Genome-wide biases in the rate and molecular spectrum of spontaneous mutations in Vibrio cholerae and Vibrio fischeri

The vast diversity in nucleotide composition and architecture among bacterial genomes may be partly explained by inherent biases in the rates and spectra of spontaneous mutations. Bacterial genomes with multiple chromosomes are relatively unusual but some are relevant to human health, none more so than the causative agent of cholera, Vibrio cholerae. Here, we present the genome-wide mutation spectra in wild-type and mismatch repair (MMR) defective backgrounds of two Vibrio species, the low-GC% squid symbiont V. fischeri and the pathogen V. cholerae, collected under conditions that greatly minimize the efficiency of natural selection. In apparent contrast to their high diversity in nature, both wild-type V. fischeri and V. cholerae have among the lowest rates for base-substitution mutations (bpsms) and insertion-deletion mutations (indels) that have been measured, below 10-3/genome/generation. V. fischeri and V. cholerae have distinct mutation spectra, but both are AT-biased and produce a surprising number of multi-nucleotide indels. Furthermore, the loss of a functional MMR system caused the mutation spectra of these species to converge, implying that the MMR system itself contributes to species-specific mutation patterns. Bpsm and indel rates varied among genome regions, but do not explain the more rapid evolutionary rates of genes on chromosome 2, which likely result from weaker purifying selection. More generally, the very low mutation rates of Vibrio species correlate inversely with their immense population sizes and suggest that selection may not only have maximized replication fidelity but also optimized other polygenic traits relative to the constraints of genetic drift.

Download Full-text

Periodic variation of mutation rates in bacterial genomes associated with replication timing

10.1101/195818 ◽

2017 ◽

Author(s):

Marcus M. Dillon ◽

Way Sung ◽

Michael Lynch ◽

Vaughn S. Cooper

Keyword(s):

Vibrio Fischeri ◽

Bacterial Species ◽

Replication Timing ◽

Small Chromosome ◽

Burkholderia Cenocepacia ◽

Mutation Rates ◽

Base Substitution ◽

Large Chromosome ◽

Bacterial Genomes ◽

Substitution Mutation

ABSTRACTThe causes and consequences of spatiotemporal variation in mutation rates remains to be explored in nearly all organisms. Here we examine relationships between local mutation rates and replication timing in three bacterial species whose genomes have multiple chromosomes:Vibrio fischeri, Vibrio cholerae, andBurkholderia cenocepacia. Following five evolution experiments with these bacteria conducted in the near-absence of natural selection, the genomes of clones from each lineage were sequenced and analyzed to identify variation in mutation rates and spectra. In lineages lacking mismatch repair, base-substitution mutation rates vary in a mirrored wave-like pattern on opposing replichores of the large chromosome ofV. fischeriandV. cholerae, where concurrently replicated regions experience similar base-substitution mutation rates. The base-substitution mutation rates on the small chromosome are less variable in both species but occur at similar rates as the concurrently replicated regions of the large chromosome. Neither nucleotide composition nor frequency of nucleotide motifs differed among regions experiencing high and low base-substitution rates, which along with the inferred ~800 Kb wave period suggests that the source of the periodicity is not sequence-specific but rather a systematic process related to the cell cycle. These results support the notion that base-substitution mutation rates are likely to vary systematically across many bacterial genomes, which exposes certain genes to elevated deleterious mutational load.

Download Full-text

Mutational Landscape of Spontaneous Base Substitutions and Small Indels in Experimental Caenorhabditis elegans Populations of Differing Size

10.1101/529214 ◽

2019 ◽

Author(s):

Anke Konrad ◽

Meghan J. Brady ◽

Ulfar Bergthorsson ◽

Vaishali Katju

Keyword(s):

Caenorhabditis Elegans ◽

Evolutionary Process ◽

Purifying Selection ◽

Base Substitution ◽

Experimental Investigations ◽

Spontaneous Mutations ◽

Small Indels ◽

Mtdna Mutations ◽

Base Substitutions ◽

Population Sizes

ABSTRACTExperimental investigations into the rates and fitness effects of spontaneous mutations are fundamental for our understanding of the evolutionary process. To gain insights into the molecular and fitness consequences of spontaneous mutations, we conducted a mutation accumulation (MA) experiment at varying population sizes in the nematode Caenorhabditis elegans, evolving 35 lines in parallel for 409 generations at three population sizes (N = 1, 10, 100 individuals). Here, we focus on nuclear SNPs and small indels under minimal influence of selection, as well as their accrual rates in larger populations under greater selection efficacy. The spontaneous rates of base substitutions and small indels are 1.84 × 10−9 substitutions and 6.84 × 10−10 changes /site/generation, respectively. Small indels exhibit a deletion-bias with deletions exceeding insertions by three-fold. Notably, there was no correlation between the frequency of base substitutions, nonsynonymous substitutions or small indels with population size. These results contrast with our previous analysis of mtDNA mutations and nuclear copy-number changes in these MA lines, and suggest that nuclear base substitutions and small indels are under less stringent purifying selection compared to the former mutational classes. A transition bias was observed in exons as was a near universal base substitution bias towards A/T. Strongly context-dependent base substitutions, where 5’–T and 3’–As increase the frequency of A/T → T/A transversions, especially at the boundaries of A or T homopolymeric runs, manifest as higher mutation rates in (i) introns and intergenic regions relative to exons, (ii) chromosomal cores versus arms and tips, and (iii) germline-expressed genes.

Download Full-text

The rate and molecular spectrum of spontaneous mutations in the GC-rich multi-chromosome genome ofBurkholderia cenocepacia

10.1101/011841 ◽

2014 ◽

Author(s):

Marcus M Dillon ◽

Way Sung ◽

Michael Lynch ◽

Vaughn S Cooper

Keyword(s):

Prokaryotic Genome ◽

Gc Content ◽

Nucleotide Composition ◽

Burkholderia Cenocepacia ◽

Model Organisms ◽

Base Substitution ◽

Spontaneous Mutations ◽

Mutation Pressure ◽

Genome Wide ◽

A Genome

Spontaneous mutations are ultimately essential for evolutionary change and are also the root cause of many diseases. However, until recently, both biological and technical barriers have prevented detailed analyses of mutation profiles, constraining our understanding of the mutation process to a few model organisms and leaving major gaps in our understanding of the role of genome content and structure on mutation. Here, we present a genome-wide view of the molecular mutation spectrum in Burkholderia cenocepacia, a clinically relevant pathogen with high %GC-content and multiple chromosomes. We find that B. cenocepacia has low genome-wide mutation rates with insertion-deletion mutations biased towards deletions, consistent with the idea that deletion pressure reduces prokaryotic genome sizes. Unlike prior studies of other organisms, mutations in B. cenocepacia are not AT-biased, which suggests that at least some genomes with high %GC-content experience unusual base-substitution mutation pressure. Importantly, we also observe variation in both the rates and spectra of mutations among chromosomes and elevated G:C>T:A transversions in late-replicating regions. Thus, although some patterns of mutation appear to be highly conserved across cellular life, others vary between species and even between chromosomes of the same species, potentially influencing the evolution of nucleotide composition and genome architecture.

Download Full-text

Mutational Landscape of Spontaneous Base Substitutions and Small Indels in Experimental Caenorhabditis elegans Populations of Differing Size

Genetics ◽

10.1534/genetics.119.302054 ◽

2019 ◽

Vol 212 (3) ◽

pp. 837-854 ◽

Cited By ~ 5

Author(s):

Anke Konrad ◽

Meghan J. Brady ◽

Ulfar Bergthorsson ◽

Vaishali Katju

Keyword(s):

Caenorhabditis Elegans ◽

Evolutionary Process ◽

Purifying Selection ◽

Base Substitution ◽

Experimental Investigations ◽

Spontaneous Mutations ◽

Small Indels ◽

Base Substitutions ◽

Population Sizes ◽

Copy Number Changes

Experimental investigations into the rates and fitness effects of spontaneous mutations are fundamental to our understanding of the evolutionary process. To gain insights into the molecular and fitness consequences of spontaneous mutations, we conducted a mutation accumulation (MA) experiment at varying population sizes in the nematode Caenorhabditis elegans, evolving 35 lines in parallel for 409 generations at three population sizes (N = 1, 10, and 100 individuals). Here, we focus on nuclear SNPs and small insertion/deletions (indels) under minimal influence of selection, as well as their accrual rates in larger populations under greater selection efficacy. The spontaneous rates of base substitutions and small indels are 1.84 (95% C.I. ± 0.14) × 10−9 substitutions and 6.84 (95% C.I. ± 0.97) × 10−10 changes/site/generation, respectively. Small indels exhibit a deletion bias with deletions exceeding insertions by threefold. Notably, there was no correlation between the frequency of base substitutions, nonsynonymous substitutions, or small indels with population size. These results contrast with our previous analysis of mitochondrial DNA mutations and nuclear copy-number changes in these MA lines, and suggest that nuclear base substitutions and small indels are under less stringent purifying selection compared to the former mutational classes. A transition bias was observed in exons as was a near universal base substitution bias toward A/T. Strongly context-dependent base substitutions, where 5′−Ts and 3′−As increase the frequency of A/T → T/A transversions, especially at the boundaries of A or T homopolymeric runs, manifest as higher mutation rates in (i) introns and intergenic regions relative to exons, (ii) chromosomal cores vs. arms and tips, and (iii) germline-expressed genes.

Download Full-text

Estimation of the Genome-Wide Mutation Rate and Spectrum in the Archaeal Species Haloferax volcanii

Genetics ◽

10.1534/genetics.120.303299 ◽

2020 ◽

Vol 215 (4) ◽

pp. 1107-1116 ◽

Cited By ~ 1

Author(s):

Sibel Kucukyildirim ◽

Megan Behringer ◽

Emily M. Williams ◽

Thomas G. Doak ◽

Michael Lynch

Keyword(s):

Molecular Mechanisms ◽

Model Organism ◽

Optimal Growth ◽

Haloferax Volcanii ◽

Base Substitution ◽

Spontaneous Mutations ◽

Evolutionary Forces ◽

Genome Wide ◽

Archaeal Species ◽

A Genome

Organisms adapted to life in extreme habitats (extremophiles) can further our understanding of the mechanisms of genetic stability, particularly replication and repair. Despite the harsh environmental conditions they endure, these extremophiles represent a great deal of the Earth’s biodiversity. Here, for the first time in a member of the archaeal domain, we report a genome-wide assay of spontaneous mutations in the halophilic species Haloferax volcanii using a direct and unbiased method: mutation accumulation experiments combined with deep whole-genome sequencing. H. volcanii is a key model organism not only for the study of halophilicity, but also for archaeal biology in general. Our methods measure the genome-wide rate, spectrum, and spatial distribution of spontaneous mutations. The estimated base substitution rate of 3.15 × 10−10 per site per generation, or 0.0012 per genome per generation, is similar to the value found in mesophilic prokaryotes (optimal growth at ∼20–45°). This study contributes to a comprehensive phylogenetic view of how evolutionary forces and molecular mechanisms shape the rate and molecular spectrum of mutations across the tree of life.

Download Full-text

Developing a Genetic System in Deinococcus radiodurans for Analyzing Mutations

Genetics ◽

10.1093/genetics/166.2.661 ◽

2004 ◽

Vol 166 (2) ◽

pp. 661-668

Author(s):

Mandy Kim ◽

Erika Wolff ◽

Tiffany Huang ◽

Lilit Garibyan ◽

Ashlee M Earl ◽

...

Keyword(s):

Dna Sequences ◽

Deinococcus Radiodurans ◽

Genetic System ◽

Base Change ◽

Base Substitution ◽

Wild Type ◽

Base Pairs ◽

E Coli ◽

Radiation Induced ◽

Induced Mutagenesis

Abstract We have applied a genetic system for analyzing mutations in Escherichia coli to Deinococcus radiodurans, an extremeophile with an astonishingly high resistance to UV- and ionizing-radiation-induced mutagenesis. Taking advantage of the conservation of the β-subunit of RNA polymerase among most prokaryotes, we derived again in D. radiodurans the rpoB/Rif r system that we developed in E. coli to monitor base substitutions, defining 33 base change substitutions at 22 different base pairs. We sequenced >250 mutations leading to Rif r in D. radiodurans derived spontaneously in wild-type and uvrD (mismatch-repair-deficient) backgrounds and after treatment with N-methyl-N′-nitro-N-nitrosoguanidine (NTG) and 5-azacytidine (5AZ). The specificities of NTG and 5AZ in D. radiodurans are the same as those found for E. coli and other organisms. There are prominent base substitution hotspots in rpoB in both D. radiodurans and E. coli. In several cases these are at different points in each organism, even though the DNA sequences surrounding the hotspots and their corresponding sites are very similar in both D. radiodurans and E. coli. In one case the hotspots occur at the same site in both organisms.

Download Full-text

cot1: A Regulator of Arabidopsis Trichome Initiation

Genetics ◽

10.1093/genetics/149.2.565 ◽

1998 ◽

Vol 149 (2) ◽

pp. 565-577

Author(s):

Daniel B Szymanski ◽

Daniel A Klis ◽

John C Larkin ◽

M David Marks

Keyword(s):

Cell Fate ◽

Gene Dosage ◽

Signal Transduction Pathway ◽

Spatial Arrangement ◽

Complex Signal ◽

Chromosome 2 ◽

Wild Type ◽

Trichome Formation ◽

In The Wild ◽

Leaf Trichome

Abstract In Arabidopsis, the timing and spatial arrangement of trichome initiation is tightly regulated and requires the activity of the GLABROUS1 (GL1) gene. The COTYLEDON TRICHOME 1 (COT1) gene affects trichome initiation during late stages of leaf development and is described in this article. In the wild-type background, cot1 has no observable effect on trichome initiation. GL1 overexpression in wild-type plants leads to a modest number of ectopic trichomes and to a decrease in trichome number on the adaxial leaf surface. The cot1 mutation enhances GL1-overexpression-dependent ectopic trichome formation and also induces increased leaf trichome initiation. The expressivity of the cot1 phenotype is sensitive to cot1 and 35S::GL1 gene dosage, and the most severe phenotypes are observed when cot1 and 35S::GL1 are homozygous. The COT1 locus is located on chromosome 2 15.3 cM north of er. Analysis of the interaction between cot1, try, and 35S::GL1 suggests that COT1 is part of a complex signal transduction pathway that regulates GL1-dependent adoption of the trichome cell fate.

Download Full-text

Genome-Wide Identification of Soybean ABC Transporters Relate to Aluminum Toxicity

International Journal of Molecular Sciences ◽

10.3390/ijms22126556 ◽

2021 ◽

Vol 22 (12) ◽

pp. 6556

Author(s):

Junjun Huang ◽

Xiaoyu Li ◽

Xin Chen ◽

Yaru Guo ◽

Weihong Liang ◽

...

Keyword(s):

Gene Expression ◽

Abc Transporters ◽

Developmental Stages ◽

Aluminum Toxicity ◽

Expression Profiles ◽

Gene Expression Profiles ◽

Purifying Selection ◽

Biotic Stresses ◽

Genome Wide ◽

New Germplasm

ATP-binding cassette (ABC) transporter proteins are a gene super-family in plants and play vital roles in growth, development, and response to abiotic and biotic stresses. The ABC transporters have been identified in crop plants such as rice and buckwheat, but little is known about them in soybean. Soybean is an important oil crop and is one of the five major crops in the world. In this study, 255 ABC genes that putatively encode ABC transporters were identified from soybean through bioinformatics and then categorized into eight subfamilies, including 7 ABCAs, 52 ABCBs, 48 ABCCs, 5 ABCDs, 1 ABCEs, 10 ABCFs, 111 ABCGs, and 21 ABCIs. Their phylogenetic relationships, gene structure, and gene expression profiles were characterized. Segmental duplication was the main reason for the expansion of the GmABC genes. Ka/Ks analysis suggested that intense purifying selection was accompanied by the evolution of GmABC genes. The genome-wide collinearity of soybean with other species showed that GmABCs were relatively conserved and that collinear ABCs between species may have originated from the same ancestor. Gene expression analysis of GmABCs revealed the distinct expression pattern in different tissues and diverse developmental stages. The candidate genes GmABCB23, GmABCB25, GmABCB48, GmABCB52, GmABCI1, GmABCI5, and GmABCI13 were responsive to Al toxicity. This work on the GmABC gene family provides useful information for future studies on ABC transporters in soybean and potential targets for the cultivation of new germplasm resources of aluminum-tolerant soybean.

Download Full-text

MetR-Regulated Vibrio cholerae Metabolism Is Required for Virulence

mBio ◽

10.1128/mbio.00236-12 ◽

2012 ◽

Vol 3 (5) ◽

Cited By ~ 28

Author(s):

Ryan W. Bogard ◽

Bryan W. Davies ◽

John J. Mekalanos

Keyword(s):

Amino Acid ◽

Vibrio Cholerae ◽

Virulence Factor ◽

Current Knowledge ◽

Intestinal Colonization ◽

Wild Type ◽

Pathogenic Potential ◽

Content Type ◽

Mouse Intestine

ABSTRACTLysR-type transcriptional regulators (LTTRs) are the largest, most diverse family of prokaryotic transcription factors, with regulatory roles spanning metabolism, cell growth and division, and pathogenesis. Using a sequence-defined transposon mutant library, we screened a panel ofV. choleraeEl Tor mutants to identify LTTRs required for host intestinal colonization. Surprisingly, out of 38 LTTRs, only one severely affected intestinal colonization in the suckling mouse model of cholera: the methionine metabolism regulator, MetR. Genetic analysis of genes influenced by MetR revealed thatglyA1andmetJwere also required for intestinal colonization. Chromatin immunoprecipitation of MetR and quantitative reverse transcription-PCR (qRT-PCR) confirmed interaction with and regulation ofglyA1, indicating that misregulation ofglyA1is likely responsible for the colonization defect observed in themetRmutant. TheglyA1mutant was auxotrophic for glycine but exhibited wild-type trimethoprim sensitivity, making folate deficiency an unlikely cause of its colonization defect. MetJ regulatory mutants are not auxotrophic but are likely altered in the regulation of amino acid-biosynthetic pathways, including those for methionine, glycine, and serine, and this misregulation likely explains its colonization defect. However, mutants defective in methionine, serine, and cysteine biosynthesis exhibited wild-type virulence, suggesting that these amino acids can be scavenged in vivo. Taken together, our results suggest that glycine biosynthesis may be required to alleviate an in vivo nutritional restriction in the mouse intestine; however, additional roles for glycine may exist. Irrespective of the precise nature of this requirement, this study illustrates the importance of pathogen metabolism, and the regulation thereof, as a virulence factor.IMPORTANCEVibrio choleraecontinues to be a severe cause of morbidity and mortality in developing countries. Identification ofV. choleraefactors critical to disease progression offers the potential to develop or improve upon therapeutics and prevention strategies. To increase the efficiency of virulence factor discovery, we employed a regulator-centric approach to multiplex our in vivo screening capabilities and allow whole regulons inV. choleraeto be interrogated for pathogenic potential. We identified MetR as a new virulence regulator and serine hydroxymethyltransferase GlyA1 as a new MetR-regulated virulence factor, both required byV. choleraeto colonize the infant mouse intestine. Bacterial metabolism is a prerequisite to virulence, and current knowledge of in vivo metabolism of pathogens is limited. Here, we expand the known role of amino acid metabolism and regulation in virulence and offer new insights into the in vivo metabolic requirements ofV. choleraewithin the mouse intestine.

Download Full-text

FlrA, a σ54-Dependent Transcriptional Activator in Vibrio fischeri, Is Required for Motility and Symbiotic Light-Organ Colonization

Journal of Bacteriology ◽

10.1128/jb.185.12.3547-3557.2003 ◽

2003 ◽

Vol 185 (12) ◽

pp. 3547-3557 ◽

Cited By ~ 47

Author(s):

Deborah S. Millikan ◽

Edward G. Ruby

Keyword(s):

Vibrio Fischeri ◽

Transcriptional Activator ◽

Pcr Analysis ◽

Euprymna Scolopes ◽

Wild Type ◽

Flagellar Regulon ◽

Colonization Factors ◽

Recognition Motifs ◽

Arbitrarily Primed Pcr ◽

Organ Colonization

ABSTRACT Flagellum-mediated motility of Vibrio fischeri is an essential factor in the bacterium's ability to colonize its host, the Hawaiian squid Euprymna scolopes. To begin characterizing the nature of the flagellar regulon, we have cloned a gene, designated flrA, from V. fischeri that encodes a putative σ54-dependent transcriptional activator. Genetic arrangement of the flrA locus in V. fischeri is similar to motility master-regulator operons of Vibrio cholerae and Vibrio parahaemolyticus. In addition, examination of regulatory regions of a number of flagellar operons in V. fischeri revealed apparent σ54 recognition motifs, suggesting that the flagellar regulatory hierarchy is controlled by a similar mechanism to that described in V. cholerae. However, in contrast to its closest known relatives, flrA mutant strains of V. fischeri ES114 were completely abolished in swimming capability. Although flrA provided in trans restored motility to the flrA mutant, the complemented strain was unable to reach wild-type levels of symbiotic colonization in juvenile squid, suggesting a possible role for the proper expression of FlrA in regulating symbiotic colonization factors in addition to those required for motility. Comparative RNA arbitrarily primed PCR analysis of the flrA mutant and its wild-type parent revealed several differentially expressed transcripts. These results define a regulon that includes both flagellar structural genes and other genes apparently not involved in flagellum elaboration or function. Thus, the transcriptional activator FlrA plays an essential role in regulating motility, and apparently in modulating other symbiotic functions, in V. fischeri.

Download Full-text