryotic cells can be used as host cells for in-cell NMR, prokaryotic in-cell NMR, particularly employing commonly used protein overexpression systems in Escherichia coli cells, is the most accessible approach. In this chapter we describe general procedures for obtaining in-cell NMR spectra in E. coli cells.

Effects of Predation by Protists on Prokaryotic Community Function, Structure, and Diversity in Anaerobic Granular Sludge.

PubMed

Hirakata, Yuga; Oshiki, Mamoru; Kuroda, Kyohei; Hatamoto, Masashi; Kubota, Kengo; Yamaguchi, Takashi; Harada, Hideki; Araki, Nobuo

2016-09-29

Predation by protists is top-down pressure that regulates prokaryotic abundance, community function, structure, and diversity in natural and artificial ecosystems. Although the effects of predation by protists have been studied in aerobic ecosystems, they are poorly understood in anoxic environments. We herein studied the influence of predation by Metopus and Caenomorpha ciliates-ciliates frequently found in anoxic ecosystems-on prokaryotic community function, structure, and diversity. Metopus and Caenomorpha ciliates were cocultivated with prokaryotic assemblages (i.e., anaerobic granular sludge) in an up-flow anaerobic sludge blanket (UASB) reactor for 171 d. Predation by these ciliates increased the methanogenic activities of granular sludge, which constituted 155% of those found in a UASB reactor without the ciliates (i.e., control reactor). Sequencing of 16S rRNA gene amplicons using Illumina MiSeq revealed that the prokaryotic community in the UASB reactor with the ciliates was more diverse than that in the control reactor; 2,885-3,190 and 2,387-2,426 operational taxonomic units (>97% sequence similarities), respectively. The effects of predation by protists in anaerobic engineered systems have mostly been overlooked, and our results show that the influence of predation by protists needs to be examined and considered in the future for a better understanding of prokaryotic community structure and function.

[Ultrastructural basis of interactions between prokaryotes and eukaryotes in different symbiotic models].

PubMed

Sacchi, L

2004-06-01

This paper reviews the Author's contribution to the knowledge of the ultrastructural basis of the prokaryote-eukaryote interactions in different models assessed by an ultrastructural approach. In agreement with the hypothesis of the origin of eukaryotic cells, which are chimeras of several prokaryotes with different morpho-functional specializations, symbiosis had major consequence for evolution of life. In Arthropods, one of the most successful lifestyles, the presence of endosymbiotic prokaryotes, plays an important role in their metabolism. In some cases, genome integration has occurred in the endosymbiotic relationships with the host, proving that intracellular symbiosis is not merely a nutritional supplement. Intracellular symbiotic bacteria are also described in nematodes. In particular, the presence of intracellular Wolbachia in filariae, even if its function is not yet completely known, influences positively the reproductive biology and the survival of the host, as proved by antibiotic treatment against this bacterium. The ultrastructural images reported in this review were obtained using different species of cockroaches, termites, ticks and filarial nematodes. The traditional methods of transmission (TEM), scansion (SEM) and immuno electron microscopy were used. In addition, also freeze-fracture and deep-etching techniques were employed. The cockroaches and the primitive termite Mastotermes darwiniensis host symbiotic bacteria in the ovary and in specialized cells (bacteriocytes) of the fat body. These bacteria have the typical cell boundary profile of gram-negative bacteria and are enveloped in a vacuolar membrane produced by the host cell. Molecular sequence data of 16S rDNA of endosymbionts of five species of cockroaches and M. darwiniensis indicate that they are members of the Flavobacteria-bacteroides group and that the infection occurred in an ancestor common to cockroaches and termites probably after the end of the Paleozoic (250 Ma BP). The

Colonic stem cell data are consistent with the immortal model of stem cell division under non-random strand segregation.

PubMed

Walters, K

2009-06-01

Colonic stem cells are thought to reside towards the base of crypts of the colon, but their numbers and proliferation mechanisms are not well characterized. A defining property of stem cells is that they are able to divide asymmetrically, but it is not known whether they always divide asymmetrically (immortal model) or whether there are occasional symmetrical divisions (stochastic model). By measuring diversity of methylation patterns in colon crypt samples, a recent study found evidence in favour of the stochastic model, assuming random segregation of stem cell DNA strands during cell division. Here, the effect of preferential segregation of the template strand is considered to be consistent with the 'immortal strand hypothesis', and explore the effect on conclusions of previously published results. For a sample of crypts, it is shown how, under the immortal model, to calculate mean and variance of the number of unique methylation patterns allowing for non-random strand segregation and compare them with those observed. The calculated mean and variance are consistent with an immortal model that incorporates non-random strand segregation for a range of stem cell numbers and levels of preferential strand segregation. Allowing for preferential strand segregation considerably alters previously published conclusions relating to stem cell numbers and turnover mechanisms. Evidence in favour of the stochastic model may not be as strong as previously thought.

Prokaryotic communities in pit mud from different-aged cellars used for the production of Chinese strong-flavored liquor.

PubMed

Tao, Yong; Li, Jiabao; Rui, Junpeng; Xu, Zhancheng; Zhou, Yan; Hu, Xiaohong; Wang, Xiang; Liu, Menghua; Li, Daping; Li, Xiangzhen

2014-04-01

Chinese strong-flavored liquor (CSFL) accounts for more than 70% of all Chinese liquor production. Microbes in pit mud play key roles in the fermentation cellar for the CSFL production. However, microbial diversity, community structure, and cellar-age-related changes in pit mud are poorly understood. Here, we investigated the prokaryotic community structure and diversity in pit-mud samples with different cellar ages (1, 10, 25, and 50 years) using the pyrosequencing technique. Results indicated that prokaryotic diversity increased with cellar age until the age reached 25 years and that prokaryotic community structure changed significantly between three cellar ages (1, 10, and 25 years). Significant correlations between prokaryotic communities and environmental variables (pH, NH4(+), lactic acid, butyric acid, and caproic acid) were observed. Overall, our study results suggested that the long-term brewing operation shapes unique prokaryotic community structure and diversity as well as pit-mud chemistry. We have proposed a three-phase model to characterize the changes of pit-mud prokaryotic communities. (i) Phase I is an initial domestication period. Pit mud is characterized by abundant Lactobacillus and high lactic acid and low pH levels. (ii) Phase II is a transition period. While Lactobacillus abundance decreases dramatically, that of Bacteroidetes and methanogens increases. (iii) Phase III is a relative mature period. The prokaryotic community shows the highest diversity and capability to produce more caproic acid as a precursor for synthesis of ethyl caproate, the main flavor component in CSFL. This research provides scientific evidence to support the practical experience that old fermentation cellars produce high-quality liquor.

Prokaryotic Communities in Pit Mud from Different-Aged Cellars Used for the Production of Chinese Strong-Flavored Liquor

PubMed Central

Tao, Yong; Li, Jiabao; Rui, Junpeng; Xu, Zhancheng; Zhou, Yan; Hu, Xiaohong; Wang, Xiang; Liu, Menghua; Li, Daping

2014-01-01

Chinese strong-flavored liquor (CSFL) accounts for more than 70% of all Chinese liquor production. Microbes in pit mud play key roles in the fermentation cellar for the CSFL production. However, microbial diversity, community structure, and cellar-age-related changes in pit mud are poorly understood. Here, we investigated the prokaryotic community structure and diversity in pit-mud samples with different cellar ages (1, 10, 25, and 50 years) using the pyrosequencing technique. Results indicated that prokaryotic diversity increased with cellar age until the age reached 25 years and that prokaryotic community structure changed significantly between three cellar ages (1, 10, and 25 years). Significant correlations between prokaryotic communities and environmental variables (pH, NH4+, lactic acid, butyric acid, and caproic acid) were observed. Overall, our study results suggested that the long-term brewing operation shapes unique prokaryotic community structure and diversity as well as pit-mud chemistry. We have proposed a three-phase model to characterize the changes of pit-mud prokaryotic communities. (i) Phase I is an initial domestication period. Pit mud is characterized by abundant Lactobacillus and high lactic acid and low pH levels. (ii) Phase II is a transition period. While Lactobacillus abundance decreases dramatically, that of Bacteroidetes and methanogens increases. (iii) Phase III is a relative mature period. The prokaryotic community shows the highest diversity and capability to produce more caproic acid as a precursor for synthesis of ethyl caproate, the main flavor component in CSFL. This research provides scientific evidence to support the practical experience that old fermentation cellars produce high-quality liquor. PMID:24487528

Conservation of coevolving protein interfaces bridges prokaryote-eukaryote homologies in the twilight zone.

PubMed

Rodriguez-Rivas, Juan; Marsili, Simone; Juan, David; Valencia, Alfonso

2016-12-27

Protein-protein interactions are fundamental for the proper functioning of the cell. As a result, protein interaction surfaces are subject to strong evolutionary constraints. Recent developments have shown that residue coevolution provides accurate predictions of heterodimeric protein interfaces from sequence information. So far these approaches have been limited to the analysis of families of prokaryotic complexes for which large multiple sequence alignments of homologous sequences can be compiled. We explore the hypothesis that coevolution points to structurally conserved contacts at protein-protein interfaces, which can be reliably projected to homologous complexes with distantly related sequences. We introduce a domain-centered protocol to study the interplay between residue coevolution and structural conservation of protein-protein interfaces. We show that sequence-based coevolutionary analysis systematically identifies residue contacts at prokaryotic interfaces that are structurally conserved at the interface of their eukaryotic counterparts. In turn, this allows the prediction of conserved contacts at eukaryotic protein-protein interfaces with high confidence using solely mutational patterns extracted from prokaryotic genomes. Even in the context of high divergence in sequence (the twilight zone), where standard homology modeling of protein complexes is unreliable, our approach provides sequence-based accurate information about specific details of protein interactions at the residue level. Selected examples of the application of prokaryotic coevolutionary analysis to the prediction of eukaryotic interfaces further illustrate the potential of this approach.

BLAST Ring Image Generator (BRIG): simple prokaryote genome comparisons

PubMed Central

2011-01-01

Background Visualisation of genome comparisons is invaluable for helping to determine genotypic differences between closely related prokaryotes. New visualisation and abstraction methods are required in order to improve the validation, interpretation and communication of genome sequence information; especially with the increasing amount of data arising from next-generation sequencing projects. Visualising a prokaryote genome as a circular image has become a powerful means of displaying informative comparisons of one genome to a number of others. Several programs, imaging libraries and internet resources already exist for this purpose, however, most are either limited in the number of comparisons they can show, are unable to adequately utilise draft genome sequence data, or require a knowledge of command-line scripting for implementation. Currently, there is no freely available desktop application that enables users to rapidly visualise comparisons between hundreds of draft or complete genomes in a single image. Results BLAST Ring Image Generator (BRIG) can generate images that show multiple prokaryote genome comparisons, without an arbitrary limit on the number of genomes compared. The output image shows similarity between a central reference sequence and other sequences as a set of concentric rings, where BLAST matches are coloured on a sliding scale indicating a defined percentage identity. Images can also include draft genome assembly information to show read coverage, assembly breakpoints and collapsed repeats. In addition, BRIG supports the mapping of unassembled sequencing reads against one or more central reference sequences. Many types of custom data and annotations can be shown using BRIG, making it a versatile approach for visualising a range of genomic comparison data. BRIG is readily accessible to any user, as it assumes no specialist computational knowledge and will perform all required file parsing and BLAST comparisons automatically. Conclusions

BLAST Ring Image Generator (BRIG): simple prokaryote genome comparisons.

PubMed

Alikhan, Nabil-Fareed; Petty, Nicola K; Ben Zakour, Nouri L; Beatson, Scott A

2011-08-08

Visualisation of genome comparisons is invaluable for helping to determine genotypic differences between closely related prokaryotes. New visualisation and abstraction methods are required in order to improve the validation, interpretation and communication of genome sequence information; especially with the increasing amount of data arising from next-generation sequencing projects. Visualising a prokaryote genome as a circular image has become a powerful means of displaying informative comparisons of one genome to a number of others. Several programs, imaging libraries and internet resources already exist for this purpose, however, most are either limited in the number of comparisons they can show, are unable to adequately utilise draft genome sequence data, or require a knowledge of command-line scripting for implementation. Currently, there is no freely available desktop application that enables users to rapidly visualise comparisons between hundreds of draft or complete genomes in a single image. BLAST Ring Image Generator (BRIG) can generate images that show multiple prokaryote genome comparisons, without an arbitrary limit on the number of genomes compared. The output image shows similarity between a central reference sequence and other sequences as a set of concentric rings, where BLAST matches are coloured on a sliding scale indicating a defined percentage identity. Images can also include draft genome assembly information to show read coverage, assembly breakpoints and collapsed repeats. In addition, BRIG supports the mapping of unassembled sequencing reads against one or more central reference sequences. Many types of custom data and annotations can be shown using BRIG, making it a versatile approach for visualising a range of genomic comparison data. BRIG is readily accessible to any user, as it assumes no specialist computational knowledge and will perform all required file parsing and BLAST comparisons automatically. There is a clear need for a user

DNA gyrase with a single catalytic tyrosine can catalyze DNA supercoiling by a nicking-closing mechanism

PubMed Central

Gubaev, Airat; Weidlich, Daniela; Klostermeier, Dagmar

2016-01-01

The topological state of DNA is important for replication, recombination and transcription, and is regulated in vivo by DNA topoisomerases. Gyrase introduces negative supercoils into DNA at the expense of ATP hydrolysis. It is the accepted view that gyrase achieves supercoiling by a strand passage mechanism, in which double-stranded DNA is cleaved, and a second double-stranded segment is passed through the gap, converting a positive DNA node into a negative node. We show here that gyrase with only one catalytic tyrosine that cleaves a single strand of its DNA substrate can catalyze DNA supercoiling without strand passage. We propose an alternative mechanism for DNA supercoiling via nicking and closing of DNA that involves trapping, segregation and relaxation of two positive supercoils. In contrast to DNA supercoiling, ATP-dependent relaxation and decatenation of DNA by gyrase lacking the C-terminal domains require both tyrosines and strand passage. Our results point towards mechanistic plasticity of gyrase and might pave the way for finding novel and specific mechanism-based gyrase inhibitors. PMID:27557712

Mechanisms of bacterial DNA replication restart

PubMed Central

Windgassen, Tricia A; Wessel, Sarah R; Bhattacharyya, Basudeb

2018-01-01

Abstract Multi-protein DNA replication complexes called replisomes perform the essential process of copying cellular genetic information prior to cell division. Under ideal conditions, replisomes dissociate only after the entire genome has been duplicated. However, DNA replication rarely occurs without interruptions that can dislodge replisomes from DNA. Such events produce incompletely replicated chromosomes that, if left unrepaired, prevent the segregation of full genomes to daughter cells. To mitigate this threat, cells have evolved ‘DNA replication restart’ pathways that have been best defined in bacteria. Replication restart requires recognition and remodeling of abandoned replication forks by DNA replication restart proteins followed by reloading of the replicative DNA helicase, which subsequently directs assembly of the remaining replisome subunits. This review summarizes our current understanding of the mechanisms underlying replication restart and the proteins that drive the process in Escherichia coli (PriA, PriB, PriC and DnaT). PMID:29202195

Archaeal replicative primases can perform translesion DNA synthesis.

PubMed

Jozwiakowski, Stanislaw K; Borazjani Gholami, Farimah; Doherty, Aidan J

2015-02-17

DNA replicases routinely stall at lesions encountered on the template strand, and translesion DNA synthesis (TLS) is used to rescue progression of stalled replisomes. This process requires specialized polymerases that perform translesion DNA synthesis. Although prokaryotes and eukaryotes possess canonical TLS polymerases (Y-family Pols) capable of traversing blocking DNA lesions, most archaea lack these enzymes. Here, we report that archaeal replicative primases (Pri S, primase small subunit) can also perform TLS. Archaeal Pri S can bypass common oxidative DNA lesions, such as 8-Oxo-2'-deoxyguanosines and UV light-induced DNA damage, faithfully bypassing cyclobutane pyrimidine dimers. Although it is well documented that archaeal replicases specifically arrest at deoxyuracils (dUs) due to recognition and binding to the lesions, a replication restart mechanism has not been identified. Here, we report that Pri S efficiently replicates past dUs, even in the presence of stalled replicase complexes, thus providing a mechanism for maintaining replication bypass of these DNA lesions. Together, these findings establish that some replicative primases, previously considered to be solely involved in priming replication, are also TLS proficient and therefore may play important roles in damage tolerance at replication forks.

Systematic Transfer of Prokaryotic Sensors and Circuits to Mammalian Cells

PubMed Central

2015-01-01

Prokaryotic regulatory proteins respond to diverse signals and represent a rich resource for building synthetic sensors and circuits. The TetR family contains >105 members that use a simple mechanism to respond to stimuli and bind distinct DNA operators. We present a platform that enables the transfer of these regulators to mammalian cells, which is demonstrated using human embryonic kidney (HEK293) and Chinese hamster ovary (CHO) cells. The repressors are modified to include nuclear localization signals (NLS) and responsive promoters are built by incorporating multiple operators. Activators are also constructed by modifying the protein to include a VP16 domain. Together, this approach yields 15 new regulators that demonstrate 19- to 551-fold induction and retain both the low levels of crosstalk in DNA binding specificity observed between the parent regulators in Escherichia coli, as well as their dynamic range of activity. By taking advantage of the DAPG small molecule sensing mediated by the PhlF repressor, we introduce a new inducible system with 50-fold induction and a threshold of 0.9 μM DAPG, which is comparable to the classic Dox-induced TetR system. A set of NOT gates is constructed from the new repressors and their response function quantified. Finally, the Dox- and DAPG- inducible systems and two new activators are used to build a synthetic enhancer (fuzzy AND gate), requiring the coordination of 5 transcription factors organized into two layers. This work introduces a generic approach for the development of mammalian genetic sensors and circuits to populate a toolbox that can be applied to diverse applications from biomanufacturing to living therapeutics. PMID:25360681

Systematic transfer of prokaryotic sensors and circuits to mammalian cells.

PubMed

Stanton, Brynne C; Siciliano, Velia; Ghodasara, Amar; Wroblewska, Liliana; Clancy, Kevin; Trefzer, Axel C; Chesnut, Jonathan D; Weiss, Ron; Voigt, Christopher A

2014-12-19

Prokaryotic regulatory proteins respond to diverse signals and represent a rich resource for building synthetic sensors and circuits. The TetR family contains >10(5) members that use a simple mechanism to respond to stimuli and bind distinct DNA operators. We present a platform that enables the transfer of these regulators to mammalian cells, which is demonstrated using human embryonic kidney (HEK293) and Chinese hamster ovary (CHO) cells. The repressors are modified to include nuclear localization signals (NLS) and responsive promoters are built by incorporating multiple operators. Activators are also constructed by modifying the protein to include a VP16 domain. Together, this approach yields 15 new regulators that demonstrate 19- to 551-fold induction and retain both the low levels of crosstalk in DNA binding specificity observed between the parent regulators in Escherichia coli, as well as their dynamic range of activity. By taking advantage of the DAPG small molecule sensing mediated by the PhlF repressor, we introduce a new inducible system with 50-fold induction and a threshold of 0.9 μM DAPG, which is comparable to the classic Dox-induced TetR system. A set of NOT gates is constructed from the new repressors and their response function quantified. Finally, the Dox- and DAPG- inducible systems and two new activators are used to build a synthetic enhancer (fuzzy AND gate), requiring the coordination of 5 transcription factors organized into two layers. This work introduces a generic approach for the development of mammalian genetic sensors and circuits to populate a toolbox that can be applied to diverse applications from biomanufacturing to living therapeutics.

Broken replication forks trigger heritable DNA breaks in the terminus of a circular chromosome

PubMed Central

Possoz, Christophe; Durand, Adeline; Desfontaines, Jean-Michel; Barre, François-Xavier; Leach, David R. F.

2018-01-01

It was recently reported that the recBC mutants of Escherichia coli, deficient for DNA double-strand break (DSB) repair, have a decreased copy number of their terminus region. We previously showed that this deficit resulted from DNA loss after post-replicative breakage of one of the two sister-chromosome termini at cell division. A viable cell and a dead cell devoid of terminus region were thus produced and, intriguingly, the reaction was transmitted to the following generations. Using genome marker frequency profiling and observation by microscopy of specific DNA loci within the terminus, we reveal here the origin of this phenomenon. We observed that terminus DNA loss was reduced in a recA mutant by the double-strand DNA degradation activity of RecBCD. The terminus-less cell produced at the first cell division was less prone to divide than the one produced at the next generation. DNA loss was not heritable if the chromosome was linearized in the terminus and occurred at chromosome termini that were unable to segregate after replication. We propose that in a recB mutant replication fork breakage results in the persistence of a linear DNA tail attached to a circular chromosome. Segregation of the linear and circular parts of this “σ-replicating chromosome” causes terminus DNA breakage during cell division. One daughter cell inherits a truncated linear chromosome and is not viable. The other inherits a circular chromosome attached to a linear tail ending in the chromosome terminus. Replication extends this tail, while degradation of its extremity results in terminus DNA loss. Repeated generation and segregation of new σ-replicating chromosomes explains the heritability of post-replicative breakage. Our results allow us to determine that in E. coli at each generation, 18% of cells are subject to replication fork breakage at dispersed, potentially random, chromosomal locations. PMID:29522563

Short Term INT-Formazan Production as a Proxy for Marine Prokaryote Respiration

NASA Astrophysics Data System (ADS)

Cajal-Medrano, R.; Villegas-Mendoza, J.; Maske, H.

2016-02-01

Prokaryotes are poisoned by the tetrazolium electron transport probe INT on time scales of less than one hour, invalidating the interpretation of the rate of in vivo INT reduction to formazan as a proxy for oxygen consumption rates (Villegas-Mendoza et al. 2015). We measured oxygen consumption rate (R; µM O2 hour-1) and electron transport activity with in vivo INT formazan production (IFP, mM formazan) at 0.5 mM INT during 1 hour exposure time of natural communities and cultures of the marine bacteria Vibrio harveyi growing in batch and continuous cultures. A strong exponential relationship R = 0.20 IFP2.15 (p<0.05) with oxygen consumption and total formazan production was found over a wide range of growth rates under aerobic condition. We find that IFP and oxygen consumption increase with bacterial specific growth rates and temperature as expected from basic principles of physiology and biochemistry. Oxygen and nitrogen saturated batch cultures of V. harveyi showed that both, IFP and oxygen consumption increased for 0.8 hours but then stopped similar to natural bacterial communities supporting the above relationship of IFP to prokaryote respiration. Our method implies adding 0.5 mM INT to a plankton sample and incubating for less than 1 hour. After prokaryote separation by size filtration (0.8 mm), the formazan crystals are collected by filtration (0.2 mm) and dissolved in propanol. The absorbance at 485 nm per sample volume yields the formazan potential that is related to prokaryote respiration in the sample.

Non-Random Inversion Landscapes in Prokaryotic Genomes Are Shaped by Heterogeneous Selection Pressures

PubMed Central

Repar, Jelena; Warnecke, Tobias

2017-01-01

Abstract Inversions are a major contributor to structural genome evolution in prokaryotes. Here, using a novel alignment-based method, we systematically compare 1,651 bacterial and 98 archaeal genomes to show that inversion landscapes are frequently biased toward (symmetric) inversions around the origin–terminus axis. However, symmetric inversion bias is not a universal feature of prokaryotic genome evolution but varies considerably across clades. At the extremes, inversion landscapes in Bacillus–Clostridium and Actinobacteria are dominated by symmetric inversions, while there is little or no systematic bias favoring symmetric rearrangements in archaea with a single origin of replication. Within clades, we find strong but clade-specific relationships between symmetric inversion bias and different features of adaptive genome architecture, including the distance of essential genes to the origin of replication and the preferential localization of genes on the leading strand. We suggest that heterogeneous selection pressures have converged to produce similar patterns of structural genome evolution across prokaryotes. PMID:28407093

Exploring cultivable Bacteria from the prokaryotic community associated with the carnivorous sponge Asbestopluma hypogea.

PubMed

Dupont, Samuel; Carre-Mlouka, Alyssa; Domart-Coulon, Isabelle; Vacelet, Jean; Bourguet-Kondracki, Marie-Lise

2014-04-01

Combining culture-dependent and independent approaches, we investigated for the first time the cultivable fraction of the prokaryotic community associated with the carnivorous sponge Asbestopluma hypogea. The heterotrophic prokaryotes isolated from this tiny sponge were compared between specimens freshly collected from cave and maintained in aquarium. Overall, 67 isolates obtained in pure culture were phylogenetically affiliated to the bacterial phyla Proteobacteria, Bacteroidetes, Actinobacteria, and Firmicutes. This cultivable diversity was lower than the prokaryotic diversity obtained by previous pyrosequencing study and comparable to that of another Mediterranean demosponge, the filter-feeding Phorbas tenacior. Furthermore, using fluorescence in situ hybridization, we visualized bacterial and archaeal cells, confirming the presence of both prokaryotes in A. hypogea tissue. Approximately 16% of the bacterial isolates tested positive for chitinolytic activity, suggesting potential microbial involvement in the digestion processes of crustacean prey by this carnivorous sponge. Additionally, 6% and 16% of bacterial isolates revealed antimicrobial and antioxidant activities, respectively. One Streptomyces sp. S1CA strain was identified as a promising candidate for the production of antimicrobial and antioxidant secondary metabolites as well as chitinolytic enzymes. Implications in the context of the sponge biology and prey-feeding strategy are discussed. © 2014 Federation of European Microbiological Societies. Published by John Wiley & Sons Ltd. All rights reserved.

Mechanisms of time-based figure-ground segregation.

PubMed

Kandil, Farid I; Fahle, Manfred

2003-11-01

Figure-ground segregation can rely on purely temporal information, that is, on short temporal delays between positional changes of elements in figure and ground (Kandil, F.I. & Fahle, M. (2001) Eur. J. Neurosci., 13, 2004-2008). Here, we investigate the underlying mechanisms by measuring temporal segregation thresholds for various kinds of motion cues. Segregation can rely on monocular first-order motion (based on luminance modulation) and second-order motion cues (contrast modulation) with a high temporal resolution of approximately 20 ms. The mechanism can also use isoluminant motion with a reduced temporal resolution of 60 ms. Figure-ground segregation can be achieved even at presentation frequencies too high for human subjects to inspect successive frames individually. In contrast, when stimuli are presented dichoptically, i.e. separately to both eyes, subjects are unable to perceive any segregation, irrespective of temporal frequency. We propose that segregation in these displays is detected by a mechanism consisting of at least two stages. On the first level, standard motion or flicker detectors signal local positional changes (flips). On the second level, a segregation mechanism combines the local activities of the low-level detectors with high temporal precision. Our findings suggest that the segregation mechanism can rely on monocular detectors but not on binocular mechanisms. Moreover, the results oppose the idea that segregation in these displays is achieved by motion detectors of a higher order (motion-from-motion), but favour mechanisms sensitive to short temporal delays even without activation of higher-order motion detectors.

46 CFR 148.120 - Stowage and segregation requirements.

Code of Federal Regulations, 2011 CFR

2011-10-01

... 46 Shipping 5 2011-10-01 2011-10-01 false Stowage and segregation requirements. 148.120 Section... OF BULK SOLID MATERIALS THAT REQUIRE SPECIAL HANDLING Stowage and Segregation § 148.120 Stowage and segregation requirements. (a) Each material listed in Table 148.10 of this part must be segregated from...

A longitudinal study of administrative segregation.

PubMed

O'Keefe, Maureen L; Klebe, Kelli J; Metzner, Jeffrey; Dvoskin, Joel; Fellner, Jamie; Stucker, Alysha

2013-01-01

The use of administrative segregation for inmates with and without mental illness has generated considerable criticism. Segregated inmates are locked in single cells for 23 hours per day, are subjected to rigorous security procedures, and have restricted access to programs. In this study, we examined whether inmates in segregation would show greater deterioration over time on psychological symptoms than would comparison offenders. The subjects were male inmates, with and without mental illness, in administrative segregation, general population, or special-needs prison. Subjects completed the Brief Symptom Inventory at regular intervals for one year. Results showed differentiation between groups at the outset and statistically significant but small positive change over time across all groups. All groups showed the same change pattern such that there was not the hypothesized differential change of inmates within administrative segregation. This study advances the empirical research, but replication research is needed to make a better determination of whether and under what conditions harm may or may not occur to inmates in solitary confinement.

Continuum modelling of segregating tridisperse granular chute flow

NASA Astrophysics Data System (ADS)

Deng, Zhekai; Umbanhowar, Paul B.; Ottino, Julio M.; Lueptow, Richard M.

2018-03-01

Segregation and mixing of size multidisperse granular materials remain challenging problems in many industrial applications. In this paper, we apply a continuum-based model that captures the effects of segregation, diffusion and advection for size tridisperse granular flow in quasi-two-dimensional chute flow. The model uses the kinematics of the flow and other physical parameters such as the diffusion coefficient and the percolation length scale, quantities that can be determined directly from experiment, simulation or theory and that are not arbitrarily adjustable. The predictions from the model are consistent with experimentally validated discrete element method (DEM) simulations over a wide range of flow conditions and particle sizes. The degree of segregation depends on the Péclet number, Pe, defined as the ratio of the segregation rate to the diffusion rate, the relative segregation strength κij between particle species i and j, and a characteristic length L, which is determined by the strength of segregation between smallest and largest particles. A parametric study of particle size, κij, Pe and L demonstrates how particle segregation patterns depend on the interplay of advection, segregation and diffusion. Finally, the segregation pattern is also affected by the velocity profile and the degree of basal slip at the chute surface. The model is applicable to different flow geometries, and should be easily adapted to segregation driven by other particle properties such as density and shape.

Image segregation in strabismic amblyopia.

PubMed

Levi, Dennis M

2007-06-01

Humans with naturally occurring amblyopia show deficits thought to involve mechanisms downstream of V1. These include excessive crowding, abnormal global image processing, spatial sampling and symmetry detection and undercounting. Several recent studies suggest that humans with naturally occurring amblyopia show deficits in global image segregation. The current experiments were designed to study figure-ground segregation in amblyopic observers with documented deficits in crowding, symmetry detection, spatial sampling and counting, using similar stimuli. Observers had to discriminate the orientation of a figure (an "E"-like pattern made up of 17 horizontal Gabor patches), embedded in a 7x7 array of Gabor patches. When the 32 "background" patches are vertical, the "E" pops-out, due to segregation by orientation and performance is perfect; however, if the background patches are all, or mostly horizontal, the "E" is camouflaged, and performance is random. Using a method of constant stimuli, we varied the number of "background" patches that were vertical and measured the probability of correct discrimination of the global orientation of the E (up/down/left/right). Surprisingly, amblyopes who showed strong crowding and deficits in symmetry detection and counting, perform normally or very nearly so in this segregation task. I therefore conclude that these deficits are not a consequence of abnormal segregation of figure from background.

A Novel Method for Accurate Operon Predictions in All SequencedProkaryotes

DOE Office of Scientific and Technical Information (OSTI.GOV)

Price, Morgan N.; Huang, Katherine H.; Alm, Eric J.

2004-12-01

We combine comparative genomic measures and the distance separating adjacent genes to predict operons in 124 completely sequenced prokaryotic genomes. Our method automatically tailors itself to each genome using sequence information alone, and thus can be applied to any prokaryote. For Escherichia coli K12 and Bacillus subtilis, our method is 85 and 83% accurate, respectively, which is similar to the accuracy of methods that use the same features but are trained on experimentally characterized transcripts. In Halobacterium NRC-1 and in Helicobacterpylori, our method correctly infers that genes in operons are separated by shorter distances than they are in E.coli, andmore » its predictions using distance alone are more accurate than distance-only predictions trained on a database of E.coli transcripts. We use microarray data from sixphylogenetically diverse prokaryotes to show that combining intergenic distance with comparative genomic measures further improves accuracy and that our method is broadly effective. Finally, we survey operon structure across 124 genomes, and find several surprises: H.pylori has many operons, contrary to previous reports; Bacillus anthracis has an unusual number of pseudogenes within conserved operons; and Synechocystis PCC6803 has many operons even though it has unusually wide spacings between conserved adjacent genes.« less

28 CFR 541.24 - Disciplinary segregation status.

Code of Federal Regulations, 2011 CFR

2011-07-01

... 28 Judicial Administration 2 2011-07-01 2011-07-01 false Disciplinary segregation status. 541.24... INMATE DISCIPLINE AND SPECIAL HOUSING UNITS Special Housing Units § 541.24 Disciplinary segregation status. You may be placed in disciplinary segregation status only by the DHO as a disciplinary sanction. ...

Spatiotemporal and species variations in prokaryotic communities associated with sediments from surface-flow constructed wetlands for treating swine wastewater.

PubMed

Jia, Fen; Lai, Cui; Chen, Liang; Zeng, Guangming; Huang, Danlian; Liu, Feng; Li, Xi; Luo, Pei; Wu, Jinshui; Qin, Lei; Zhang, Chen; Cheng, Min; Xu, Piao

2017-10-01

Microorganisms are the main mechanisms of pollutants removals in constructed wetlands (CWs) used for wastewater treatment. However, the different biological processes and variations of prokaryotic community in CWs remain poorly understood. In this study, we applied a high-throughput sequencing technique to investigate the prokaryotic communities associated with sediments from pilot-scale surface-flow constructed wetlands (SFCWs) treating swine wastewater (SW) of varying strengths. Our results revealed that highly diverse prokaryotic communities were present in the SFCWs, with Proteobacteria (16.44-44.44%), Acidobacteria (3.25-24.40%), and Chloroflexi (5.77-14.43%) being the major phyla, and Nitrospira (4.14-12.02%), the most dominant genus. The prokaryotic communities in the sediments varied greatly with location and season, which markedly altered the microenvironmental conditions. Principal co-ordinates analysis indicated that SW strength significantly influenced the community structure in sediments of the SFCWs, and canonical correspondence analysis illustrated that the shifts in prokaryotic communities were strongly related to NO 3 - -N and TN in winter; and in summer with NH 4 + N, NO 3 - -N, NO 2 - -N, TN, TP, SOM, and pH. In conclusion, the use of high-throughput sequencing greatly enhanced our understanding of prokaryotic communities with different functional groups in SFCWs. Copyright © 2017 Elsevier Ltd. All rights reserved.

Cellular strategies for regulating DNA supercoiling: A single-molecule perspective

PubMed Central

Koster, Daniel A.; Crut, Aurélien; Shuman, Stewart; Bjornsti, Mary-Ann; Dekker, Nynke H.

2010-01-01

Summary Excess entangling and twisting of cellular DNA (i.e., DNA supercoiling) are problems inherent to the helical structure of double-stranded DNA. Supercoiling affects transcription, DNA replication, and chromosomal segregation. Consequently the cell must fine-tune supercoiling to optimize these key processes. Here, we summarize how supercoiling is generated and review experimental and theoretical insights into supercoil relaxation. We distinguish between the passive dissipation of supercoils by diffusion and the active removal of supercoils by topoisomerase enzymes. We also review single-molecule studies that elucidate the timescales and mechanisms of supercoil removal. PMID:20723754

A Demonstration of Sample Segregation

ERIC Educational Resources Information Center

Fritz, Mark D.; Brumbach, Stephen B.; Hartman, JudithAnn R.

2005-01-01

The demonstration of sample segregation, which is simple, and visually compelling illustrates the importance of sample handling for students studying analytical chemistry and environmental chemistry. The mixture used in this demonstration has two components, which have big particle size, and different colors, which makes the segregation graphic.

Residential Segregation and Racial Cancer Disparities: A Systematic Review.

PubMed

Landrine, Hope; Corral, Irma; Lee, Joseph G L; Efird, Jimmy T; Hall, Marla B; Bess, Jukelia J

2017-12-01

This paper provides the first review of empirical studies of segregation and black-white cancer disparities. We searched all years of PubMed (through May 2016) using these terms: racial segregation, residential segregation, neighborhood racial composition (first terms) and (second terms) cancer incidence, mortality, survival, stage at diagnosis, screening. The 17 (of 668) articles that measured both segregation and a cancer outcome were retained. Segregation contributed significantly to cancer and to racial cancer disparities in 70% of analyses, even after controlling for socioeconomic status and health insurance. Residing in segregated African-American areas was associated with higher odds of later-stage diagnosis of breast and lung cancers, higher mortality rates and lower survival rates from breast and lung cancers, and higher cumulative cancer risks associated with exposure to ambient air toxics. There were no studies of many types of cancer (e.g., cervical). Studies differed in their measure of segregation, and 40% used an invalid measure. Possible mediators of the segregation effect usually were not tested. Empirical analysis of segregation and racial cancer disparities is a recent area of research. The literature is limited to 17 studies that focused primarily on breast cancer. Studies differed in their measure of segregation, yet segregation nonetheless contributed to cancer and to racial cancer disparities in 70% of analyses. This suggests the need for further research that uses valid measures of segregation, examines a variety of types of cancers, and explores the variables that may mediate the segregation effect.

Engineering prokaryotic transcriptional activators as metabolite biosensors in yeast.

PubMed

Skjoedt, Mette L; Snoek, Tim; Kildegaard, Kanchana R; Arsovska, Dushica; Eichenberger, Michael; Goedecke, Tobias J; Rajkumar, Arun S; Zhang, Jie; Kristensen, Mette; Lehka, Beata J; Siedler, Solvej; Borodina, Irina; Jensen, Michael K; Keasling, Jay D

2016-11-01

Whole-cell biocatalysts have proven a tractable path toward sustainable production of bulk and fine chemicals. Yet the screening of libraries of cellular designs to identify best-performing biocatalysts is most often a low-throughput endeavor. For this reason, the development of biosensors enabling real-time monitoring of production has attracted attention. Here we applied systematic engineering of multiple parameters to search for a general biosensor design in the budding yeast Saccharomyces cerevisiae based on small-molecule binding transcriptional activators from the prokaryote superfamily of LysR-type transcriptional regulators (LTTRs). We identified a design supporting LTTR-dependent activation of reporter gene expression in the presence of cognate small-molecule inducers. As proof of principle, we applied the biosensors for in vivo screening of cells producing naringenin or cis,cis-muconic acid at different levels, and found that reporter gene output correlated with production. The transplantation of prokaryotic transcriptional activators into the eukaryotic chassis illustrates the potential of a hitherto untapped biosensor resource useful for biotechnological applications.

27 CFR 24.27 - Segregation of operations.

Code of Federal Regulations, 2010 CFR

2010-04-01

... 27 Alcohol, Tobacco Products and Firearms 1 2010-04-01 2010-04-01 false Segregation of operations. 24.27 Section 24.27 Alcohol, Tobacco Products and Firearms ALCOHOL AND TOBACCO TAX AND TRADE BUREAU... Segregation of operations. The appropriate TTB officer may require the proprietor to segregate operations...

27 CFR 24.27 - Segregation of operations.

Code of Federal Regulations, 2011 CFR

2011-04-01

... 27 Alcohol, Tobacco Products and Firearms 1 2011-04-01 2011-04-01 false Segregation of operations. 24.27 Section 24.27 Alcohol, Tobacco Products and Firearms ALCOHOL AND TOBACCO TAX AND TRADE BUREAU... Segregation of operations. The appropriate TTB officer may require the proprietor to segregate operations...

Mixing and segregation of microspheres in microchannel flows of mono- and bidispersed suspensions

NASA Astrophysics Data System (ADS)

Gao, C.; Xu, B.; Gilchrist, J. F.

2009-03-01

We investigate the mixing and segregation of mono- and bidispersed microsphere suspensions in microchannel flows. These flows are common in biological microelectromechanical systems (BioMEMS) applications handling blood or suspensions of DNA. Suspension transport in pressure driven flows is significantly hindered by shear-induced migration, where particles migrate away from the walls and are focused in the center due to multibody hydrodynamic interactions. The microchannels used in this study have geometries that induce chaotic advection in Newtonian fluids. Our results show that mixing in straight, herringbone and staggered herringbone channels depends strongly on volume fraction. Due to this complex interplay of advection and shear-induced migration, a staggered herringbone channel that typically results in chaotic mixing is not always effective for dispersing particles. The maximum degree of segregation is observed in a straight channel once the maximum packing fraction is reached at channel center. We modify a one-dimensional suspension balance model [R. Miller and J. Morris, J. Non-Newtonian Fluid Mech. 135, 149 (2006)] to describe the behavior at the center of the straight channel. The degree of mixing is then calculated as a function of bulk volume fraction, predicting the volume fraction that results in the maximum degree of segregation. In bidispersed suspension flow, it is shown that mixing of the larger species is enhanced in straight and staggered herringbone channels while segregation is enhanced at moderate volume fractions in herringbone channels. This suggests mixing and separations can be tailored by adjusting both the suspension properties and the channel geometry.

Resource-limited heterotrophic prokaryote production and its potential environmental impact associated with Mn nodule exploitation in the northeast equatorial pacific.

PubMed

Hyun, Jung-Ho

2006-08-01

Shipboard enrichment incubation experiments were performed to elucidate the limiting resources for heterotrophic prokaryotic production and to discuss the potential impact of bottom water and sediment discharges in relation to manganese (Mn) nodule exploitation on the heterotrophic prokaryotes in the oligotrophic northeast equatorial Pacific. Compared to an unamended control, the production of heterotrophic prokaryotes increased 25-fold in water samples supplemented with amino acids (i.e., organic carbon plus nitrogen), whereas the production increased five and two times, respectively, in samples supplemented with either glucose or ammonium alone. These results indicate that heterotrophic prokaryote production in the northeast equatorial Pacific was co-limited by the availability of dissolved organic carbon and inorganic nitrogen. In samples from the nutrient-depleted surface mixed layer (10-m depth), the addition of a slurry of bottom water and sediment doubled heterotrophic prokaryote production compared to an unamended control, whereas sonicating the slurry prior to addition quadrupled the production rate. However, little difference was observed between an unamended control and slurry-amended samples in the subsurface chlorophyll a (Chl a) maximum (SCM) layer. Thus, the impact of slurry discharge is more significant at the nutrient-depleted surface mixed layer than at the high-nutrient SCM layer. The greatly enhanced prokaryote production resulting from the addition of sonicated slurry further suggests that dissociated organic carbon may directly stimulate heterotrophic prokaryote production in the surface mixed layer. Overall, the results suggest that the surface discharge of bottom water and sediments during manganese nodule exploitation could have a significant environmental impact on the production of heterotrophic prokaryotes that are currently resource limited.

17 CFR 32.6 - Segregation.

Code of Federal Regulations, 2011 CFR

2011-04-01

... 17 Commodity and Securities Exchanges 1 2011-04-01 2011-04-01 false Segregation. 32.6 Section 32.6 Commodity and Securities Exchanges COMMODITY FUTURES TRADING COMMISSION REGULATION OF COMMODITY OPTION TRANSACTIONS § 32.6 Segregation. (a) Any person which accepts money, securities, or property from an option...

17 CFR 32.6 - Segregation.

Code of Federal Regulations, 2010 CFR

2010-04-01

... 17 Commodity and Securities Exchanges 1 2010-04-01 2010-04-01 false Segregation. 32.6 Section 32.6 Commodity and Securities Exchanges COMMODITY FUTURES TRADING COMMISSION REGULATION OF COMMODITY OPTION TRANSACTIONS § 32.6 Segregation. (a) Any person which accepts money, securities, or property from an option...

GlycoPP: A Webserver for Prediction of N- and O-Glycosites in Prokaryotic Protein Sequences

PubMed Central

Chauhan, Jagat S.; Bhat, Adil H.; Raghava, Gajendra P. S.; Rao, Alka

2012-01-01

Glycosylation is one of the most abundant post-translational modifications (PTMs) required for various structure/function modulations of proteins in a living cell. Although elucidated recently in prokaryotes, this type of PTM is present across all three domains of life. In prokaryotes, two types of protein glycan linkages are more widespread namely, N- linked, where a glycan moiety is attached to the amide group of Asn, and O- linked, where a glycan moiety is attached to the hydroxyl group of Ser/Thr/Tyr. For their biologically ubiquitous nature, significance, and technology applications, the study of prokaryotic glycoproteins is a fast emerging area of research. Here we describe new Support Vector Machine (SVM) based algorithms (models) developed for predicting glycosylated-residues (glycosites) with high accuracy in prokaryotic protein sequences. The models are based on binary profile of patterns, composition profile of patterns, and position-specific scoring matrix profile of patterns as training features. The study employ an extensive dataset of 107 N-linked and 116 O-linked glycosites extracted from 59 experimentally characterized glycoproteins of prokaryotes. This dataset includes validated N-glycosites from phyla Crenarchaeota, Euryarchaeota (domain Archaea), Proteobacteria (domain Bacteria) and validated O-glycosites from phyla Actinobacteria, Bacteroidetes, Firmicutes and Proteobacteria (domain Bacteria). In view of the current understanding that glycosylation occurs on folded proteins in bacteria, hybrid models have been developed using information on predicted secondary structures and accessible surface area in various combinations with training features. Using these models, N-glycosites and O-glycosites could be predicted with an accuracy of 82.71% (MCC 0.65) and 73.71% (MCC 0.48), respectively. An evaluation of the best performing models with 28 independent prokaryotic glycoproteins confirms the suitability of these models in predicting N- and O

Rapid phylogenetic dissection of prokaryotic community structure in tidal flat using pyrosequencing.

PubMed

Kim, Bong-Soo; Kim, Byung Kwon; Lee, Jae-Hak; Kim, Myungjin; Lim, Young Woon; Chun, Jongsik

2008-08-01

Dissection of prokaryotic community structure is prerequisite to understand their ecological roles. Various methods are available for such a purpose which amplification and sequencing of 16S rRNA genes gained its popularity. However, conventional methods based on Sanger sequencing technique require cloning process prior to sequencing, and are expensive and labor-intensive. We investigated prokaryotic community structure in tidal flat sediments, Korea, using pyrosequencing and a subsequent automated bioinformatic pipeline for the rapid and accurate taxonomic assignment of each amplicon. The combination of pyrosequencing and bioinformatic analysis showed that bacterial and archaeal communities were more diverse than previously reported in clone library studies. Pyrosequencing analysis revealed 21 bacterial divisions and 37 candidate divisions. Proteobacteria was the most abundant division in the bacterial community, of which Gamma-and Delta-Proteobacteria were the most abundant. Similarly, 4 archaeal divisions were found in tidal flat sediments. Euryarchaeota was the most abundant division in the archaeal sequences, which were further divided into 8 classes and 11 unclassified euryarchaeota groups. The system developed here provides a simple, in-depth and automated way of dissecting a prokaryotic community structure without extensive pretreatment such as cloning.

DNA damage and polyploidization.

PubMed

Chow, Jeremy; Poon, Randy Y C

2010-01-01

A growing body of evidence indicates that polyploidization triggers chromosomal instability and contributes to tumorigenesis. DNA damage is increasingly being recognized for its roles in promoting polyploidization. Although elegant mechanisms known as the DNA damage checkpoints are responsible for halting the cell cycle after DNA damage, agents that uncouple the checkpoints can induce unscheduled entry into mitosis. Likewise, defects of the checkpoints in several disorders permit mitotic entry even in the presence of DNA damage. Forcing cells with damaged DNA into mitosis causes severe chromosome segregation defects, including lagging chromosomes, chromosomal fragments and chromosomal bridges. The presence of these lesions in the cleavage plane is believed to abort cytokinesis. It is postulated that if cytokinesis failure is coupled with defects of the p53-dependent postmitotic checkpoint pathway, cells can enter S phase and become polyploids. Progress in the past several years has unraveled some of the underlying principles of these pathways and underscored the important role of DNA damage in polyploidization. Furthermore, polyploidization per se may also be an important determinant of sensitivity to DNA damage, thereby may offer an opportunity for novel therapies.

The Dimensions of Residential Segregation.

ERIC Educational Resources Information Center

Massey, Douglas S.; Denton, Nancy A.

1988-01-01

Evaluates 20 potential indicators of residential segregation using census data on Hispanics, Blacks, Asians, and non-Hispanic Whites in 60 U.S. metropolitan areas. Factor-analyzes the results to select a single best indicator for each of five dimensions of residential segregation. Contains 69 references and 22 statistical formulas. (SV)

Multilevel Modeling of Social Segregation

ERIC Educational Resources Information Center

Leckie, George; Pillinger, Rebecca; Jones, Kelvyn; Goldstein, Harvey

2012-01-01

The traditional approach to measuring segregation is based upon descriptive, non-model-based indices. A recently proposed alternative is multilevel modeling. The authors further develop the argument for a multilevel modeling approach by first describing and expanding upon its notable advantages, which include an ability to model segregation at a…

Integrated schools, segregated curriculum: effects of within-school segregation on adolescent health behaviors and educational aspirations.

PubMed

Walsemann, Katrina M; Bell, Bethany A

2010-09-01

We examined the extent to which within-school segregation, as measured by unevenness in the distribution of Black and White adolescents across levels of the English curriculum (advanced placement-international baccalaureate-honors, general, remedial, or no English), was associated with smoking, drinking, and educational aspirations, which previous studies found are related to school racial/ethnic composition. We analyzed data from wave 1 of the National Longitudinal Study of Adolescent Health, restricting our sample to non-Hispanic Blacks (n=2731) and Whites (n=4158) who from 1994 to 1995 attended high schools that enrolled Black and White students. White female students had higher predicted probabilities of smoking or drinking than did Black female students; the largest differences were in schools with high levels of within-school segregation. Black male students had higher predicted probabilities of high educational aspirations than did White male students in schools with low levels of within-school segregation; this association was attenuated for Black males attending schools with moderate or high levels of within-school segregation. Our results provide evidence that within-school segregation may influence both students' aspirations and their behaviors.

Natural Compounds as Anticancer Agents Targeting DNA Topoisomerases

PubMed Central

Jain, Chetan Kumar; Majumder, Hemanta Kumar; Roychoudhury, Susanta

2017-01-01

DNA topoisomerases are important cellular enzymes found in almost all types of living cells (eukaryotic and prokaryotic). These enzymes are essential for various DNA metabolic processes e.g. replication, transcription, recombination, chromosomal decatenation etc. These enzymes are important molecular drug targets and inhibitors of these enzymes are widely used as effective anticancer and antibacterial drugs. However, topoisomerase inhibitors have some therapeutic limitations and they exert serious side effects during cancer chemotherapy. Thus, development of novel anticancer topoisomerase inhibitors is necessary for improving cancer chemotherapy. Nature serves as a repertoire of structurally and chemically diverse molecules and in the recent years many DNA topoisomerase inhibitors have been identified from natural sources. The present review discusses anticancer properties and therapeutic importance of eighteen recently identified natural topoisomerase inhibitors (from the year 2009 to 2015). Structural characteristics of these novel inhibitors provide backbones for designing and developing new anticancer drugs. PMID:28503091

Evolution of prokaryote and eukaryote lines inferred from sequence evidence

NASA Technical Reports Server (NTRS)

Hunt, L. T.; George, D. G.; Yeh, L.-S.; Dayhoff, M. O.

1984-01-01

This paper describes the evolution of prokaryotes and early eukaryotes, including their symbiotic relationships, as inferred from phylogenetic trees of bacterial ferredoxin, 5S ribosomal RNA, ribulose-1,5-biphosphate carboxylase large chain, and mitochondrial cytochrome oxidase polypeptide II.

A mitosis-specific and R loop-driven ATR pathway promotes faithful chromosome segregation.

PubMed

Kabeche, Lilian; Nguyen, Hai Dang; Buisson, Rémi; Zou, Lee

2018-01-05

The ataxia telangiectasia mutated and Rad3-related (ATR) kinase is crucial for DNA damage and replication stress responses. Here, we describe an unexpected role of ATR in mitosis. Acute inhibition or degradation of ATR in mitosis induces whole-chromosome missegregation. The effect of ATR ablation is not due to altered cyclin-dependent kinase 1 (CDK1) activity, DNA damage responses, or unscheduled DNA synthesis but to loss of an ATR function at centromeres. In mitosis, ATR localizes to centromeres through Aurora A-regulated association with centromere protein F (CENP-F), allowing ATR to engage replication protein A (RPA)-coated centromeric R loops. As ATR is activated at centromeres, it stimulates Aurora B through Chk1, preventing formation of lagging chromosomes. Thus, a mitosis-specific and R loop-driven ATR pathway acts at centromeres to promote faithful chromosome segregation, revealing functions of R loops and ATR in suppressing chromosome instability. Copyright © 2018, American Association for the Advancement of Science.

Mutations reducing replication from R-loops suppress the defects of growth, chromosome segregation and DNA supercoiling in cells lacking topoisomerase I and RNase HI activity.

PubMed

Usongo, Valentine; Martel, Makisha; Balleydier, Aurélien; Drolet, Marc

2016-04-01

R-loop formation occurs when the nascent RNA hybridizes with the template DNA strand behind the RNA polymerase. R-loops affect a wide range of cellular processes and their use as origins of replication was the first function attributed to them. In Escherichia coli, R-loop formation is promoted by the ATP-dependent negative supercoiling activity of gyrase (gyrA and gyrB) and is inhibited by topoisomerase (topo) I (topA) relaxing transcription-induced negative supercoiling. RNase HI (rnhA) degrades the RNA moiety of R-loops. The depletion of RNase HI activity in topA null mutants was previously shown to lead to extensive DNA relaxation, due to DNA gyrase inhibition, and to severe growth and chromosome segregation defects that were partially corrected by overproducing topo III (topB). Here, DNA gyrase assays in crude cell extracts showed that the ATP-dependent activity (supercoiling) of gyrase but not its ATP-independent activity (relaxation) was inhibited in topA null cells lacking RNase HI. To characterize the cellular event(s) triggered by the absence of RNase HI, we performed a genetic screen for suppressors of the growth defect of topA rnhA null cells. Suppressors affecting genes in replication (holC2::aph and dnaT18::aph) nucleotide metabolism (dcd49::aph), RNA degradation (rne59::aph) and fimbriae synthesis (fimD22::aph) were found to reduce replication from R-loops and to restore supercoiling, thus pointing to a correlation between R-loop-dependent replication in topA rnhA mutants and the inhibition of gyrase activity and growth. Interestingly, the position of fimD on the E. coli chromosome corresponds to the site of one of the five main putative origins of replication from R-loops in rnhA null cells recently identified by next-generation sequencing, thus suggesting that the fimD22::aph mutation inactivated one of these origins. Furthermore, we show that topo III overproduction is unable to complement the growth defect of topA rnhA null mutants at low

Rapid mass segregation in small stellar clusters

NASA Astrophysics Data System (ADS)

Spera, Mario; Capuzzo-Dolcetta, Roberto

2017-12-01

In this paper we focus our attention on small-to-intermediate N-body systems that are, initially, distributed uniformly in space and dynamically `cool' (virial ratios Q=2T/|Ω| below ˜0.3). In this work, we study the mass segregation that emerges after the initial violent dynamical evolution. At this scope, we ran a set of high precision N-body simulations of isolated clusters by means of HiGPUs, our direct summation N-body code. After the collapse, the system shows a clear mass segregation. This (quick) mass segregation occurs in two phases: the first shows up in clumps originated by sub-fragmentation before the deep overall collapse; this segregation is partly erased during the deep collapse to re-emerge, abruptly, during the second phase, that follows the first bounce of the system. In this second stage, the proper clock to measure the rate of segregation is the dynamical time after virialization, which (for cold and cool systems) may be significantly different from the crossing time evaluated from initial conditions. This result is obtained for isolated clusters composed of stars of two different masses (in the ratio mh/ml=2), at varying their number ratio, and is confirmed also in presence of a massive central object (simulating a black hole of stellar size). Actually, in stellar systems starting their dynamical evolution from cool conditions, the fast mass segregation adds to the following, slow, secular segregation which is collisionally induced. The violent mass segregation is an effect persistent over the whole range of N (128 ≤ N ≤1,024) investigated, and is an interesting feature on the astronomical-observational side, too. The semi-steady state reached after virialization corresponds to a mass segregated distribution function rather than that of equipartition of kinetic energy per unit mass as it should result from violent relaxation.

43 CFR 2611.2 - Period of segregation.

Code of Federal Regulations, 2011 CFR

2011-10-01

... 43 Public Lands: Interior 2 2011-10-01 2011-10-01 false Period of segregation. 2611.2 Section 2611..., DEPARTMENT OF THE INTERIOR LAND RESOURCE MANAGEMENT (2000) CAREY ACT GRANTS Segregation Under the Carey Act: Procedures § 2611.2 Period of segregation. (a) The States are allowed 10 years from the date of the signing...

A cell engineering strategy to enhance supercoiled plasmid DNA production for gene therapy.

PubMed

Hassan, Sally; Keshavarz-Moore, Eli; Ward, John

2016-09-01

With the recent revival of the promise of plasmid DNA vectors in gene therapy, a novel synthetic biology approach was used to enhance the quantity, (yield), and quality of the plasmid DNA. Quality was measured by percentage supercoiling and supercoiling density, as well as improving segregational stability in fermentation. We examined the hypothesis that adding a Strong Gyrase binding Site (SGS) would increase DNA gyrase-mediated plasmid supercoiling. SGS from three different replicons, (the Mu bacteriophage and two plasmids, pSC101 and pBR322) were inserted into the plasmid, pUC57. Different sizes of these variants were transformed into E. coli DH5α, and their supercoiling properties and segregational stability measured. A 36% increase in supercoiling density was found in pUC57-SGS, but only when SGS was derived from the Mu phage and was the larger sized version of this fragment. These results were also confirmed at fermentation scale. Total percentage supercoiled monomer was maintained to 85-90%. A twofold increase in plasmid yield was also observed for pUC57-SGS in comparison to pUC57. pUC57-SGS displayed greater segregational stability than pUC57-cer and pUC57, demonstrating a further potential advantage of the SGS site. These findings should augment the potential of plasmid DNA vectors in plasmid DNA manufacture. Biotechnol. Bioeng. 2016;113: 2064-2071. © 2016 The Authors. Biotechnology and Bioengineering Published by Wiley Periodicals, Inc. © 2016 The Authors. Biotechnology and Bioengineering Published by Wiley Periodicals, Inc.

Transport analogy for segregation and granular rheology

NASA Astrophysics Data System (ADS)

Liu, Siying; McCarthy, Joseph J.

2017-08-01

Here, we show a direct connection between density-based segregation and granular rheology that can lead to insight into both problems. Our results exhibit a transition in the rate of segregation during simple shear that occurs at I ˜0.5 and mimics a coincident regime change in flow rheology. We propose scaling arguments that support a packing fraction criterion for this transition that can both explain our segregation results as well as unify existing literature studies of granular rheology. By recasting a segregation model in terms of rheological parameters, we establish an approach that not only collapses results for a wide range of conditions, but also yields a direct relationship between the coordination number z and the segregation velocity. Moreover, our approach predicts the precise location of the observed regime change or saturation. This suggests that it is possible to rationally design process operating conditions that lead to significantly lower segregation extents. These observations can have a profound impact on both the study of granular flow or mixing as well as industrial practice.

Segregation physics of a macroscale granular ratchet

NASA Astrophysics Data System (ADS)

Bhateja, Ashish; Sharma, Ishan; Singh, Jayant K.

2017-05-01

New experiments with multigrain mixtures in a laterally shaken, horizontal channel show complete axial segregation of species. The channel consists of multiple concatenated trapeziums, and superficially resembles microratchets wherein asymmetric geometries and potentials transport, and sort, randomly agitated microscopic particles. However, the physics of our macroscale granular ratchet is fundamentally different, as macroscopic segregation is gravity driven. Our observations are not explained by classical granular segregation theories either. Motivated by the experiments, extensive parallelized discrete element simulations reveal that the macroratchet differentiates grains through hierarchical bidirectional segregation over two different time scales: Grains rapidly sort vertically into horizontal bands spanning the channel's length that, subsequently, slowly separate axially, driven by strikingly gentle, average interfacial pressure gradients acting over long distances. At its maximum, the pressure gradient responsible for axial separation was due to a change in height of about two big grain diameters (d =7 mm) over a meter-long channel. The strong directional segregation achieved by the granular macroratchet has practical importance, while identifying the underlying new physics will further our understanding of granular segregation in industrial and geophysical processes.

Non-Random Inversion Landscapes in Prokaryotic Genomes Are Shaped by Heterogeneous Selection Pressures.

PubMed

Repar, Jelena; Warnecke, Tobias

2017-08-01

Inversions are a major contributor to structural genome evolution in prokaryotes. Here, using a novel alignment-based method, we systematically compare 1,651 bacterial and 98 archaeal genomes to show that inversion landscapes are frequently biased toward (symmetric) inversions around the origin-terminus axis. However, symmetric inversion bias is not a universal feature of prokaryotic genome evolution but varies considerably across clades. At the extremes, inversion landscapes in Bacillus-Clostridium and Actinobacteria are dominated by symmetric inversions, while there is little or no systematic bias favoring symmetric rearrangements in archaea with a single origin of replication. Within clades, we find strong but clade-specific relationships between symmetric inversion bias and different features of adaptive genome architecture, including the distance of essential genes to the origin of replication and the preferential localization of genes on the leading strand. We suggest that heterogeneous selection pressures have converged to produce similar patterns of structural genome evolution across prokaryotes. © The Author 2017. Published by Oxford University Press on behalf of the Society for Molecular Biology and Evolution.

The Big Disconnect between Segregation and Integration

ERIC Educational Resources Information Center

Verdun, Vincene

2005-01-01

The hearts and minds of the American people have been won over on the issue of segregation. However, the dilemma is that while an overwhelming majority of Americans would cringe at the idea of a racially segregated America, America remains racially segregated and racial equality is more ideal than real. Even though there is almost no legal…

The actin-like MreB cytoskeleton organizes viral DNA replication in bacteria.

PubMed

Muñoz-Espín, Daniel; Daniel, Richard; Kawai, Yoshikazu; Carballido-López, Rut; Castilla-Llorente, Virginia; Errington, Jeff; Meijer, Wilfried J J; Salas, Margarita

2009-08-11

Little is known about the organization or proteins involved in membrane-associated replication of prokaryotic genomes. Here we show that the actin-like MreB cytoskeleton of the distantly related bacteria Escherichia coli and Bacillus subtilis is required for efficient viral DNA replication. Detailed analyses of B. subtilis phage ϕ29 showed that the MreB cytoskeleton plays a crucial role in organizing phage DNA replication at the membrane. Thus, phage double-stranded DNA and components of the ϕ29 replication machinery localize in peripheral helix-like structures in a cytoskeleton-dependent way. Importantly, we show that MreB interacts directly with the ϕ29 membrane-protein p16.7, responsible for attaching viral DNA at the cell membrane. Altogether, the results reveal another function for the MreB cytoskeleton and describe a mechanism by which viral DNA replication is organized at the bacterial membrane.

Segregated Systems of Human Brain Networks.

PubMed

Wig, Gagan S

2017-12-01

The organization of the brain network enables its function. Evaluation of this organization has revealed that large-scale brain networks consist of multiple segregated subnetworks of interacting brain areas. Descriptions of resting-state network architecture have provided clues for understanding the functional significance of these segregated subnetworks, many of which correspond to distinct brain systems. The present report synthesizes accumulating evidence to reveal how maintaining segregated brain systems renders the human brain network functionally specialized, adaptable to task demands, and largely resilient following focal brain damage. The organizational properties that support system segregation are harmonious with the properties that promote integration across the network, but confer unique and important features to the brain network that are central to its function and behavior. Copyright © 2017 Elsevier Ltd. All rights reserved.

DNA methylome signature in rheumatoid arthritis.

PubMed

Nakano, Kazuhisa; Whitaker, John W; Boyle, David L; Wang, Wei; Firestein, Gary S

2013-01-01

Epigenetics can influence disease susceptibility and severity. While DNA methylation of individual genes has been explored in autoimmunity, no unbiased systematic analyses have been reported. Therefore, a genome-wide evaluation of DNA methylation loci in fibroblast-like synoviocytes (FLS) isolated from the site of disease in rheumatoid arthritis (RA) was performed. Genomic DNA was isolated from six RA and five osteoarthritis (OA) FLS lines and evaluated using the Illumina HumanMethylation450 chip. Cluster analysis of data was performed and corrected using Benjamini-Hochberg adjustment for multiple comparisons. Methylation was confirmed by pyrosequencing and gene expression was determined by qPCR. Pathway analysis was performed using the Kyoto Encyclopedia of Genes and Genomes. RA and control FLS segregated based on DNA methylation, with 1859 differentially methylated loci. Hypomethylated loci were identified in key genes relevant to RA, such as CHI3L1, CASP1, STAT3, MAP3K5, MEFV and WISP3. Hypermethylation was also observed, including TGFBR2 and FOXO1. Hypomethylation of individual genes was associated with increased gene expression. Grouped analysis identified 207 hypermethylated or hypomethylated genes with multiple differentially methylated loci, including COL1A1, MEFV and TNF. Hypomethylation was increased in multiple pathways related to cell migration, including focal adhesion, cell adhesion, transendothelial migration and extracellular matrix interactions. Confirmatory studies with OA and normal FLS also demonstrated segregation of RA from control FLS based on methylation pattern. Differentially methylated genes could alter FLS gene expression and contribute to the pathogenesis of RA. DNA methylation of critical genes suggests that RA FLS are imprinted and implicate epigenetic contributions to inflammatory arthritis.

Epigenetic Segregation of Microbial Genomes from Complex Samples Using Restriction Endonucleases HpaII and McrB.

PubMed

Liu, Guohong; Weston, Christopher Q; Pham, Long K; Waltz, Shannon; Barnes, Helen; King, Paula; Sphar, Dan; Yamamoto, Robert T; Forsyth, R Allyn

2016-01-01

We describe continuing work to develop restriction endonucleases as tools to enrich targeted genomes of interest from diverse populations. Two approaches were developed in parallel to segregate genomic DNA based on cytosine methylation. First, the methyl-sensitive endonuclease HpaII was used to bind non-CG methylated DNA. Second, a truncated fragment of McrB was used to bind CpG methylated DNA. Enrichment levels of microbial genomes can exceed 100-fold with HpaII allowing improved genomic detection and coverage of otherwise trace microbial genomes from sputum. Additionally, we observe interesting enrichment results that correlate with the methylation states not only of bacteria, but of fungi, viruses, a protist and plants. The methods presented here offer promise for testing biological samples for pathogens and global analysis of population methylomes.

Liquid-phase electroepitaxy - Dopant segregation

NASA Technical Reports Server (NTRS)

Lagowski, J.; Jastrzebski, L.; Gatos, H. C.

1980-01-01

A theoretical model is presented which accounts for the dopant segregation in liquid-phase electroepitaxy in terms of dopant transport in the liquid phase (by electromigration and diffusion), the growth velocity, and the Peltier effect at the substrate-solution interface. The contribution of dopant electromigration to the magnitude of the effective segregation coefficient is dominant in the absence of convection; the contribution of the Peltier effect becomes significant only in the presence of pronounced convection. Quantitative expressions which relate the segregation coefficient to the growth parameters also permit the determination of the diffusion constant and electromigration mobility of the dopant in the liquid phase. The model was found to be in good agreement with the measured segregation characteristics of Sn in the electroepitaxial growth of GaAs from Ga-As solutions. For Sn in Ga-As solution at 900 C the diffusion constant was found to be 4 x 10 to the -5 sq cm/s and the electromigration velocity (toward the substrate with a positive polarity 2 x 10 to the -5 cm/s current density of 10 A/sq cm.

Prokaryotic Argonaute proteins: novel genome-editing tools?

PubMed

Hegge, Jorrit W; Swarts, Daan C; van der Oost, John

2018-01-01

Argonaute proteins constitute a highly diverse family of nucleic acid-guided proteins. They were first discovered in eukaryotes as key proteins in RNA interference systems, but homologous prokaryotic Argonaute proteins (pAgos) have also been found in archaea and bacteria. In this Progress article, we focus on long pAgo variants, a class of pAgos that are involved in nucleic acid-guided host defence against invading nucleic acids, and discuss the potential of pAgos in genome editing.

Multifunctional centromere binding factor 1 is essential for chromosome segregation in the human pathogenic yeast Candida glabrata.

PubMed

Stoyan, T; Gloeckner, G; Diekmann, S; Carbon, J

2001-08-01

The CBF1 (centromere binding factor 1) gene of Candida glabrata was cloned by functional complementation of the methionine biosynthesis defect of a Saccharomyces cerevisiae cbf1 deletion mutant. The C. glabrata-coded protein, CgCbf1, contains a basic-helix-loop-helix leucine zipper domain and has features similar to those of other budding yeast Cbf1 proteins. CgCbf1p binds in vitro to the centromere DNA element I (CDEI) sequence GTCACATG with high affinity (0.9 x 10(9) M(-1)). Bandshift experiments revealed a pattern of protein-DNA complexes on CgCEN DNA different from that known for S. cerevisiae. We examined the effect of altering the CDEI binding site on CEN plasmid segregation, using a newly developed colony-sectoring assay. Internal deletion of the CDEI binding site led only to a fivefold increase in rates of plasmid loss, indicating that direct binding of Cbf1p to the centromere DNA is not required for full function. Additional deletion of sequences to the left of CDEI, however, led to a 70-fold increase in plasmid loss rates. Deletion of the CBF1 gene proved to be lethal in C. glabrata. C. glabrata cells containing the CBF1 gene under the influence of a shutdown promoter (tetO-ScHOP) arrested their growth after 5 h of cultivation in the presence of the reactive drug doxycycline. DAPI (4',6'-diamidino-2-phenylindole) staining of the arrested cells revealed a significant increase in the number of large-budded cells with single nuclei, 2C DNA content, and short spindles, indicating a defect in the G(2)/M transition of the cell cycle. Thus, we conclude that Cbf1p is required for chromosome segregation in C. glabrata.

Reduced prokaryotic heterotrophic production at in situ pressure conditions in the dark ocean

NASA Astrophysics Data System (ADS)

Amano-Sato, Chie; Sintes, Eva; Reinthaler, Thomas; Utsumi, Motoo; Herndl, Gerhard J.

2017-04-01

Prokaryotic heterotrophic production (PHP) is a key process in the ocean's biological carbon cycle. About 50% of the oceanic PHP takes place in the dark ocean characterized by low temperature and high hydrostatic pressure, which increases by 1 MPa (10 atm) every 100 m depth. However, rate measurements of PHP are usually performed under atmospheric pressure conditions. Yet, the difference in pressure conditions and the handling of the samples on board may introduce biases in the PHP measurements. To determine PHP at in situ conditions, we developed an in situ microbial incubator (ISMI) designed to autonomously sample and incubate seawater down to a depth of 4000 m. Natural prokaryotic communities from the North Atlantic and Pacific Oceans were incubated in the ISMI with 5 nM 3H-leucine at different depths ranging between 10 and 3200 m. For comparison, atmospheric pressure incubations at in situ temperature were also conducted. PHP and single cell activity assessed by microautoradiography combined with catalyzed reporter deposition fluorescence in situ hybridization (MICRO-CARD-FISH) were determined. PHP obtained under in situ pressure conditions was generally lower than under atmospheric pressure conditions, suggesting that incubation under atmospheric pressure on board stimulates activity of dark ocean prokaryotes. The ratio between the bulk PHP obtained under in situ and under atmospheric pressure conditions decreased with depth. Moreover, MICRO-CARD-FISH revealed that some specific prokaryotic groups are apparently more affected by the hydrostatic pressure condition than others. Our results suggest that PHP in the dark ocean might be lower than assumed based on measurements under surface pressure conditions.

The First Prokaryotic Trehalose Synthase Complex Identified in the Hyperthermophilic Crenarchaeon Thermoproteus tenax

PubMed Central

Bräsen, Christopher; Hensel, Reinhard; Lupas, Andrei N.; Brinkmann, Henner; Siebers, Bettina

2013-01-01

The role of the disaccharide trehalose, its biosynthesis pathways and their regulation in Archaea are still ambiguous. In Thermoproteus tenax a fused trehalose-6-phosphate synthase/phosphatase (TPSP), consisting of an N-terminal trehalose-6-phosphate synthase (TPS) and a C-terminal trehalose-6-phosphate phosphatase (TPP) domain, was identified. The tpsp gene is organized in an operon with a putative glycosyltransferase (GT) and a putative mechanosensitive channel (MSC). The T. tenax TPSP exhibits high phosphatase activity, but requires activation by the co-expressed GT for bifunctional synthase-phosphatase activity. The GT mediated activation of TPS activity relies on the fusion of both, TPS and TPP domain, in the TPSP enzyme. Activation is mediated by complex-formation in vivo as indicated by yeast two-hybrid and crude extract analysis. In combination with first evidence for MSC activity the results suggest a sophisticated stress response involving TPSP, GT and MSC in T. tenax and probably in other Thermoproteales species. The monophyletic prokaryotic TPSP proteins likely originated via a single fusion event in the Bacteroidetes with subsequent horizontal gene transfers to other Bacteria and Archaea. Furthermore, evidence for the origin of eukaryotic TPSP fusions via HGT from prokaryotes and therefore a monophyletic origin of eukaryotic and prokaryotic fused TPSPs is presented. This is the first report of a prokaryotic, archaeal trehalose synthase complex exhibiting a much more simple composition than the eukaryotic complex described in yeast. Thus, complex formation and a complex-associated regulatory potential might represent a more general feature of trehalose synthesizing proteins. PMID:23626675

Exploring DNA variant segregation types in pooled genome sequencing enables effective mapping of weeping trait in Malus

USDA-ARS?s Scientific Manuscript database

In recent years, next generation sequencing (NGS) based bulked segregant analysis (BSA) has become a powerful approach for allele discovery in non-model plant species. However, challenges remain, particular for out-crossing species with complex genomes. Here, the genetic control of a weeping bran...

Factors shaping workplace segregation between natives and immigrants.

PubMed

Strömgren, Magnus; Tammaru, Tiit; Danzer, Alexander M; van Ham, Maarten; Marcińczak, Szymon; Stjernström, Olof; Lindgren, Urban

2014-04-01

Research on segregation of immigrant groups is increasingly turning its attention from residential areas toward other important places, such as the workplace, where immigrants can meet and interact with members of the native population. This article examines workplace segregation of immigrants. We use longitudinal, georeferenced Swedish population register data, which enables us to observe all immigrants in Sweden for the period 1990-2005 on an annual basis. We compare estimates from ordinary least squares with fixed-effects regressions to quantify the extent of immigrants' self-selection into specific workplaces, neighborhoods, and partnerships, which may bias more naïve ordinary least squares results. In line with previous research, we find lower levels of workplace segregation than residential segregation. The main finding is that low levels of residential segregation reduce workplace segregation, even after we take into account intermarriage with natives as well as unobserved characteristics of immigrants' such as willingness and ability to integrate into the host society. Being intermarried with a native reduces workplace segregation for immigrant men but not for immigrant women.

Zinc-binding Domain of the Bacteriophage T7 DNA Primase Modulates Binding to the DNA Template*

PubMed Central

Lee, Seung-Joo; Zhu, Bin; Akabayov, Barak; Richardson, Charles C.

2012-01-01

The zinc-binding domain (ZBD) of prokaryotic DNA primases has been postulated to be crucial for recognition of specific sequences in the single-stranded DNA template. To determine the molecular basis for this role in recognition, we carried out homolog-scanning mutagenesis of the zinc-binding domain of DNA primase of bacteriophage T7 using a bacterial homolog from Geobacillus stearothermophilus. The ability of T7 DNA primase to catalyze template-directed oligoribonucleotide synthesis is eliminated by substitution of any five-amino acid residue-long segment within the ZBD. The most significant defect occurs upon substitution of a region (Pro-16 to Cys-20) spanning two cysteines that coordinate the zinc ion. The role of this region in primase function was further investigated by generating a protein library composed of multiple amino acid substitutions for Pro-16, Asp-18, and Asn-19 followed by genetic screening for functional proteins. Examination of proteins selected from the screening reveals no change in sequence-specific recognition. However, the more positively charged residues in the region facilitate DNA binding, leading to more efficient oligoribonucleotide synthesis on short templates. The results suggest that the zinc-binding mode alone is not responsible for sequence recognition, but rather its interaction with the RNA polymerase domain is critical for DNA binding and for sequence recognition. Consequently, any alteration in the ZBD that disturbs its conformation leads to loss of DNA-dependent oligoribonucleotide synthesis. PMID:23024359

The Segregation Academy and the Law

ERIC Educational Resources Information Center

Champagne, Anthony M.

1973-01-01

A case study of one private school which functions as a segregation academy'' was done in order to learn more about what segregation academies are, how they operate, and how they relate to the community. (Author/JM)

Comparison of Prokaryotic Diversity in Cold, Oligotrophic Remote Lakes of Chilean Patagonia.

PubMed

Aguayo, Paulina; González, Paulina; Campos, Víctor; Maugeri, Teresa L; Papale, Maria; Gugliandolo, Concetta; Martinez, Miguel A

2017-05-01

The prokaryotic abundance and diversity in three cold, oligotrophic Patagonian lakes (Témpanos, Las Torres and Mercedes) in the northern region Aysén (Chile) were compared in winter and summer using 16S rRNA fluorescence in situ hybridization and PCR-denaturing gradient gel electrophoresis technique. Prokaryotic abundances, numerically dominated by Bacteria, were quite similar in the three lakes, but higher in sediments than in waters, and they were also higher in summer than in winter. The relative contribution of Archaea was greater in waters than in sediments, and in winter rather than in summer. Despite the phylogenetic analysis indicated that most sequences were affiliated to a few taxonomic groups, mainly referred to Proteobacteria (consisting of Beta-, Alpha- and Gammaproteobacteria) and Euryarchaeota (mainly related to uncultured methanogens), their relative abundances differed in each sample, resulting in different bacterial and archaeal assemblages. In winter, the abundance of the dominant bacterial phylotypes were mainly regulated by the increasing levels of total organic carbon in waters. Archaeal abundance and richness appeared mostly influenced by pH in winter and total nitrogen content in summer. The prokaryotic community composition at Témpanos lake, located most northerly and closer to a glacier, greatly differed in respect to the other two lakes. In this lake was detected the highest bacterial diversity, being Betaproteobacteria the most abundant group, whereas Alphaproteobacteria were distinctive of Mercedes. Archaeal community associated with sediments was mainly represent by members related to the order of Methanosarcinales at Mercedes and Las Torres lakes, and by Crenarchaeota at Témpanos lake. Our results indicate that the proximity to the glacier and the seasonality shape the composition of the prokaryotic communities in these remote lakes. These results may be used as baseline information to follow the microbial community responses to

The Influence of pH on Prokaryotic Cell Size and Temperature

NASA Astrophysics Data System (ADS)

Sundararajan, D.; Gutierrez, F.; Heim, N. A.; Payne, J.

2015-12-01

The pH of a habitat is essential to an organism's growth and success in its environment. Although most organisms maintain a neutral internal pH, their environmental pH can vary greatly. However, little research has been done concerning an organism's environmental pH across a wide range of taxa. We studied pH tolerance in prokaryotes and its relationship with biovolume, taxonomic classification, and ideal temperature. We had three hypotheses: pH and temperature are not correlated; pH tolerance is similar within taxonomic groups; and extremophiles have small cell sizes. To test these hypotheses, we used pH, size, and taxonomic data from The Prokaryotes. We found that the mean optimum external pH was neutral for prokaryotes as a whole and when divided by domain, phylum, and class. Using ANOVA to test for pH within and among group variances, we found that variation of pH in domains, phyla, classes, and families was greater than between them. pH and size did not show much of a correlation, except that the largest and smallest sized prokaryotes had nearly neutral pH. This seems significant because extremophiles need to divert more of their energy from growth to maintain a neutral internal pH. Acidophiles showed a larger range of optimum pH values than alkaliphiles. A similar result was seen with the minimum and maximum pH values of acidophiles and alkaliphiles. While acidophiles were spread out and had some alkaline maximum values, alkaliphiles had smaller ranges, and unlike some acidophiles that had pH minimums close to zero, alkaliphile pH maximums did not go beyond a pH of 12. No statistically significant differences were found between sizes of acidophiles and alkaliphiles. However, optimum temperatures of acidophiles and alkaliphiles did have a statistically significant difference. pH and temperature had a negative correlation. Therefore, pH seems to have a correlation with cell size, temperature, and taxonomy to some extent.

Sequence conservation from human to prokaryotes of Surf1, a protein involved in cytochrome c oxidase assembly, deficient in Leigh syndrome.

PubMed

Poyau, A; Buchet, K; Godinot, C

1999-12-03

The human SURF1 gene encoding a protein involved in cytochrome c oxidase (COX) assembly, is mutated in most patients presenting Leigh syndrome associated with COX deficiency. Proteins homologous to the human Surf1 have been identified in nine eukaryotes and six prokaryotes using database alignment tools, structure prediction and/or cDNA sequencing. Their sequence comparison revealed a remarkable Surf1 conservation during evolution and put forward at least four highly conserved domains that should be essential for Surf1 function. In Paracoccus denitrificans, the Surf1 homologue is found in the quinol oxidase operon, suggesting that Surf1 is associated with a primitive quinol oxidase which belongs to the same superfamily as cytochrome oxidase.

Marker genes that are less conserved in their sequences are useful for predicting genome-wide similarity levels between closely related prokaryotic strains

DOE Office of Scientific and Technical Information (OSTI.GOV)

Lan, Yemin; Rosen, Gail; Hershberg, Ruth

The 16s rRNA gene is so far the most widely used marker for taxonomical classification and separation of prokaryotes. Since it is universally conserved among prokaryotes, it is possible to use this gene to classify a broad range of prokaryotic organisms. At the same time, it has often been noted that the 16s rRNA gene is too conserved to separate between prokaryotes at finer taxonomic levels. In this paper, we examine how well levels of similarity of 16s rRNA and 73 additional universal or nearly universal marker genes correlate with genome-wide levels of gene sequence similarity. We demonstrate that themore » percent identity of 16s rRNA predicts genome-wide levels of similarity very well for distantly related prokaryotes, but not for closely related ones. In closely related prokaryotes, we find that there are many other marker genes for which levels of similarity are much more predictive of genome-wide levels of gene sequence similarity. Finally, we show that the identities of the markers that are most useful for predicting genome-wide levels of similarity within closely related prokaryotic lineages vary greatly between lineages. However, the most useful markers are always those that are least conserved in their sequences within each lineage. In conclusion, our results show that by choosing markers that are less conserved in their sequences within a lineage of interest, it is possible to better predict genome-wide gene sequence similarity between closely related prokaryotes than is possible using the 16s rRNA gene. We point readers towards a database we have created (POGO-DB) that can be used to easily establish which markers show lowest levels of sequence conservation within different prokaryotic lineages.« less

Marker genes that are less conserved in their sequences are useful for predicting genome-wide similarity levels between closely related prokaryotic strains

DOE PAGES

Lan, Yemin; Rosen, Gail; Hershberg, Ruth

2016-05-03

The 16s rRNA gene is so far the most widely used marker for taxonomical classification and separation of prokaryotes. Since it is universally conserved among prokaryotes, it is possible to use this gene to classify a broad range of prokaryotic organisms. At the same time, it has often been noted that the 16s rRNA gene is too conserved to separate between prokaryotes at finer taxonomic levels. In this paper, we examine how well levels of similarity of 16s rRNA and 73 additional universal or nearly universal marker genes correlate with genome-wide levels of gene sequence similarity. We demonstrate that themore » percent identity of 16s rRNA predicts genome-wide levels of similarity very well for distantly related prokaryotes, but not for closely related ones. In closely related prokaryotes, we find that there are many other marker genes for which levels of similarity are much more predictive of genome-wide levels of gene sequence similarity. Finally, we show that the identities of the markers that are most useful for predicting genome-wide levels of similarity within closely related prokaryotic lineages vary greatly between lineages. However, the most useful markers are always those that are least conserved in their sequences within each lineage. In conclusion, our results show that by choosing markers that are less conserved in their sequences within a lineage of interest, it is possible to better predict genome-wide gene sequence similarity between closely related prokaryotes than is possible using the 16s rRNA gene. We point readers towards a database we have created (POGO-DB) that can be used to easily establish which markers show lowest levels of sequence conservation within different prokaryotic lineages.« less

49 CFR 176.146 - Segregation from non-hazardous materials.

Code of Federal Regulations, 2010 CFR

2010-10-01

... 49 Transportation 2 2010-10-01 2010-10-01 false Segregation from non-hazardous materials. 176.146... VESSEL Detailed Requirements for Class 1 (Explosive) Materials Segregation § 176.146 Segregation from non... for “away from” segregation apply. (2) An explosive substance or article which has a secondary...

49 CFR 176.146 - Segregation from non-hazardous materials.

Code of Federal Regulations, 2011 CFR

2011-10-01

... 49 Transportation 2 2011-10-01 2011-10-01 false Segregation from non-hazardous materials. 176.146... VESSEL Detailed Requirements for Class 1 (Explosive) Materials Segregation § 176.146 Segregation from non... for “away from” segregation apply. (2) An explosive substance or article which has a secondary...

Prokaryotic silicon utilizing microorganisms in the biosphere

NASA Astrophysics Data System (ADS)

Gupta, D.; Das, S.

2012-12-01

Although a little study has been done to determine the silicon utilizing prokaryotes, our previous experiments indicated that almost all Gram-positive bacteria are silicon utilizing; one of them, Streptococci survived exposure on the lunar surface for a long period in experiment done by others. Our initial experiments with these Gram positive microorganisms showed that there were limited growths of these microorganisms on carbon free silicate medium probably with the help of some carry over carbon and nitrogen during cultivation procedures. However, increase in growth rate after repeated subcultures could not be explained at present. The main groups of prokaryotes which were found silicon utilizing microorganisms were Mycobacterium, Bacillus, Nocardia, Streptomyces, Staphylococcus, Streptococcus, Lactobacillus, and Clostridium. In a another previous study by us when silicon level was studied in such grown up cells on carbon "free" silicate medium by electron prove microanalyser, it was found that silicon in cells grown on carbon "free" silicate medium was much higher (24.9%) than those grown on conventional carbon based medium (0.84%). However, these initial findings are encouraging for our future application of this group of organisms on extraterrestrial surfaces for artificial micro-ecosystem formation. It was found that when electropositive elements are less in extraterrestrial situation, then polymerization of silicon-oxygen profusion may occur easily, particularly in carbon and nitrogen paucity in the rocky worlds of the Universe.

Detecting uber-operons in prokaryotic genomes

PubMed Central

Che, Dongsheng; Li, Guojun; Mao, Fenglou; Wu, Hongwei; Xu, Ying

2006-01-01

We present a study on computational identification of uber-operons in a prokaryotic genome, each of which represents a group of operons that are evolutionarily or functionally associated through operons in other (reference) genomes. Uber-operons represent a rich set of footprints of operon evolution, whose full utilization could lead to new and more powerful tools for elucidation of biological pathways and networks than what operons have provided, and a better understanding of prokaryotic genome structures and evolution. Our prediction algorithm predicts uber-operons through identifying groups of functionally or transcriptionally related operons, whose gene sets are conserved across the target and multiple reference genomes. Using this algorithm, we have predicted uber-operons for each of a group of 91 genomes, using the other 90 genomes as references. In particular, we predicted 158 uber-operons in Escherichia coli K12 covering 1830 genes, and found that many of the uber-operons correspond to parts of known regulons or biological pathways or are involved in highly related biological processes based on their Gene Ontology (GO) assignments. For some of the predicted uber-operons that are not parts of known regulons or pathways, our analyses indicate that their genes are highly likely to work together in the same biological processes, suggesting the possibility of new regulons and pathways. We believe that our uber-operon prediction provides a highly useful capability and a rich information source for elucidation of complex biological processes, such as pathways in microbes. All the prediction results are available at our Uber-Operon Database: , the first of its kind. PMID:16682449

A Dual-Intein Autoprocessing Domain that Directs Synchronized Protein Co-Expression in Both Prokaryotes and Eukaryotes

PubMed Central

Zhang, Bei; Rapolu, Madhusudhan; Liang, Zhibin; Han, Zhenlin; Williams, Philip G.; Su, Wei Wen

2015-01-01

Being able to coordinate co-expression of multiple proteins is necessary for a variety of important applications such as assembly of protein complexes, trait stacking, and metabolic engineering. Currently only few options are available for multiple recombinant protein co-expression, and most of them are not applicable to both prokaryotic and eukaryotic hosts. Here, we report a new polyprotein vector system that is based on a pair of self-excising mini-inteins fused in tandem, termed the dual-intein (DI) domain, to achieve synchronized co-expression of multiple proteins. The DI domain comprises an Ssp DnaE mini-intein N159A mutant and an Ssp DnaB mini-intein C1A mutant connected in tandem by a peptide linker to mediate efficient release of the flanking proteins via autocatalytic cleavage. Essentially complete release of constituent proteins, GFP and RFP (mCherry), from a polyprotein precursor, in bacterial, mammalian, and plant hosts was demonstrated. In addition, successful co-expression of GFP with chloramphenicol acetyltransferase, and thioredoxin with RFP, respectively, further substantiates the general applicability of the DI polyprotein system. Collectively, our results demonstrate the DI-based polyprotein technology as a highly valuable addition to the molecular toolbox for multi-protein co-expression which finds vast applications in biotechnology, biosciences, and biomedicine. PMID:25712612

Proteogenomic Analysis of Polymorphisms and Gene Annotation Divergences in Prokaryotes using a Clustered Mass Spectrometry-Friendly Database*

PubMed Central

de Souza, Gustavo A.; Arntzen, Magnus Ø.; Fortuin, Suereta; Schürch, Anita C.; Målen, Hiwa; McEvoy, Christopher R. E.; van Soolingen, Dick; Thiede, Bernd; Warren, Robin M.; Wiker, Harald G.

2011-01-01

Precise annotation of genes or open reading frames is still a difficult task that results in divergence even for data generated from the same genomic sequence. This has an impact in further proteomic studies, and also compromises the characterization of clinical isolates with many specific genetic variations that may not be represented in the selected database. We recently developed software called multistrain mass spectrometry prokaryotic database builder (MSMSpdbb) that can merge protein databases from several sources and be applied on any prokaryotic organism, in a proteomic-friendly approach. We generated a database for the Mycobacterium tuberculosis complex (using three strains of Mycobacterium bovis and five of M. tuberculosis), and analyzed data collected from two laboratory strains and two clinical isolates of M. tuberculosis. We identified 2561 proteins, of which 24 were present in M. tuberculosis H37Rv samples, but not annotated in the M. tuberculosis H37Rv genome. We were also able to identify 280 nonsynonymous single amino acid polymorphisms and confirm 367 translational start sites. As a proof of concept we applied the database to whole-genome DNA sequencing data of one of the clinical isolates, which allowed the validation of 116 predicted single amino acid polymorphisms and the annotation of 131 N-terminal start sites. Moreover we identified regions not present in the original M. tuberculosis H37Rv sequence, indicating strain divergence or errors in the reference sequence. In conclusion, we demonstrated the potential of using a merged database to better characterize laboratory or clinical bacterial strains. PMID:21030493

Information dynamics in living systems: prokaryotes, eukaryotes, and cancer.

PubMed

Frieden, B Roy; Gatenby, Robert A

2011-01-01

Living systems use information and energy to maintain stable entropy while far from thermodynamic equilibrium. The underlying first principles have not been established. We propose that stable entropy in living systems, in the absence of thermodynamic equilibrium, requires an information extremum (maximum or minimum), which is invariant to first order perturbations. Proliferation and death represent key feedback mechanisms that promote stability even in a non-equilibrium state. A system moves to low or high information depending on its energy status, as the benefit of information in maintaining and increasing order is balanced against its energy cost. Prokaryotes, which lack specialized energy-producing organelles (mitochondria), are energy-limited and constrained to an information minimum. Acquisition of mitochondria is viewed as a critical evolutionary step that, by allowing eukaryotes to achieve a sufficiently high energy state, permitted a phase transition to an information maximum. This state, in contrast to the prokaryote minima, allowed evolution of complex, multicellular organisms. A special case is a malignant cell, which is modeled as a phase transition from a maximum to minimum information state. The minimum leads to a predicted power-law governing the in situ growth that is confirmed by studies measuring growth of small breast cancers. We find living systems achieve a stable entropic state by maintaining an extreme level of information. The evolutionary divergence of prokaryotes and eukaryotes resulted from acquisition of specialized energy organelles that allowed transition from information minima to maxima, respectively. Carcinogenesis represents a reverse transition: of an information maximum to minimum. The progressive information loss is evident in accumulating mutations, disordered morphology, and functional decline characteristics of human cancers. The findings suggest energy restriction is a critical first step that triggers the genetic

Ocean acidification effect on prokaryotic metabolism tested in two diverse trophic regimes in the Mediterranean Sea

NASA Astrophysics Data System (ADS)

Celussi, Mauro; Malfatti, Francesca; Annalisa, Franzo; Gazeau, Frédéric; Giannakourou, Antonia; Pitta, Paraskevi; Tsiola, Anastasia; Del Negro, Paola

2017-02-01

Notwithstanding the increasing amount of researches on the effect of ocean acidification (OA) on marine ecosystems, no consent has emerged on its consequences on many prokaryote-mediated processes. Two mesocosm experiments were performed in coastal Mediterranean areas with different trophic status: the summer oligotrophic Bay of Calvi (BC, Corsica, France) and the winter mesotrophic Bay of Villefranche (BV, France). During these experiments, nine enclosures (∼54 m3) were deployed: 3 unamended controls and 6 elevated CO2, following a gradient up to 1250 μatm. We present results involving free-living viral and prokaryotic standing stocks, bacterial carbon production, abundance of highly active cells (CTC+), and degradation processes (beta-glucosidase, chitinase, leucine-aminopeptidase, lipase and alkaline phosphatase activities). The experiments revealed clear differences in the response of the two prokaryotic communities to CO2 manipulation. Only abundances of heterotrophic prokaryotes, viruses and lipase activity were not affected by CO2 manipulation at both locations. On the contrary, the percent of CTC+ was positively correlated to CO2 only in BC, concomitantly to a bulk reduction of [3H]-leucine uptake. The other tested parameters showed a different response at the two sites suggesting that the trophic regime of the systems plays a fundamental role on the effect of OA on prokaryotes through indirect modifications of the available substrate. Modified degradation rates may affect considerably the export of organic matter to the seafloor and thus ecosystem functioning within the water column. Our results highlight the need to further analyse the consequences of OA in oligotrophic ecosystems with particular focus on dissolved organic matter.

PSP: rapid identification of orthologous coding genes under positive selection across multiple closely related prokaryotic genomes.

PubMed

Su, Fei; Ou, Hong-Yu; Tao, Fei; Tang, Hongzhi; Xu, Ping

2013-12-27

With genomic sequences of many closely related bacterial strains made available by deep sequencing, it is now possible to investigate trends in prokaryotic microevolution. Positive selection is a sub-process of microevolution, in which a particular mutation is favored, causing the allele frequency to continuously shift in one direction. Wide scanning of prokaryotic genomes has shown that positive selection at the molecular level is much more frequent than expected. Genes with significant positive selection may play key roles in bacterial adaption to different environmental pressures. However, selection pressure analyses are computationally intensive and awkward to configure. Here we describe an open access web server, which is designated as PSP (Positive Selection analysis for Prokaryotic genomes) for performing evolutionary analysis on orthologous coding genes, specially designed for rapid comparison of dozens of closely related prokaryotic genomes. Remarkably, PSP facilitates functional exploration at the multiple levels by assignments and enrichments of KO, GO or COG terms. To illustrate this user-friendly tool, we analyzed Escherichia coli and Bacillus cereus genomes and found that several genes, which play key roles in human infection and antibiotic resistance, show significant evidence of positive selection. PSP is freely available to all users without any login requirement at: http://db-mml.sjtu.edu.cn/PSP/. PSP ultimately allows researchers to do genome-scale analysis for evolutionary selection across multiple prokaryotic genomes rapidly and easily, and identify the genes undergoing positive selection, which may play key roles in the interactions of host-pathogen and/or environmental adaptation.

41 CFR 109-1.5106 - Segregation of personal property.

Code of Federal Regulations, 2011 CFR

2011-01-01

... 41 Public Contracts and Property Management 3 2011-01-01 2011-01-01 false Segregation of personal...-INTRODUCTION 1.51-Personal Property Management Standards and Practices § 109-1.5106 Segregation of personal...) The segregation of the property would materially hinder the progress of the work (i.e., segregation is...

41 CFR 109-1.5106 - Segregation of personal property.

Code of Federal Regulations, 2010 CFR

2010-07-01

... 41 Public Contracts and Property Management 3 2010-07-01 2010-07-01 false Segregation of personal...-INTRODUCTION 1.51-Personal Property Management Standards and Practices § 109-1.5106 Segregation of personal...) The segregation of the property would materially hinder the progress of the work (i.e., segregation is...

Dynamics of DNA replication during premeiosis and early meiosis in wheat.

PubMed

Rey, María-Dolores; Prieto, Pilar

2014-01-01

Meiosis is a specialised cell division that involves chromosome replication, two rounds of chromosome segregation and results in the formation of the gametes. Meiotic DNA replication generally precedes chromosome pairing, recombination and synapsis in sexually developing eukaryotes. In this work, replication has been studied during premeiosis and early meiosis in wheat using flow cytometry, which has allowed the quantification of the amount of DNA in wheat anther in each phase of the cell cycle during premeiosis and each stage of early meiosis. Flow cytometry has been revealed as a suitable and user-friendly tool to detect and quantify DNA replication during early meiosis in wheat. Chromosome replication was detected in wheat during premeiosis and early meiosis until the stage of pachytene, when chromosomes are associated in pairs to further recombine and correctly segregate in the gametes. In addition, the effect of the Ph1 locus, which controls chromosome pairing and affects replication in wheat, was also studied by flow cytometry. Here we showed that the Ph1 locus plays an important role on the length of meiotic DNA replication in wheat, particularly affecting the rate of replication during early meiosis in wheat.

Dynamics of DNA Replication during Premeiosis and Early Meiosis in Wheat

PubMed Central

Rey, María-Dolores; Prieto, Pilar

2014-01-01

Meiosis is a specialised cell division that involves chromosome replication, two rounds of chromosome segregation and results in the formation of the gametes. Meiotic DNA replication generally precedes chromosome pairing, recombination and synapsis in sexually developing eukaryotes. In this work, replication has been studied during premeiosis and early meiosis in wheat using flow cytometry, which has allowed the quantification of the amount of DNA in wheat anther in each phase of the cell cycle during premeiosis and each stage of early meiosis. Flow cytometry has been revealed as a suitable and user-friendly tool to detect and quantify DNA replication during early meiosis in wheat. Chromosome replication was detected in wheat during premeiosis and early meiosis until the stage of pachytene, when chromosomes are associated in pairs to further recombine and correctly segregate in the gametes. In addition, the effect of the Ph1 locus, which controls chromosome pairing and affects replication in wheat, was also studied by flow cytometry. Here we showed that the Ph1 locus plays an important role on the length of meiotic DNA replication in wheat, particularly affecting the rate of replication during early meiosis in wheat. PMID:25275307

Fracture zones in the Mid Atlantic Ridge lead to alterations in prokaryotic and viral parameters in deep-water masses

PubMed Central

Muck, Simone; Griessler, Thomas; Köstner, Nicole; Klimiuk, Adam; Winter, Christian; Herndl, Gerhard J.

2014-01-01

We hypothesized that mixing zones of deep-water masses act as ecotones leading to alterations in microbial diversity and activity due to changes in the biogeochemical characteristics of these boundary systems. We determined the changes in prokaryotic and viral abundance and production in the Vema Fracture Zone (VFZ) of the subtropical North Atlantic Ocean, where North Atlantic Deep Water (NADW) and Antarctic Bottom Water (AABW) are funneled through this narrow canyon and therefore, are subjected to intense vertical mixing. Consequently, salinity, potential temperature, oxygen, PO4, SiO4, NO3 were altered in the NADW inside the VFZ as compared to the NADW outside of the VFZ. Also, viral abundance, lytic viral production (VP) and the virus-to-prokaryote ratio (VPR) were elevated in the NADW in the VFZ as compared to the NADW outside the VFZ. In contrast to lytic VP, lysogenic VP and both the frequency of lytically (FIC) and lysogenically infected cells (FLC) did not significantly differ between in- and outside the VFZ. Generally, FIC was higher than FLC throughout the water column. Prokaryotic (determined by T-RFLP) and viral (determined by RAPD-PCR) community composition was depth-stratified inside and outside the VFZ. The viral community was more modified both with depth and over distance inside the VFZ as compared to the northern section and to the prokaryotic communities. However, no clusters of prokaryotic and viral communities characteristic for the VFZ were identified. Based on our observations, we conclude that turbulent mixing of the deep water masses impacts not only the physico-chemical parameters of the mixing zone but also the interaction between viruses and prokaryotes due to a stimulation of the overall activity. However, only minor effects of deep water mixing were observed on the community composition of the dominant prokaryotes and viruses. PMID:24917857

Structure and interaction with phospholipids of a prokaryotic lipoxygenase from Pseudomonas aeruginosa

PubMed Central

Garreta, Albert; Val-Moraes, Silvana P.; García-Fernández, Queralt; Busquets, Montserrat; Juan, Carlos; Oliver, Antonio; Ortiz, Antonio; Gaffney, Betty J.; Fita, Ignacio; Manresa, Àngels; Carpena, Xavi

2013-01-01

Lipoxygenases (LOXs), which are essential in eukaryotes, have no confirmed function in prokaryotes that are devoid of polyunsaturated fatty acids. The structure of a secretable LOX from Pseudomonas aeruginosa (Pa_LOX), the first available from a prokaryote, presents significant differences with respect to eukaryotic LOXs, including a cluster of helices acting as a lid to the active center. The mobility of the lid and the structural variability of the N-terminal region of Pa_LOX was confirmed by comparing 2 crystal forms. The binding pocket contains a phosphatidylethanolamine phospholipid with branches of 18 (sn-1) and 14/16 (sn-2) carbon atoms in length. Carbon atoms from the sn-1 chain approach the catalytic iron in a manner that sheds light on how the enzymatic reaction might proceed. The findings in these studies suggest that Pa_LOX has the capacity to extract and modify unsaturated phospholipids from eukaryotic membranes, allowing this LOX to play a role in the interaction of P. aeruginosa with host cells.—Garreta, A., Val-Moraes, S. P., García-Fernández, Q., Montserrat Busquets, C. J., Oliver, A., Ortiz, A., Gaffney, B. J., Fita, I., Manresa, A., Carpena, X. Structure and interaction with phospholipids of a prokaryotic lipoxygenase from Pseudomonas aeruginosa. PMID:23985801

Cellular Viscosity in Prokaryotes and Thermal Stability of Low Molecular Weight Biomolecules.

PubMed

Cuecas, Alba; Cruces, Jorge; Galisteo-López, Juan F; Peng, Xiaojun; Gonzalez, Juan M

2016-08-23

Some low molecular weight biomolecules, i.e., NAD(P)H, are unstable at high temperatures. The use of these biomolecules by thermophilic microorganisms has been scarcely analyzed. Herein, NADH stability has been studied at different temperatures and viscosities. NADH decay increased at increasing temperatures. At increasing viscosities, NADH decay rates decreased. Thus, maintaining relatively high cellular viscosity in cells could result in increased stability of low molecular weight biomolecules (i.e., NADH) at high temperatures, unlike what was previously deduced from studies in diluted water solutions. Cellular viscosity was determined using a fluorescent molecular rotor in various prokaryotes covering the range from 10 to 100°C. Some mesophiles showed the capability of changing cellular viscosity depending on growth temperature. Thermophiles and extreme thermophiles presented a relatively high cellular viscosity, suggesting this strategy as a reasonable mechanism to thrive under these high temperatures. Results substantiate the capability of thermophiles and extreme thermophiles (growth range 50-80°C) to stabilize and use generally considered unstable, universal low molecular weight biomolecules. In addition, this study represents a first report, to our knowledge, on cellular viscosity measurements in prokaryotes and it shows the dependency of prokaryotic cellular viscosity on species and growth temperature. Copyright © 2016 Biophysical Society. Published by Elsevier Inc. All rights reserved.

A SHORT SEQUENCE IMMEDIATELY UPSTREAM OF THE INTERNAL REPEAT ELEMENTS IS CRITICAL FOR KSHV LANA MEDIATED DNA REPLICATION AND IMPACTS EPISOME PERSISTENCE

PubMed Central

León Vázquez, Erika De; Juillard, Franceline; Rosner, Bernard; Kaye, Kenneth M.

2013-01-01

Kaposi’s sarcoma-associated herpesvirus LANA (1162 residues) mediates episomal persistence of viral genomes during latency. LANA mediates viral DNA replication and segregates episomes to daughter nuclei. A 59 residue deletion immediately upstream of the internal repeat elements rendered LANA highly deficient for DNA replication and modestly deficient for the ability to segregate episomes, while smaller deletions did not. The 59 amino acid deletion reduced LANA episome persistence by ~14-fold, while sequentially smaller deletions resulted in ~3-fold, or no deficiency. Three distinct LANA regions reorganized heterochromatin, one of which contains the deleted sequence, but the deletion did not abolish LANA’s ability to alter chromatin. Therefore, this work identifies a short internal LANA sequence that is critical for DNA replication, has modest effects on episome segregation, and substantially impacts episome persistence; this region may exert its effects through an interacting host cell protein(s). PMID:24314665

Ethnic Segregation in Arizona Charter Schools.

ERIC Educational Resources Information Center

Cobb, Casey D.; Glass, Gene V.

1999-01-01

Addressed whether Arizona charter schools were more ethnically segregated than traditional public schools by studying 55 urban and 57 rural charter schools. Nearly half showed evidence of substantial ethnic segregation, and charter schools were higher in white enrollment than other public schools. (SLD)

An integrative and applicable phylogenetic footprinting framework for cis-regulatory motifs identification in prokaryotic genomes.

PubMed

Liu, Bingqiang; Zhang, Hanyuan; Zhou, Chuan; Li, Guojun; Fennell, Anne; Wang, Guanghui; Kang, Yu; Liu, Qi; Ma, Qin

2016-08-09

Phylogenetic footprinting is an important computational technique for identifying cis-regulatory motifs in orthologous regulatory regions from multiple genomes, as motifs tend to evolve slower than their surrounding non-functional sequences. Its application, however, has several difficulties for optimizing the selection of orthologous data and reducing the false positives in motif prediction. Here we present an integrative phylogenetic footprinting framework for accurate motif predictions in prokaryotic genomes (MP(3)). The framework includes a new orthologous data preparation procedure, an additional promoter scoring and pruning method and an integration of six existing motif finding algorithms as basic motif search engines. Specifically, we collected orthologous genes from available prokaryotic genomes and built the orthologous regulatory regions based on sequence similarity of promoter regions. This procedure made full use of the large-scale genomic data and taxonomy information and filtered out the promoters with limited contribution to produce a high quality orthologous promoter set. The promoter scoring and pruning is implemented through motif voting by a set of complementary predicting tools that mine as many motif candidates as possible and simultaneously eliminate the effect of random noise. We have applied the framework to Escherichia coli k12 genome and evaluated the prediction performance through comparison with seven existing programs. This evaluation was systematically carried out at the nucleotide and binding site level, and the results showed that MP(3) consistently outperformed other popular motif finding tools. We have integrated MP(3) into our motif identification and analysis server DMINDA, allowing users to efficiently identify and analyze motifs in 2,072 completely sequenced prokaryotic genomes. The performance evaluation indicated that MP(3) is effective for predicting regulatory motifs in prokaryotic genomes. Its application may enhance

Non-random Mis-segregation of Human Chromosomes.

PubMed

Worrall, Joseph Thomas; Tamura, Naoka; Mazzagatti, Alice; Shaikh, Nadeem; van Lingen, Tineke; Bakker, Bjorn; Spierings, Diana Carolina Johanna; Vladimirou, Elina; Foijer, Floris; McClelland, Sarah Elizabeth

2018-06-12

A common assumption is that human chromosomes carry equal chances of mis-segregation during compromised cell division. Human chromosomes vary in multiple parameters that might generate bias, but technological limitations have precluded a comprehensive analysis of chromosome-specific aneuploidy. Here, by imaging specific centromeres coupled with high-throughput single-cell analysis as well as single-cell sequencing, we show that aneuploidy occurs non-randomly following common treatments to elevate chromosome mis-segregation. Temporary spindle disruption leads to elevated mis-segregation and aneuploidy of a subset of chromosomes, particularly affecting chromosomes 1 and 2. Unexpectedly, we find that a period of mitotic delay weakens centromeric cohesion and promotes chromosome mis-segregation and that chromosomes 1 and 2 are particularly prone to suffer cohesion fatigue. Our findings demonstrate that inherent properties of individual chromosomes can bias chromosome mis-segregation and aneuploidy rates, with implications for studies on aneuploidy in human disease. Copyright © 2018 The Author(s). Published by Elsevier Inc. All rights reserved.

Large-Scale Distribution and Activity of Prokaryotes in Deep-Sea Surface Sediments of the Mediterranean Sea and the Adjacent Atlantic Ocean

PubMed Central

Giovannelli, Donato; Molari, Massimiliano; d’Errico, Giuseppe; Baldrighi, Elisa; Pala, Claudia; Manini, Elena

2013-01-01

The deep-sea represents a substantial portion of the biosphere and has a major influence on carbon cycling and global biogeochemistry. Benthic deep-sea prokaryotes have crucial roles in this ecosystem, with their recycling of organic matter from the photic zone. Despite this, little is known about the large-scale distribution of prokaryotes in the surface deep-sea sediments. To assess the influence of environmental and trophic variables on the large-scale distribution of prokaryotes, we investigated the prokaryotic assemblage composition (Bacteria to Archaea and Euryarchaeota to Crenarchaeota ratio) and activity in the surface deep-sea sediments of the Mediterranean Sea and the adjacent North Atlantic Ocean. Prokaryotic abundance and biomass did not vary significantly across the Mediterranean Sea; however, there were depth-related trends in all areas. The abundance of prokaryotes was positively correlated with the sedimentary concentration of protein, an indicator of the quality and bioavailability of organic matter. Moving eastwards, the Bacteria contribution to the total prokaryotes decreased, which appears to be linked to the more oligotrophic conditions of the Eastern Mediterranean basins. Despite the increased importance of Archaea, the contributions of Crenarchaeota Marine Group I to the total pool was relatively constant across the investigated stations, with the exception of Matapan-Vavilov Deep, in which Euryarchaeota Marine Group II dominated. Overall, our data suggest that deeper areas of the Mediterranean Sea share more similar communities with each other than with shallower sites. Freshness and quality of sedimentary organic matter were identified through Generalized Additive Model analysis as the major factors for describing the variation in the prokaryotic community structure and activity in the surface deep-sea sediments. Longitude was also important in explaining the observed variability, which suggests that the overlying water masses might have a

Bioinformatics analysis of disordered proteins in prokaryotes

PubMed Central

2011-01-01

Background A significant number of proteins have been shown to be intrinsically disordered, meaning that they lack a fixed 3 D structure or contain regions that do not posses a well defined 3 D structure. It has also been proven that a protein's disorder content is related to its function. We have performed an exhaustive analysis and comparison of the disorder content of proteins from prokaryotic organisms (i.e., superkingdoms Archaea and Bacteria) with respect to functional categories they belong to, i.e., Clusters of Orthologous Groups of proteins (COGs) and groups of COGs-Cellular processes (Cp), Information storage and processing (Isp), Metabolism (Me) and Poorly characterized (Pc). We also analyzed the disorder content of proteins with respect to various genomic, metabolic and ecological characteristics of the organism they belong to. We used correlations and association rule mining in order to identify the most confident associations between specific modalities of the characteristics considered and disorder content. Results Bacteria are shown to have a somewhat higher level of protein disorder than archaea, except for proteins in the Me functional group. It is demonstrated that the Isp and Cp functional groups in particular (L-repair function and N-cell motility and secretion COGs of proteins in specific) possess the highest disorder content, while Me proteins, in general, posses the lowest. Disorder fractions have been confirmed to have the lowest level for the so-called order-promoting amino acids and the highest level for the so-called disorder promoters. For each pair of organism characteristics, specific modalities are identified with the maximum disorder proteins in the corresponding organisms, e.g., high genome size-high GC content organisms, facultative anaerobic-low GC content organisms, aerobic-high genome size organisms, etc. Maximum disorder in archaea is observed for high GC content-low genome size organisms, high GC content-facultative anaerobic

Bioinformatics analysis of disordered proteins in prokaryotes.

PubMed

Pavlović-Lažetić, Gordana M; Mitić, Nenad S; Kovačević, Jovana J; Obradović, Zoran; Malkov, Saša N; Beljanski, Miloš V

2011-03-02

A significant number of proteins have been shown to be intrinsically disordered, meaning that they lack a fixed 3 D structure or contain regions that do not posses a well defined 3 D structure. It has also been proven that a protein's disorder content is related to its function. We have performed an exhaustive analysis and comparison of the disorder content of proteins from prokaryotic organisms (i.e., superkingdoms Archaea and Bacteria) with respect to functional categories they belong to, i.e., Clusters of Orthologous Groups of proteins (COGs) and groups of COGs-Cellular processes (Cp), Information storage and processing (Isp), Metabolism (Me) and Poorly characterized (Pc). We also analyzed the disorder content of proteins with respect to various genomic, metabolic and ecological characteristics of the organism they belong to. We used correlations and association rule mining in order to identify the most confident associations between specific modalities of the characteristics considered and disorder content. Bacteria are shown to have a somewhat higher level of protein disorder than archaea, except for proteins in the Me functional group. It is demonstrated that the Isp and Cp functional groups in particular (L-repair function and N-cell motility and secretion COGs of proteins in specific) possess the highest disorder content, while Me proteins, in general, posses the lowest. Disorder fractions have been confirmed to have the lowest level for the so-called order-promoting amino acids and the highest level for the so-called disorder promoters. For each pair of organism characteristics, specific modalities are identified with the maximum disorder proteins in the corresponding organisms, e.g., high genome size-high GC content organisms, facultative anaerobic-low GC content organisms, aerobic-high genome size organisms, etc. Maximum disorder in archaea is observed for high GC content-low genome size organisms, high GC content-facultative anaerobic or aquatic or

Fate of DNA encoding hygromycin resistance after meiosis in transformed strains of Gibberella fujikuroi (Fusarium moniliforme).

PubMed Central

Leslie, J F; Dickman, M B

1991-01-01

Stability of foreign DNA transformed into a novel host is an important parameter in decisions to permit the release of genetically engineered microorganisms into the environment. Meiotic instability of transformed DNA has been reported in fungi such as Ascobolus, Aspergillus, and Neurospora. We used strains of Gibberella fujikuroi (Fusarium moniliforme) transformed with the hygr gene from Escherichia coli to study meiotic stability of foreign DNA in this plant pathogenic fungus. Crosses with single-copy transformants segregated hygr:hygs in a 1:1 manner consistent with that expected for a Mendelian locus in a haploid organism. Multicopy transformants, however, segregated hygr:hygs in a 1:2 manner that was not consistent with Mendelian expectations for a chromosomal marker, even though two unrelated auxotrophic nuclear genes were segregating normally. Segregation ratios in crosses in which hygr was introduced via the male parent did not differ significantly from crosses in which the transformed strain served as the female parent. Some of the sensitive progeny from the crosses with the multicopy transformants carried hygr sequences. When these phenotypically sensitive progeny were crossed with a wild-type strain that carried no hygr sequences, some of the progeny were phenotypically hygr. Some progeny from some crosses were more resistant to hygromycin than were their sibs or the transformant strains that served as their parents. Transformants passaged through a maize plant only rarely segregated progeny with the high levels of resistance. The mechanism underlying these genetic instabilities is not clear but may involve unequal crossing over or methylation or both. Further work with cloned genes with homology to sequences already present in the Fusarium genome is warranted. Images PMID:1854200

Nonmagnetotactic Multicellular Prokaryotes from Low-Saline, Nonmarine Aquatic Environments and Their Unusual Negative Phototactic Behavior▿ †

PubMed Central

Lefèvre, Christopher T.; Abreu, Fernanda; Lins, Ulysses; Bazylinski, Dennis A.

2010-01-01

Magnetotactic multicellular prokaryotes (MMPs) are unique magnetotactic bacteria of the Deltaproteobacteria class and the first found to biomineralize the magnetic mineral greigite (Fe3S4). Thus far they have been reported only from marine habitats. We questioned whether MMPs exist in low-saline, nonmarine environments. MMPs were observed in samples from shallow springs in the Great Boiling Springs geothermal field and Pyramid Lake, both located in northwestern Nevada. The temperature at all sites was ambient, and salinities ranged from 5 to 11 ppt. These MMPs were not magnetotactic and did not contain magnetosomes (called nMMPs here). nMMPs ranged from 7 to 11 μm in diameter, were composed of about 40 to 60 Gram-negative cells, and were motile by numerous flagella that covered each cell on one side, characteristics similar to those of MMPs. 16S rRNA gene sequences of nMMPs show that they form a separate phylogenetic branch within the MMP group in the Deltaproteobacteria class, probably representing a single species. nMMPs exhibited a negative phototactic behavior to white light and to wavelengths of ≤480 nm (blue). We devised a “light racetrack” to exploit this behavior, which was used to photoconcentrate nMMPs for specific purposes (e.g., DNA extraction) even though their numbers were low in the sample. Our results show that the unique morphology of the MMP is not restricted to marine and magnetotactic prokaryotes. Discovery of nonmagnetotactic forms of the MMP might support the hypothesis that acquisition of the magnetosome genes involves horizontal gene transfer. To our knowledge, this is the first report of phototaxis in bacteria of the Deltaproteobacteria class. PMID:20363801

Skewed segregation of the mtDNA nt 8993 (T-->G) mutation in human oocytes.

PubMed Central

Blok, R B; Gook, D A; Thorburn, D R; Dahl, H H

1997-01-01

Rapid changes in mtDNA variants between generations have led to the bottleneck theory, which proposes a dramatic reduction in mtDNA numbers during early oogenesis. We studied oocytes from a woman with heteroplasmic expression of the mtDNA nt 8993 (T-->G) mutation. Of seven oocytes analyzed, one showed no evidence of the mutation, and the remaining six had a mutant load > 95%. This skewed expression of the mutation in oocytes is not compatible with the conventional bottleneck theory. A possible explanation is that, during amplification of mtDNA in the developing oocyte, mtDNA from one mitochondrion is preferentially amplified. Thus, subsequent mature oocytes may contain predominantly wild-type or mutant mitochondrial genomes. Images Figure 2 Figure 3 PMID:9199572

Thermodynamic impact of abasic sites on simulated translesion DNA synthesis.

PubMed

Malina, Jaroslav; Brabec, Viktor

2014-06-16

Loss of a base in DNA and the creation of an abasic (apurinic/apyrimidinic, AP) site is a frequent lesion that may occur spontaneously, or as a consequence of the action of DNA-damaging agents. The AP lesion is mutagenic or lethal if not repaired. We report a systematic thermodynamic investigation by differential scanning calorimetry on the evolution, during primer extension, of a model AP site in chemically simulated DNA translesion synthesis. Incorporation of dAMP (deoxyadenosine monophosphate), as well as dTMP (deoxythymidine monophosphate), opposite an AP site is enthalpically unfavorable, although incorporation of dTMP is more enthalpically unfavorable than that of dAMP. This finding is in a good agreement with experimental data showing that AP sites block various DNA polymerases of eukaryotic and prokaryotic origin and that, if bypassed, dAMP is preferentially inserted, whereas insertion of dTMP is less likely. The results emphasize the importance of thermodynamic contributions to the insertion of nucleotides opposite an AP site by DNA polymerases. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

PICH and BLM limit histone association with anaphase centromeric DNA threads and promote their resolution

PubMed Central

Ke, Yuwen; Huh, Jae-Wan; Warrington, Ross; Li, Bing; Wu, Nan; Leng, Mei; Zhang, Junmei; Ball, Haydn L; Li, Bing; Yu, Hongtao

2011-01-01

Centromeres nucleate the formation of kinetochores and are vital for chromosome segregation during mitosis. The SNF2 family helicase PICH (Plk1-interacting checkpoint helicase) and the BLM (the Bloom's syndrome protein) helicase decorate ultrafine histone-negative DNA threads that link the segregating sister centromeres during anaphase. The functions of PICH and BLM at these threads are not understood, however. Here, we show that PICH binds to BLM and enables BLM localization to anaphase centromeric threads. PICH- or BLM-RNAi cells fail to resolve these threads in anaphase. The fragmented threads form centromeric-chromatin-containing micronuclei in daughter cells. Anaphase threads in PICH- and BLM-RNAi cells contain histones and centromere markers. Recombinant purified PICH has nucleosome remodelling activities in vitro. We propose that PICH and BLM unravel centromeric chromatin and keep anaphase DNA threads mostly free of nucleosomes, thus allowing these threads to span long distances between rapidly segregating centromeres without breakage and providing a spatiotemporal window for their resolution. PMID:21743438

Metabolic rescue in pluripotent cells from patients with mtDNA disease.

PubMed

Ma, Hong; Folmes, Clifford D L; Wu, Jun; Morey, Robert; Mora-Castilla, Sergio; Ocampo, Alejandro; Ma, Li; Poulton, Joanna; Wang, Xinjian; Ahmed, Riffat; Kang, Eunju; Lee, Yeonmi; Hayama, Tomonari; Li, Ying; Van Dyken, Crystal; Gutierrez, Nuria Marti; Tippner-Hedges, Rebecca; Koski, Amy; Mitalipov, Nargiz; Amato, Paula; Wolf, Don P; Huang, Taosheng; Terzic, Andre; Laurent, Louise C; Izpisua Belmonte, Juan Carlos; Mitalipov, Shoukhrat

2015-08-13

Mitochondria have a major role in energy production via oxidative phosphorylation, which is dependent on the expression of critical genes encoded by mitochondrial (mt)DNA. Mutations in mtDNA can cause fatal or severely debilitating disorders with limited treatment options. Clinical manifestations vary based on mutation type and heteroplasmy (that is, the relative levels of mutant and wild-type mtDNA within each cell). Here we generated genetically corrected pluripotent stem cells (PSCs) from patients with mtDNA disease. Multiple induced pluripotent stem (iPS) cell lines were derived from patients with common heteroplasmic mutations including 3243A>G, causing mitochondrial encephalomyopathy and stroke-like episodes (MELAS), and 8993T>G and 13513G>A, implicated in Leigh syndrome. Isogenic MELAS and Leigh syndrome iPS cell lines were generated containing exclusively wild-type or mutant mtDNA through spontaneous segregation of heteroplasmic mtDNA in proliferating fibroblasts. Furthermore, somatic cell nuclear transfer (SCNT) enabled replacement of mutant mtDNA from homoplasmic 8993T>G fibroblasts to generate corrected Leigh-NT1 PSCs. Although Leigh-NT1 PSCs contained donor oocyte wild-type mtDNA (human haplotype D4a) that differed from Leigh syndrome patient haplotype (F1a) at a total of 47 nucleotide sites, Leigh-NT1 cells displayed transcriptomic profiles similar to those in embryo-derived PSCs carrying wild-type mtDNA, indicative of normal nuclear-to-mitochondrial interactions. Moreover, genetically rescued patient PSCs displayed normal metabolic function compared to impaired oxygen consumption and ATP production observed in mutant cells. We conclude that both reprogramming approaches offer complementary strategies for derivation of PSCs containing exclusively wild-type mtDNA, through spontaneous segregation of heteroplasmic mtDNA in individual iPS cell lines or mitochondrial replacement by SCNT in homoplasmic mtDNA-based disease.

Prokaryotic dynamics and heterotrophic metabolism in a deep convection site of Eastern Mediterranean Sea (the Southern Adriatic Pit)

NASA Astrophysics Data System (ADS)

Azzaro, M.; La Ferla, R.; Maimone, G.; Monticelli, L. S.; Zaccone, R.; Civitarese, G.

2012-08-01

We report on investigations of prokaryotic abundance, biomass, extracellular enzymatic activity, prokaryotic heterotrophic production and respiration in the full water column (˜1200 m) of a deep convection site (the Southern Adriatic Pit), carried out on six cruises in 2006-2008. Prokaryotic abundance (PA) varied vertically and temporally and ranged from 1.2 to 20.4×105 cell ml-1. Cell volumes, generally increased with depth; the lowest mean cell volume was observed in a period with no active convective process (Feb-07) and the highest in a period of stratification (Jun-08) following the convection process occurred in Feb-08. Prokaryotic biomass decreased with the depth and was related with both seasonal cycles of organic matter and hydrological processes. The picophytoplankton ranged in the upper layer (UL) from 0.089 to 10.71×104 cell ml-1. Cells were also recorded till 500 m depth in Feb-08 and this finding could be linked to water convection occurred in the Southern Adriatic Pit in that month. In UL the variations of enzymatic activities as well as leucine-aminopeptidase/ß-glucosidase ratio showed a seasonal trend probably linked to the productive processes of the photic layer. An inverse relation between alkaline phosphatase activity (APA) and phosphate concentrations was found (APA=0.0003PO4-1.7714, R2=0.333, P<0.05). Generally cell-specific enzymatic activities increased with depth as did cell-specific carbon dioxide production rates, while cell-specific prokaryotic heterotrophic production had an opposite trend. High values of prokaryotic growth efficiency registered in the deep layers in Nov-06 reflected a supply of preformed C transported within the deep water masses. Overall, in 2007 when no convective phenomenon was observed, the variability of prokaryotic metabolism was governed by the seasonal cycle of the organic matter, while in Nov-06 and Jun-08 the dynamics of deep water ventilation influenced the trend along the water column of many microbial

Iridescence of a Marine Bacterium and Classification of Prokaryotic Structural Colors

PubMed Central

Vukusic, Peter; Luke, Stephen

2012-01-01

Iridescence is a property of structural color that is occasionally encountered in higher eukaryotes but that has been poorly documented in the prokaryotic kingdom. In the present work, we describe a marine bacterium, identified as Cellulophaga lytica, isolated from the surface of an anemone, that exhibits bright green iridescent colonies under direct epi-illumination. This phenomenon has not previously been investigated in detail. In this study, color changes of C. lytica colonies were observed at various angles of direct illumination or observation. Its iridescent green appearance was dominant on various growth media. Red and violet colors were also discerned on colony edges. Remarkable C. lytica bacterial iridescence was revealed and characterized using high-resolution optical spectrometry. In addition to this, by culturing other bacterial strains to which various forms of faintly iridescent traits have previously been attributed, we identify four principal appearance characteristics of structural color in prokaryotes. A new general classification of bacterial iridescence is therefore proposed in this study. Furthermore, a specific separate class is described for iridescent C. lytica strains because they exhibit what is so far a unique intense glitter-like iridescence in reflection. C. lytica is the first prokaryote discovered to produce the same sort of intense iridescence under direct illumination as that associated with higher eukaryotes, like some insects and birds. Due to the nature of bacterial biology, cultivation, and ubiquity, this discovery may be of significant interest for both ecological and nanoscience endeavors. PMID:22267664

Chromosomal rearrangement segregating with adrenoleukodystrophy: associated changes in color vision.

PubMed Central

Alpern, M; Sack, G H; Krantz, D H; Jenness, J; Zhang, H; Moser, H W

1993-01-01

A patient from a large kindred with adrenoleukodystrophy showed profound disturbance of color ordering, color matching, increment thresholds, and luminosity. Except for color matching, his performance was similar to blue-cone "monochromacy," an X chromosome-linked recessive retinal dystrophy in which color vision is dichromatic, mediated by the visual pigments of rods and short-wave-sensitive cones. Color matching, however, indicated that an abnormal rudimentary visual pigment was also present. This may reflect the presence of a recombinant visual pigment protein or altered regulation of residual pigment genes, due to DNA changes--deletion of the long-wave pigment gene and reorganized sequences 5' to the pigment gene cluster--that segregate with the metabolic defect in this kindred. PMID:8415729

Subpopulation-proteomics in prokaryotic populations.

PubMed

Jahn, Michael; Seifert, Jana; von Bergen, Martin; Schmid, Andreas; Bühler, Bruno; Müller, Susann

2013-02-01

Clonal microbial cells do not behave in an identical manner and form subpopulations during cultivation. Besides varying micro-environmental conditions, cell inherent features like cell cycle dependent localization and concentration of regulatory proteins as well as epigenetic properties are well accepted mechanisms creating cell heterogeneity. Another suspected reason is molecular noise on the transcriptional and translational level. A promising tool to unravel reasons for cell heterogeneity is the combination of cell sorting and subpopulation proteomics. This review summarizes recent developments in prokaryotic single-cell analytics and provides a workflow for selection of single cells, low cell number mass spectrometry, and proteomics evaluation. This approach is useful for understanding the dependency of individual cell decisions on inherent protein profiles. Copyright © 2012 Elsevier Ltd. All rights reserved.

Segregation-mobility feedback for bidisperse shallow granular flows: Towards understanding segregation in geophysical flows

NASA Astrophysics Data System (ADS)

Thornton, A.; Denissen, I.; Weinhart, T.; Van der Vaart, K.

2017-12-01

The flow behaviour of shallow granular chute flows for uniform particles is well-described by the hstop-rheology [1]. Geophysical flows, however, are often composed of highly non-uniform particles that differ in particle (size, shape, composition) or contact (friction, dissipation, cohesion) properties. The flow behaviour of such mixtures can be strongly influenced by particle segregation effects. Here, we study the influence of particle size-segregation on the flow behaviour of bidisperse flows using experiments and the discrete particle method. We use periodic DPM to derive hstop-rheology for the bi-dispersed granular shallow layer equations, and study their dependence on the segregation profile. In the periodic box simulations, size-segregation results in an upward coarsening of the size distribution with the largest grains collecting at the top of the flow. In geophysical flows, the fact the flow velocity is greatest at the top couples with the vertical segregation to preferentially transported large particles to the front. The large grains may be overrun, resegregated towards the surface and recirculated before being shouldered aside into lateral levees. Theoretically it has been suggested this process should lead to a breaking size-segregation (BSS) wave located between a large-particle-rich front and a small-particle-rich tail [2,3]. In the BSS wave large particles that have been overrun rise up again to the free-surface while small particles sink to the bed. We present evidence for the existences of the BSS wave. This is achieved through the study of three-dimensional bidisperse granular flows in a moving-bed channel. Our analysis demonstrates a relation between the concentration of small particles in the flow and the amount of basal slip, in which the structure of the BSS wave plays a key role. This leads to a feedback between the mean bulk flow velocity and the process of size-segregation. Ultimately, these findings shed new light on the recirculation of

Yeast aconitase binds and provides metabolically coupled protection to mitochondrial DNA.

PubMed

Chen, Xin Jie; Wang, Xiaowen; Butow, Ronald A

2007-08-21

Aconitase (Aco1p) is a multifunctional protein: It is an enzyme of the tricarboxylic acid cycle. In animal cells, Aco1p also is a cytosolic protein binding to mRNAs to regulate iron metabolism. In yeast, Aco1p was identified as a component of mtDNA nucleoids. Here we show that yeast Aco1p protects mtDNA from excessive accumulation of point mutations and ssDNA breaks and suppresses reductive recombination of mtDNA. Aconitase binds to both ds- and ssDNA, with a preference for GC-containing sequences. Therefore, mitochondria are opportunistic organelles that seize proteins, such as metabolic enzymes, for construction of the nucleoid, an mtDNA maintenance/segregation apparatus.

Structure and information in spatial segregation

PubMed Central

2017-01-01

Ethnoracial residential segregation is a complex, multiscalar phenomenon with immense moral and economic costs. Modeling the structure and dynamics of segregation is a pressing problem for sociology and urban planning, but existing methods have limitations. In this paper, we develop a suite of methods, grounded in information theory, for studying the spatial structure of segregation. We first advance existing profile and decomposition methods by posing two related regionalization methods, which allow for profile curves with nonconstant spatial scale and decomposition analysis with nonarbitrary areal units. We then formulate a measure of local spatial scale, which may be used for both detailed, within-city analysis and intercity comparisons. These methods highlight detailed insights in the structure and dynamics of urban segregation that would be otherwise easy to miss or difficult to quantify. They are computationally efficient, applicable to a broad range of study questions, and freely available in open source software. PMID:29078323

Structure and information in spatial segregation.

PubMed

Chodrow, Philip S

2017-10-31

Ethnoracial residential segregation is a complex, multiscalar phenomenon with immense moral and economic costs. Modeling the structure and dynamics of segregation is a pressing problem for sociology and urban planning, but existing methods have limitations. In this paper, we develop a suite of methods, grounded in information theory, for studying the spatial structure of segregation. We first advance existing profile and decomposition methods by posing two related regionalization methods, which allow for profile curves with nonconstant spatial scale and decomposition analysis with nonarbitrary areal units. We then formulate a measure of local spatial scale, which may be used for both detailed, within-city analysis and intercity comparisons. These methods highlight detailed insights in the structure and dynamics of urban segregation that would be otherwise easy to miss or difficult to quantify. They are computationally efficient, applicable to a broad range of study questions, and freely available in open source software. Published under the PNAS license.

HOW POPULATION STRUCTURE SHAPES NEIGHBORHOOD SEGREGATION*

PubMed Central

Bruch, Elizabeth E.

2014-01-01

This study investigates how choices about social affiliation based on one attribute can exacerbate or attenuate segregation on another correlated attribute. The specific application is the role of racial and economic factors in generating patterns of racial residential segregation. I identify three population parameters—between-group inequality, within-group inequality, and relative group size—that determine how income inequality between race groups affects racial segregation. I use data from the Panel Study of Income Dynamics to estimate models of individual-level residential mobility, and incorporate these estimates into agent-based models. I then simulate segregation dynamics under alternative assumptions about: (1) the relative size of minority groups; and (2) the degree of correlation between race and income among individuals. I find that income inequality can have offsetting effects at the high and low ends of the income distribution. I demonstrate the empirical relevance of the simulation results using fixed-effects, metro-level regressions applied to 1980-2000 U.S. Census data. PMID:25009360

Eukaryotic major facilitator superfamily transporter modeling based on the prokaryotic GlpT crystal structure.

PubMed

Lemieux, M Joanne

2007-01-01

The major facilitator superfamily (MFS) of transporters represents the largest family of secondary active transporters and has a diverse range of substrates. With structural information for four MFS transporters, we can see a strong structural commonality suggesting, as predicted, a common architecture for MFS transporters. The rate for crystal structure determination of MFS transporters is slow, making modeling of both prokaryotic and eukaryotic transporters more enticing. In this review, models of eukaryotic transporters Glut1, G6PT, OCT1, OCT2 and Pho84, based on the crystal structures of the prokaryotic GlpT, based on the crystal structure of LacY are discussed. The techniques used to generate the different models are compared. In addition, the validity of these models and the strategy of using prokaryotic crystal structures to model eukaryotic proteins are discussed. For comparison, E. coli GlpT was modeled based on the E. coli LacY structure and compared to the crystal structure of GlpT demonstrating that experimental evidence is essential for accurate modeling of membrane proteins.

Structures of benthic prokaryotic communities and their hydrolytic enzyme activities resuspended from samples of intertidal mudflats: An experimental approach

NASA Astrophysics Data System (ADS)

Mallet, Clarisse; Agogué, Hélène; Bonnemoy, Frédérique; Guizien, Katell; Orvain, Francis; Dupuy, Christine

2014-09-01

Resuspended sediment can increase plankton biomass and the growth of bacteria, thus influencing the coastal planktonic microbial food web. But little is known about resuspension itself: is it a single massive change or a whole series of events and how does it affect the quantity and quality of resuspended prokaryotic cells? We simulated the sequential erosion of mud cores to better understand the fate and role of benthic prokaryotes resuspended in the water column. We analyzed the total, attached and free-living prokaryotic cells resuspended, their structure and the activities of their hydrolytic enzymes in terms of the biotic and abiotic factors that affect the composition of microphytobenthic biofilm. Free living prokaryotes were resuspended during the fluff layer erosion phase (for shear velocities below 5 cm · s- 1) regardless of the bed sediment composition. At the higher shear velocities, resuspended prokaryotes were attached to particulate matter. Free and attached cells are thus unevenly distributed, scattered throughout the organic matter (OM) in the uppermost mm of the sediment. Only 10-27% of the total cells initially resuspended were living and most of the Bacteria were Cyanobacteria and Gamma-proteobacteria; their numbers increased to over 30% in parallel with the hydrolytic enzyme activity at highest shear velocity. These conditions released prokaryotic cells having different functions that lie deep in the sediment; the most important of them are Archaea. Finally, composition of resuspended bacterial populations varied with resuspension intensity, and intense resuspension events boosted the microbial dynamics and enzyme activities in the bottom layers of sea water.

Analysis of Minor Component Segregation in Ternary Powder Mixtures

NASA Astrophysics Data System (ADS)

Asachi, Maryam; Hassanpour, Ali; Ghadiri, Mojtaba; Bayly, Andrew

2017-06-01

In many powder handling operations, inhomogeneity in powder mixtures caused by segregation could have significant adverse impact on the quality as well as economics of the production. Segregation of a minor component of a highly active substance could have serious deleterious effects, an example is the segregation of enzyme granules in detergent powders. In this study, the effects of particle properties and bulk cohesion on the segregation tendency of minor component are analysed. The minor component is made sticky while not adversely affecting the flowability of samples. The segregation extent is evaluated using image processing of the photographic records taken from the front face of the heap after the pouring process. The optimum average sieve cut size of components for which segregation could be reduced is reported. It is also shown that the extent of segregation is significantly reduced by applying a thin layer of liquid to the surfaces of minor component, promoting an ordered mixture.

Self-organized Segregation on the Grid

NASA Astrophysics Data System (ADS)

Omidvar, Hamed; Franceschetti, Massimo

2018-02-01

We consider an agent-based model with exponentially distributed waiting times in which two types of agents interact locally over a graph, and based on this interaction and on the value of a common intolerance threshold τ , decide whether to change their types. This is equivalent to a zero-temperature ising model with Glauber dynamics, an asynchronous cellular automaton with extended Moore neighborhoods, or a Schelling model of self-organized segregation in an open system, and has applications in the analysis of social and biological networks, and spin glasses systems. Some rigorous results were recently obtained in the theoretical computer science literature, and this work provides several extensions. We enlarge the intolerance interval leading to the expected formation of large segregated regions of agents of a single type from the known size ɛ >0 to size ≈ 0.134. Namely, we show that for 0.433< τ < 1/2 (and by symmetry 1/2<τ <0.567), the expected size of the largest segregated region containing an arbitrary agent is exponential in the size of the neighborhood. We further extend the interval leading to expected large segregated regions to size ≈ 0.312 considering "almost segregated" regions, namely regions where the ratio of the number of agents of one type and the number of agents of the other type vanishes quickly as the size of the neighborhood grows. In this case, we show that for 0.344 < τ ≤ 0.433 (and by symmetry for 0.567 ≤ τ <0.656) the expected size of the largest almost segregated region containing an arbitrary agent is exponential in the size of the neighborhood. This behavior is reminiscent of supercritical percolation, where small clusters of empty sites can be observed within any sufficiently large region of the occupied percolation cluster. The exponential bounds that we provide also imply that complete segregation, where agents of a single type cover the whole grid, does not occur with high probability for p=1/2 and the range of

Positive regulation of meiotic DNA double-strand break formation by activation of the DNA damage checkpoint kinase Mec1(ATR).

PubMed

Gray, Stephen; Allison, Rachal M; Garcia, Valerie; Goldman, Alastair S H; Neale, Matthew J

2013-07-31

During meiosis, formation and repair of programmed DNA double-strand breaks (DSBs) create genetic exchange between homologous chromosomes-a process that is critical for reductional meiotic chromosome segregation and the production of genetically diverse sexually reproducing populations. Meiotic DSB formation is a complex process, requiring numerous proteins, of which Spo11 is the evolutionarily conserved catalytic subunit. Precisely how Spo11 and its accessory proteins function or are regulated is unclear. Here, we use Saccharomyces cerevisiae to reveal that meiotic DSB formation is modulated by the Mec1(ATR) branch of the DNA damage signalling cascade, promoting DSB formation when Spo11-mediated catalysis is compromised. Activation of the positive feedback pathway correlates with the formation of single-stranded DNA (ssDNA) recombination intermediates and activation of the downstream kinase, Mek1. We show that the requirement for checkpoint activation can be rescued by prolonging meiotic prophase by deleting the NDT80 transcription factor, and that even transient prophase arrest caused by Ndt80 depletion is sufficient to restore meiotic spore viability in checkpoint mutants. Our observations are unexpected given recent reports that the complementary kinase pathway Tel1(ATM) acts to inhibit DSB formation. We propose that such antagonistic regulation of DSB formation by Mec1 and Tel1 creates a regulatory mechanism, where the absolute frequency of DSBs is maintained at a level optimal for genetic exchange and efficient chromosome segregation.

Silver-doped calcium phosphate nanoparticles: synthesis, characterization, and toxic effects toward mammalian and prokaryotic cells.

PubMed

Peetsch, Alexander; Greulich, Christina; Braun, Dieter; Stroetges, Christian; Rehage, Heinz; Siebers, Bettina; Köller, Manfred; Epple, Matthias

2013-02-01

Spherical silver-doped calcium phosphate nanoparticles were synthesized in a co-precipitation route from calcium nitrate/silver nitrate and ammonium phosphate in a continuous process and colloidally stabilized by carboxymethyl cellulose. Nanoparticles with 0.39 wt% silver content and a diameter of about 50-60 nm were obtained. The toxic effects toward mammalian and prokaryotic cells were determined by viability tests and determination of the minimal inhibitory and minimal bactericidal concentrations (MIC and MBC). Three mammalian cells lines, i.e. human mesenchymal stem cells (hMSC) and blood peripheral mononuclear cells (PBMC, monocytes and T-lymphocytes), and two prokaryotic strains, i.e. Escherichia coli (E. coli) and Staphylococcus aureus (S. aureus) were used. Silver-doped calcium phosphate nanoparticles and silver acetate showed similar effect toward mammalian and prokaryotic cells with toxic silver concentrations in the range of 1-3 μg mL(-1). Copyright © 2012 Elsevier B.V. All rights reserved.

CRISPR: From Prokaryotic Immune Systems to Plant Genome Editing Tools.

PubMed

Bandyopadhyay, Anindya; Mazumdar, Shamik; Yin, Xiaojia; Quick, William Paul

2017-01-01

The clustered regularly interspaced short palindromic repeats (CRISPR) system is a prokaryotic adaptive immune system that has the ability to identify specific locations on the bacteriophage (phage) genome to create breaks in it, and internalize the phage genome fragments in its own genome as CRISPR arrays for memory-dependent resistance. Although CRISPR has been used in the dairy industry for a long time, it recently gained importance in the field of genome editing because of its ability to precisely target locations in a genome. This system has further been modified to locate and target any region of a genome of choice due to modifications in the components of the system. By changing the nucleotide sequence of the 20-nucleotide target sequence in the guide RNA, targeting any location is possible. It has found an application in the modification of plant genomes with its ability to generate mutations and insertions, thus helping to create new varieties of plants. With the ability to introduce specific sequences into the plant genome after cleavage by the CRISPR system and subsequent DNA repair through homology-directed repair (HDR), CRISPR ensures that genome editing can be successfully applied in plants, thus generating stronger and more improved traits. Also, the use of the CRISPR editing system can generate plants that are transgene-free and have mutations that are stably inherited, thus helping to circumvent current GMO regulations.

Stochastic correlative firing for figure-ground segregation.

PubMed

Chen, Zhe

2005-03-01

Segregation of sensory inputs into separate objects is a central aspect of perception and arises in all sensory modalities. The figure-ground segregation problem requires identifying an object of interest in a complex scene, in many cases given binaural auditory or binocular visual observations. The computations required for visual and auditory figure-ground segregation share many common features and can be cast within a unified framework. Sensory perception can be viewed as a problem of optimizing information transmission. Here we suggest a stochastic correlative firing mechanism and an associative learning rule for figure-ground segregation in several classic sensory perception tasks, including the cocktail party problem in binaural hearing, binocular fusion of stereo images, and Gestalt grouping in motion perception.

Sound segregation via embedded repetition is robust to inattention.

PubMed

Masutomi, Keiko; Barascud, Nicolas; Kashino, Makio; McDermott, Josh H; Chait, Maria

2016-03-01

The segregation of sound sources from the mixture of sounds that enters the ear is a core capacity of human hearing, but the extent to which this process is dependent on attention remains unclear. This study investigated the effect of attention on the ability to segregate sounds via repetition. We utilized a dual task design in which stimuli to be segregated were presented along with stimuli for a "decoy" task that required continuous monitoring. The task to assess segregation presented a target sound 10 times in a row, each time concurrent with a different distractor sound. McDermott, Wrobleski, and Oxenham (2011) demonstrated that repetition causes the target sound to be segregated from the distractors. Segregation was queried by asking listeners whether a subsequent probe sound was identical to the target. A control task presented similar stimuli but probed discrimination without engaging segregation processes. We present results from 3 different decoy tasks: a visual multiple object tracking task, a rapid serial visual presentation (RSVP) digit encoding task, and a demanding auditory monitoring task. Load was manipulated by using high- and low-demand versions of each decoy task. The data provide converging evidence of a small effect of attention that is nonspecific, in that it affected the segregation and control tasks to a similar extent. In all cases, segregation performance remained high despite the presence of a concurrent, objectively demanding decoy task. The results suggest that repetition-based segregation is robust to inattention. (c) 2016 APA, all rights reserved).

Losing Ground: School Segregation in Massachuestts

ERIC Educational Resources Information Center

Ayscue, Jennifer B.; Greenberg, Alyssa

2013-01-01

Though once a leader in school integration, Massachusetts has regressed over the last two decades as its students of color have experienced intensifying school segregation. This report investigates trends in school segregation in Massachusetts by examining concentration, exposure, and evenness measures by both race and class. First, the report…

Sex Segregation in Undergraduate Engineering Majors

ERIC Educational Resources Information Center

Litzler, Elizabeth

2010-01-01

Gender inequality in engineering persists in spite of women reaching parity in college enrollments and degrees granted. To date, no analyses of educational sex segregation have comprehensively examined segregation within one discipline. To move beyond traditional methods of studying the long-standing stratification by field of study in higher…

Residential Segregation in Texas in 1980.

ERIC Educational Resources Information Center

Hwang, Sean-Shong; Murdock, Steve H.

1982-01-01

Between 1970 and 1980 racial and ethnic segregation for major Texas cities declined for all groups, but declines were small between Anglo and Spanish groups. Segregation is unaffected by variation in size of city, percent of population that is Spanish or Black, or central city status. (Author/AM)