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John R Battista

Publications and source records attributed to John R Battista.

15 recordsLinked to original sources
PubMed2005

Truepera radiovictrix gen. nov., sp. nov., a new radiation resistant species and the proposal of Trueperaceae fam. nov.

Two isolates, belonging to a new species of a novel genus of the Phylum "Deinococcus/Thermus ", were recovered from hot spring runoffs on the Island of São Miguel in the Azores. Strains RQ-24(T) and TU-8 are the first cultured representatives of a distinct phylogenetic lineage within this phylum. These strains form orange/red colonies, spherical-shaped cells, have an optimum growth temperature of about 50 degrees C, an optimum pH for growth between about 7.5 and 9.5, and do not grow at pH below 6.5 or above pH 11.2. These organisms grow in complex media without added NaCl, but have a maximum growth rate in media with 1.0% NaCl and grow in media containing up to 6.0% NaCl. The organisms are extremely ionizing radiation resistant; 60% of the cells survive 5.0 kGy. These strains are chemoorganotrophic and aerobic; do not grow in Thermus medium under anaerobic conditions with or without nitrate as electron acceptor and glucose as a source of carbon and energy, but ferment glucose to D-lactate without formation of gas. The organisms assimilate a large variety of sugars, organic acids and amino acids. Fatty acids are predominantly iso- and anteiso-branched; long chain 1,2 diols were also found in low relative proportions; menaquinone 8 (MK-8) is the primary respiratory quinone. Peptidoglycan was not detected. Based on 16S rRNA gene sequence analysis, physiological, biochemical and chemical analysis we describe a new species of one novel genus represented by strain RQ-24(T) (CIP 108686(T)=LMG 22925(T)=DSM 17093(T)) for which we propose the name Truepera radiovictrix. We also propose the family Trueperaceae fam. nov. to accommodate this new genus.

Amino Acids↗
PubMed2005

A ring-like nucleoid is not necessary for radioresistance in the Deinococcaceae.

BACKGROUND: Transmission electron microscopy images of Deinococcus radiodurans R1 suggest that the nucleoid of this species exists as a "ring-like" body, and have led to speculation that this structure contributes to the radioresistance of the species. Since extreme radioresistance is characteristic of six other species of Deinococcus, we have attempted to correlate nucleoid morphology and radioresistance by determining whether the genomic DNA of each of these species exhibit similar structures. RESULTS: The nucleoid morphologies of seven recognized species of Deinococcus, the radioresistant bacterium Rubrobacter radiotolerans, and the more radiosensitive deinococcal relative Thermus aquaticus were evaluated using epifluorescence and deconvolution techniques. Although the nucleoids of Deinococcus murrayi, Deinococcus proteolyticus, Deinococcus radiophilus, and Deinococcus grandis have structures similar to D. radiodurans, the majority of nucleoids found in Deinococcus radiopugnans and Deinococcus geothermalis lack any specific organization. The nucleoid of R. radiotolerans consists of multiple highly condensed spheres of DNA scattered throughout the cell. The genomic DNA of Thermus aquaticus is uniformly distributed throughout the cell. CONCLUSION: There is no obvious relationship between the shape of a species' nucleoid and extreme radioresistance. However, the genomes of all extremely radioresistance species examined are highly condensed relative to more radiosensitive species. Whether DNA in this tightly packed configuration contributes to the radioresistance of these bacteria remains unknown, but this common structural feature appears to limit diffusion of fragments generated post-irradiation even in cells incapable of repairing strand breaks.

DNA, Bacterial↗
PubMed2005

Deinococcus radiodurans - the consummate survivor.

Relatively little is known about the biochemical basis of the capacity of Deinococcus radiodurans to endure the genetic insult that results from exposure to ionizing radiation and can include hundreds of DNA double-strand breaks. However, recent reports indicate that this species compensates for extensive DNA damage through adaptations that allow cells to avoid the potentially detrimental effects of DNA strand breaks. It seems that D. radiodurans uses mechanisms that limit DNA degradation and that restrict the diffusion of DNA fragments that are produced following irradiation, to preserve genetic integrity. These mechanisms also increase the efficiency of the DNA-repair proteins.

DNA Repair↗
PubMed2005

HspR is a global negative regulator of heat shock gene expression in Deinococcus radiodurans.

The HspR protein functions as a negative regulator of chaperone and protease gene expression in a diversity of bacteria. Here we have identified, cloned and deleted the Deinococcus radiodurans HspR homologue, DR0934. Delta hspR mutants exhibit moderate growth defects when shifted to mild heat shock temperatures, but are severely impaired for survival at 48 degrees C. Using quantitative reverse transcription polymerase chain reaction and global transcriptional analysis, we have identified 14 genes that are derepressed in the absence of stress in the delta hspR background, 11 of which encode predicted chaperones and proteases, including dnaKJgrpE, ftsH, lonB, hsp20 and clpB. Promoter mapping indicated that the transcription of these genes initiates from a promoter bearing a sigma70-type consensus, and that putative HspR binding sites (HAIR) were present in the 5'-untranslated regions. Electrophoretic mobility shift assays indicated that HspR binds to these promoters at the HAIR site in vitro. These results strongly suggest that DR0934 encodes the HspR-like global negative regulator of D. radiodurans that directly represses chaperone and protease gene expression by binding to the HAIR site in close proximity to promoter regions.

5' Untranslated Regions↗
PubMed2005

Extensive diversity of ionizing-radiation-resistant bacteria recovered from Sonoran Desert soil and description of nine new species of the genus Deinococcus obtained from a single soil sample.

The ionizing-radiation-resistant fractions of two soil bacterial communities were investigated by exposing an arid soil from the Sonoran Desert and a nonarid soil from a Louisiana forest to various doses of ionizing radiation using a (60)Co source. The numbers of surviving bacteria decreased as the dose of gamma radiation to which the soils were exposed increased. Bacterial isolates surviving doses of 30 kGy were recovered from the Sonoran Desert soil, while no isolates were recovered from the nonarid forest soil after exposure to doses greater than 13 kGy. The phylogenetic diversities of the surviving culturable bacteria were compared for the two soils using 16S rRNA gene sequence analysis. In addition to a bacterial population that was more resistant to higher doses of ionizing radiation, the diversity of the isolates was greater in the arid soil. The taxonomic diversity of the isolates recovered was found to decrease as the level of ionizing-radiation exposure increased. Bacterial isolates of the genera Deinococcus, Geodermatophilus, and Hymenobacter were still recovered from the arid soil after exposure to doses of 17 to 30 kGy. The recovery of large numbers of extremely ionizing-radiation-resistant bacteria from an arid soil and not from a nonarid soil provides further ecological support for the hypothesis that the ionizing-radiation resistance phenotype is a consequence of the evolution of other DNA repair systems that protect cells against commonly encountered environmental stressors, such as desiccation. The diverse group of bacterial strains isolated from the arid soil sample included 60 Deinococcus strains, the characterization of which revealed nine novel species of this genus.

DNA, Bacterial↗
PubMed2005

Global transcriptional and proteomic analysis of the Sig1 heat shock regulon of Deinococcus radiodurans.

The sig1 gene, predicted to encode an extracytoplasmic function-type heat shock sigma factor of Deinococcus radiodurans, has been shown to play a central role in the positive regulation of the heat shock operons groESL and dnaKJ. To determine if Sig1 is required for the regulation of additional heat shock genes, we monitored the global transcriptional and proteomic profiles of a D. radiodurans R1 sig1 mutant and wild-type cells in response to elevated temperature stress. Thirty-one gene products were identified that showed heat shock induction in the wild type but not in the sig1 mutant. Quantitative real-time PCR experiments verified the transcriptional requirement of Sig1 for the heat shock induction of the mRNA of five of these genes-dnaK, groES, DR1314, pspA, and hsp20. hsp20 appears to encode a new member of the small heat shock protein superfamily, DR1314 is predicted to encode a hypothetical protein with no recognizable orthologs, and pspA is predicted to encode a protein involved in maintenance of membrane integrity. Deletion mutation analysis demonstrated the importance in heat shock protection of hsp20 and DR1314. The promoters of dnaKJE, groESL, DR1314, pspA, and hsp20 were mapped and, combined with computer-based pattern searches of the upstream regions of the 26 other Sig1 regulon members, these results suggested that Sig1 might recognize both sigma70-type and sigma(W)-type promoter consensus sequences. These results expand the D. radiodurans Sig1 heat shock regulon to include 31 potential new members, including not only factors with cytoplasmic functions, such as groES and dnaK, but also those with extracytoplasmic functions, like pspA.

Bacterial Proteins↗
PubMed2004

Preserving genome integrity: the DdrA protein of Deinococcus radiodurans R1.

The bacterium Deinococcus radiodurans can withstand extraordinary levels of ionizing radiation, reflecting an equally extraordinary capacity for DNA repair. The hypothetical gene product DR0423 has been implicated in the recovery of this organism from DNA damage, indicating that this protein is a novel component of the D. radiodurans DNA repair system. DR0423 is a homologue of the eukaryotic Rad52 protein. Following exposure to ionizing radiation, DR0423 expression is induced relative to an untreated control, and strains carrying a deletion of the DR0423 gene exhibit increased sensitivity to ionizing radiation. When recovering from ionizing-radiation-induced DNA damage in the absence of nutrients, wild-type D. radiodurans reassembles its genome while the mutant lacking DR0423 function does not. In vitro, the purified DR0423 protein binds to single-stranded DNA with an apparent affinity for 3' ends, and protects those ends from nuclease degradation. We propose that DR0423 is part of a DNA end-protection system that helps to preserve genome integrity following exposure to ionizing radiation. We designate the DR0423 protein as DNA damage response A protein.

Bacterial Proteins↗
PubMed2004

Developing a genetic system in Deinococcus radiodurans for analyzing mutations.

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 beta-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.

Amino Acid Sequence↗
PubMed2004

Cysteinyl-tRNA(Cys) formation in Methanocaldococcus jannaschii: the mechanism is still unknown.

Most organisms form Cys-tRNA(Cys), an essential component for protein synthesis, through the action of cysteinyl-tRNA synthetase (CysRS). However, the genomes of Methanocaldococcus jannaschii, Methanothermobacter thermautotrophicus, and Methanopyrus kandleri do not contain a recognizable cysS gene encoding CysRS. It was reported that M. jannaschii prolyl-tRNA synthetase (C. Stathopoulos, T. Li, R. Longman, U. C. Vothknecht, H. D. Becker, M. Ibba, and D. Söll, Science 287:479-482, 2000; R. S. Lipman, K. R. Sowers, and Y. M. Hou, Biochemistry 39:7792-7798, 2000) or the M. jannaschii MJ1477 protein (C. Fabrega, M. A. Farrow, B. Mukhopadhyay, V. de Crécy-Lagard, A. R. Ortiz, and P. Schimmel, Nature 411:110-114, 2001) provides the "missing" CysRS activity for in vivo Cys-tRNA(Cys) formation. These conclusions were supported by complementation of temperature-sensitive Escherichia coli cysS(Ts) strain UQ818 with archaeal proS genes (encoding prolyl-tRNA synthetase) or with the Deinococcus radiodurans DR0705 gene, the ortholog of the MJ1477 gene. Here we show that E. coli UQ818 harbors a mutation (V27E) in CysRS; the largest differences compared to the wild-type enzyme are a fourfold increase in the K(m) for cysteine and a ninefold reduction in the k(cat) for ATP. While transformants of E. coli UQ818 with archaeal and bacterial cysS genes grew at a nonpermissive temperature, growth was also supported by elevated intracellular cysteine levels, e.g., by transformation with an E. coli cysE allele (encoding serine acetyltransferase) or by the addition of cysteine to the culture medium. An E. coli cysS deletion strain permitted a stringent complementation test; growth could be supported only by archaeal or bacterial cysS genes and not by archaeal proS genes or the D. radiodurans DR0705 gene. Construction of a D. radiodurans DR0705 deletion strain showed this gene to be dispensable. However, attempts to delete D. radiodurans cysS failed, suggesting that this is an essential Deinococcus gene. These results imply that it is not established that proS or MJ1477 gene products catalyze Cys-tRNA(Cys) synthesis in M. jannaschii. Thus, the mechanism of Cys-tRNA(Cys) formation in M. jannaschii still remains to be discovered.

Amino Acyl-tRNA Synthetases↗
PubMed2004

Analysis of Deinococcus radiodurans's transcriptional response to ionizing radiation and desiccation reveals novel proteins that contribute to extreme radioresistance.

During the first hour after a sublethal dose of ionizing radiation, 72 genes were upregulated threefold or higher in D. radiodurans R1. Thirty-three of these loci were also among a set of 73 genes expressed in R1 cultures recovering from desiccation. The five transcripts most highly induced in response to each stress are the same and encode proteins of unknown function. The genes (ddrA, ddrB, ddrC, ddrD, and pprA) corresponding to these transcripts were deleted, both alone and in all possible two-way combinations. Characterization of the mutant strains defines three epistasis groups that reflect different cellular responses to ionizing radiation-induced damage. The ddrA and ddrB gene products have complementary activities and inactivating both loci generates a strain that is more sensitive to ionizing radiation than strains in which either single gene has been deleted. These proteins appear to mediate efficient RecA-independent processes connected to ionizing radiation resistance. The pprA gene product is not necessary for homologous recombination during natural transformation, but nevertheless may participate in a RecA-dependent process during recovery from radiation damage. These characterizations clearly demonstrate that novel mechanisms significantly contribute to the ionizing radiation resistance in D. radiodurans.

Deinococcus↗
PubMed2003

Using DNA microarray data to understand the ionizing radiation resistance of Deinococcus radiodurans.

In a recent paper, Liu et al. documented the changes in gene expression as stationary phase Deinococcus radiodurans cultures recover from acute exposure to gamma radiation. Given that the biochemical details of the response of D. radiodurans to ionizing radiation are poorly understood, this work represents an important first step towards achieving an understanding of the ionizing radiation resistance in this species.

Computational Biology↗
PubMed2002

Global analysis of the Deinococcus radiodurans proteome by using accurate mass tags.

Understanding biological systems and the roles of their constituents is facilitated by the ability to make quantitative, sensitive, and comprehensive measurements of how their proteome changes, e.g., in response to environmental perturbations. To this end, we have developed a high-throughput methodology to characterize an organism's dynamic proteome based on the combination of global enzymatic digestion, high-resolution liquid chromatographic separations, and analysis by Fourier transform ion cyclotron resonance mass spectrometry. The peptides produced serve as accurate mass tags for the proteins and have been used to identify with high confidence >61% of the predicted proteome for the ionizing radiation-resistant bacterium Deinococcus radiodurans. This fraction represents the broadest proteome coverage for any organism to date and includes 715 proteins previously annotated as either hypothetical or conserved hypothetical.

Bacteria↗
PubMed2002

The IrrE protein of Deinococcus radiodurans R1 is a novel regulator of recA expression.

IRS24 is a DNA damage-sensitive strain of Deinococcus radiodurans strain 302 carrying a mutation in an uncharacterized locus designated irrE. Five overlapping cosmids capable of restoring ionizing radiation resistance to IRS24 were isolated from a genomic library. The ends of each cloned insert were sequenced, and these sequences were used to localize irrE to a 970-bp region on chromosome I of D. radiodurans R1. The irrE gene corresponds to coding sequence DR0167 in the R1 genome. The irrE gene encodes a 35,000-Da protein that has no similarity to any previously characterized peptide. The irrE locus of R1 was also inactivated by transposon mutagenesis, and this strain was sensitive to ionizing radiation, UV light, and mitomycin C. Preliminary findings indicate that IrrE is a novel regulatory protein that stimulates transcription of the recA gene following exposure to ionizing radiation.

Bacterial Proteins↗
PubMed2002

Genetic evidence that the uvsE gene product of Deinococcus radiodurans R1 is a UV damage endonuclease.

An in vitro transposition system, developed to facilitate gene disruption in Deinococcus radiodurans R1, has been used to inactivate the gene designated dr1819 in uvrA-1(+) and uvrA-1 backgrounds. dr1819 encodes a protein with homology to a UV DNA damage endonuclease expressed by Schizosaccharomyces pombe. Interruption of dr1819 greatly sensitizes the uvrA-1 strain but not the uvrA-1(+) strain to UV light, indicating that the dr1819 gene product is a component in a DNA repair pathway that can compensate for the loss of nucleotide excision repair in this species. Clones of dr1819 will restore UV resistance to UVS78, a uvrA-1 uvsE strain, indicating that dr1819 and uvsE are the same locus.

Bacterial Proteins↗