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P Dion

Publications and source records attributed to P Dion.

At least 19 recordsLinked to original sources
PubMed1996

Influence of alkyltransferase activity and chromosomal locus on mutational hotspots in Chinese hamster ovary cells.

High-density mutational spectra have been established for exon 3 of the gene encoding adenine phosphoribosyltransferase (APRT) of the Chinese hamster ovary (CHO) cell line derivative D422 and closely related and/or modified lines by using the mutagen ethyl methanesulfonate (EMS). The total number of selectable sites (GC-->AT transitions yielding a selectable APRT- phenotype) was estimated at 31 based on our own accumulated data base of 136 sequenced exon 3 mutations and on literature reports. D422 and two other APRT hemizygous lines each yielded very similar spectra and showed two populations of mutable sites: (i) 24 "baseline" sites that followed the Poisson distribution and therefore were equally susceptible to mutation and (ii) two hotspots, one comprising a cluster at nucleotides 1293-1309 and the other at nucleotide 1365. Collectively, the latter sites were about 10-fold more frequently mutated than the others. CHO cells are mer- as they lack the repair enzyme O6-methylguanidine methyltransferase (EC 2.1.1.63). In modified repair-proficient CHO cells, the distribution of mutations among all of the 31 sites was random, with only 3 of the 19 GC-->AT transitions in the above hotspots. To determine whether the distribution was locus-dependent, two independent lines carrying single copies of transfected APRT genes were generated from a derivative of D422 carrying a deletion in the endogenous APRT gene. Nucleotides 1293-1309 were again no longer preferentially mutated, but the site at nucleotide 1365 was still a hotspot. We conclude that mutational spectra in mer- cells are at least in part locus dependent and that some sequences are particularly susceptible to EMS mutagenesis and perhaps also to methyltransferase repair.

Adenine Phosphoribosyltransferase↗
PubMed1995

Genetic analysis of nonpathogenic Agrobacterium tumefaciens mutants arising in crown gall tumors.

Little is known about the effect of the host on the genetic stability of bacterial plant pathogens. Crown gall, a plant disease caused by Agrobacterium tumefaciens, may represent a useful model to study this effect. Indeed, our previous observations on the natural occurrence and origin of nonpathogenic agrobacteria suggest that the host plant might induce loss of pathogenicity in populations of A. tumefaciens. Here we report that five different A. tumefaciens strains initially isolated from apple tumors produced up to 99% nonpathogenic mutants following their reintroduction into axenic apple plants. Two of these five strains were also found to produce mutants on pear and/or blackberry plants. Generally, the mutants of the apple isolate D10B/87 were altered in the tumor-inducing plasmid, harboring either deletions in this plasmid or point mutations in the regulatory virulence gene virG. Most of the mutants originating from the same tumor appeared to be of clonal origin, implying that the host plants influenced agrobacterial populations by favoring growth of nonpathogenic mutants over that of wild-type cells. This hypothesis was confirmed by coinoculation of apple rootstocks with strain D10B/87 and a nonpathogenic mutant.

Acetophenones↗
PubMed1994

Novel narrow-host-range vectors for direct cloning of foreign DNA in Pseudomonas.

Narrow-host-range vectors, based on an indigenous replicon and containing a multiple cloning site, have been constructed in a Pseudomonas host capable of growth on unusual substrates. The new cloning vectors yield sufficient amounts of DNA for preparative purposes and belong to an incompatibility group different from that of the incP and incQ broad-host-range vectors. One of these vectors, named pDB47F, was used to clone, directly in Pseudomonas, DNA fragments from Agrobacterium, Pseudomonas, and Rhizobium. A clone containing Agrobacterium and KmR gene sequences was transformed with a higher efficiency than an RSF1010-derived vector (by as much as 1250-fold) in four out of five Pseudomonas strains tested. The considerable efficiency obtained with this system makes possible the direct cloning and phenotypic selection of foreign DNA in Pseudomonas.

Base Sequence↗
PubMed1994

Effect of ionizing dose rate on the radioresistance of some food pathogenic bacteria.

Food pathogenic bacteria including Listeria monocytogenes (1A1 and ATCC 19111), Staphylococcus aureus (GD13 and ATCC 13565), Escherichia coli O157:H7 (ATCC 35150), Salmonella typhimurium, Yersinia enterocolitica, Vibrio parahaemolyticus, and Campylobacter jejuni were exposed to various rates of ionizing radiation (0.78, 2.6, and 22 kGy/h) emitted by three different 60Co irradiators. D10 values (D10 is the radiation dose required to eliminate 90% of a bacterial population (one logarithmic cycle reduction)) were calculated for the various strains and growth conditions tested. A covariance analysis of these results revealed that the dose rates studied had no significant influence on the radiosensitivity of these bacteria. At all dose rates, the bacteria were more radiosensitive when irradiated in a saline solution (0.85% NaCl) than in a chicken breast meat suspension. The growth phase of the bacterial population had a variable influence on its radioresistance. For L. monocytogenes 1A1, Staphylococcus aureus ATCC 13565, E. coli O157:H7, Y. enterocolitica, and V. parahaemolyticus, radioresistance was not significantly different in the exponential and stationary phases. Populations of L. monocytogenes ATCC 19111 and Staphylococcus aureus GD13 were significantly more resistant in the stationary phase (D10 = 0.23 and 0.12 kGy, respectively) than in the exponential phase (D10 = 0.17 and 0.09 kGy, respectively). Among the pathogenic bacteria investigated in this study, the most radioresistant was L. monocytogenes (D10 = 0.16-0.38 kGy, Gram-positive bacilli) and the most radiosensitive was V. parahaemolyticus (D10 = 0.03-0.04 kGy, halophilic Gram-negative bacilli).

Animals↗
PubMed1994

Identification of variability of ribosomal DNA spacer from Pseudomonas soil isolates.

The polymerase chain reaction was used to amplify the spacer region located between the 16S and 23S ribosomal RNA genes of strains of Pseudomonas fluorescens and Pseudomonas putida isolated from peat bog, canola field, or arctic plants. Some of spacer region of four of the P. fluorescens strains examined, strains 64-3, 63-28, QP5, and R17-FP2, was about 515 base pairs (bp) in length, and contained the genes for tRNA(Ile) and tRNA(Ala). The DNA sequences of two strains from canola, 64-3 and 63-28, differed at only two positions. The sequences of the peat bog strains QP5 and R17-FP2 were identical. However, differences were noted between the DNA sequence common to the pair of strains 64-3 and 63-28 and the corresponding common sequence for strains QP5 and R17-FP2. These differences were mainly concentrated in two DNA segments of 10 and 19 bp, respectively. A probe for the 19-bp variable segment that occurs in the ribosomal spacer of strains QP5 and R17-FP2 recognized total DNA from these two strains, but not DNA from other bacteria of different origins. These results suggest the existence of a limited degree of variability within the 16S-23S ribosomal DNA spacer region, and that this variability may be useful to the recognition of particular Pseudomonas strains from environmental samples.

Base Sequence↗
PubMed1993

Dynamic structure of Agrobacterium tumefaciens Ti plasmids.

Agrobacterium tumefaciens C58F is a variant of strain C58 which generates a high proportion of avirulent mutants in the presence of the virulence (vir) gene inducer acetosyringone. These mutants are altered in the Ti plasmid and do not respond to the acetosyringone signal (C. Fortin, E. W. Nester, and P. Dion, J. Bacteriol. 174:5676-5685, 1992). The physical organization of the Ti plasmid was compared in strain C58 and its variant. One feature distinguishing pTiC58F from its parent plasmid was the presence of the insertion element IS426. Three copies of this element were detected in the strain C58 chromosome, whereas two additional copies were found in strain C58F, including one copy in the Ti plasmid. This particular copy of IS426 was associated with the region of arginine and nopaline catabolism of pTiC58F. Most of the avirulent mutants recovered following growth of strain C58F in the presence of acetosyringone were complemented by clones carrying either virA or virG. Element IS426 was no longer found in the arginine and nopaline catabolism region of the Ti plasmids from the virA and virG mutants, but it resided in the particular KpnI fragment containing the modified vir locus. Behavior of a strain C58F derivative, which was inactivated in a chromosomal component required for the response to acetosyringone, was consistent with the possibility that vir gene induction is essential to the massive production of avirulent mutants.

Acetophenones↗
PubMed1992

Growth inhibition and loss of virulence in cultures of Agrobacterium tumefaciens treated with acetosyringone.

Acetosyringone, a phenolic inducer of the virulence (vir) genes of Agrobacterium tumefaciens, inhibited the growth of the nopaline-type strains T37 and C58 incubated under acidic conditions. In the course of a 6-day incubation with acetosyringone, avirulent clones were produced in different proportions by strains T37 and C58 and also by a spontaneous variant of strain C58, denominated C58F. The proportion of avirulent clones in acetosyringone-treated cultures often exceeded 50% for strains T37 and C58F and was of the order of 1% for strain C58. Control cultures not exposed to acetosyringone did not yield avirulent clones. Two other vir inducers, sinapinic acid and syringaldehyde, also inhibited growth and promoted accumulation of avirulent clones in cultures of strains C58F and T37. On the other hand, various acetosyringone analogs reported not to induce the vir genes did not act as growth inhibitors. All of the T37 and most of the C58F avirulent clones examined still carried a Ti plasmid. In all instances examined, avirulent clones still carrying a Ti plasmid were mutated in this plasmid. Mutants of strain C58F lacked the capacity to induce a virB::lacZ fusion in the presence of acetosyringone.

Acetophenones↗
PubMed1992

Diauxic growth of Agrobacterium tumefaciens 15955 on succinate and mannopine.

Diauxic growth was observed upon incubation of Agrobacterium tumefaciens 15955 on a mixture of succinate and mannopine as the carbon source. Diauxic growth was also observed when either fumarate or L-malate was mixed with mannopine. No diauxie was detectable when A. tumefaciens 15955 was grown on a mixture of mannopine and glucose, fructose, sucrose, or L-arabinose. Preferential utilization of succinate was observed in the initial growth phase of diauxie, whereas the final growth phase occurred at the expense of mannopine. Cells harvested during the initial growth phase exhibited a capacity for uptake of [14C]succinate but not of [14C] mannopine. A capacity for [14C]mannopine uptake was expressed during the final growth phase. Extracts from cells grown on a mixture of succinate and mannopine exhibited a low level of mannopine cyclase activity in the initial phase of diauxie. This activity increased substantially in the final phase of growth. Added succinate had no effect on the rate of [14C]mannopine uptake or mannopine cyclase activities of cells previously grown on mannopine. Diauxie was also observed during growth of strain 15955 on a mixture of succinate and octopine.

Agrobacterium tumefaciens↗
PubMed1992

Spontaneous mutation conferring the ability to catabolize mannopine in Agrobacterium tumefaciens.

Two nopaline-type strains of Agrobacterium tumefaciens, C58 and T37, as well as strain A136, which is a Ti plasmid-cured derivative of strain C58, gave rise to spontaneous mutants that were able to grow on mannopine. The observation of mutagenesis with strain A136 demonstrated that the ability to acquire this new catabolic potential was independent of the presence of a Ti plasmid. The mutants were isolated after 4 weeks of incubation on minimal medium containing mannopine as the sole carbon source. They also utilized mannopinic acid, but not agropine or agropinic acid. In addition, the spontaneous mutant LM136, but not its parent strain A136, degraded many mannityl opine analogs. [14C]mannopine disappeared in the presence of LM136 cells which had been pregrown on opine or nonopine substrates. These results suggested that the catabolic system of this mutant was not subject to a stringent regulation. A clone conferring the ability to utilize mannopine on a recipient pseudomonad was selected from a genomic library from both the mutant LM136 and its parent strain. Only the LM136 clone was expressed in the parent Agrobacterium strain A136. Southern analysis showed that the genes for mannopine catabolism in the spontaneous mutants differed from the corresponding Ti plasmid-encoded genes of octopine-type or agropine-type Agrobacterium strains. Cells of LM136 utilized [14C]mannopine without generating detectable amounts of intracellular agropine. In contrast, a major fraction of the radioactivity recovered from cells of the octopine-type strain Ach5, after incubation on [14C]mannopine, was in the form of agropine.

Agrobacterium tumefaciens↗
PubMed1991

Mannopine and mannopinic acid as substrates for Arthrobacter sp. strain MBA209 and Pseudomonas putida NA513.

The characteristics of mannopine and mannopinic acid utilization by Agrobacterium tumefaciens B6S3, Arthrobacter sp. strain MBA209, and Pseudomonas putida NA513 were studied. Strain B6S3 utilized the four mannityl opines, mannopine, mannopinic acid, agropine, and agropinic acid. It also utilized several mannityl opine analogs, which were modified in either the sugar or the amino acid moiety. It utilized mannopine more rapidly after preincubation on mannopine, mannopinic acid, or glutamine than after pregrowth on glucose, mannose, or mannitol. Strains MBA209 and NA513 utilized mannopine and mannopinic acid, but not the other two mannityl opines. They utilized few mannityl opine analogs, sometimes because of failure to utilize the products of initial cleavage of the analog. Utilization of mannopine and mannopinic acid by strain NA513 was strictly dependent on prior growth on these substrates. A spontaneous regulatory variant of strain NA513 remained unable to utilize most of the mannityl opine analogs. Glutamine, mannose, and several analogs had no inhibitory effect on [14C]mannopine utilization by strain NA513.

Arthrobacter↗
PubMed1990

Tumor necrosis factor mediated cytolysis requires the adenovirus E1a protein but not the transformed phenotype.

Adenovirus transformed cells are susceptible to lysis by human recombinant tumor necrosis factor (TNF). This susceptibility correlates with the presence of E1a in these cells. A flat revertant cell line which expresses a biologically functional E1a but not the transformed phenotype was nevertheless susceptible to TNF. However, flat revertants retransformed by 5-azacytidine, without concomitant reactivation of E1a, were resistant to TNF-alpha. This result suggests TNF susceptibility is not transformation but E1a dependent. To study the mechanism of cytolysis in these cell lines, we examined the possibility that changes in the transcription of E1a were brought about by TNF, as it was reported in the case of a c-myc transformed cell line. The results showed that TNF did not affect either E1a or c-myc transcription in our cells during the development of the cytotoxic response.

Adenoviridae↗
PubMed1989

Non-p-glycoprotein-mediated multidrug resistance in detransformed rat cells selected for resistance to methylglyoxal bis(guanylhydrazone).

Three independent variants (G2, G4, G5), resistant to methylglyoxal bis(guanylhydrazone), an anticancer drug, have been isolated by single step selection from an adenovirus-transformed rat brain cell line (1). These variants display selective cross-resistance to several natural product drugs of dissimilar structure and action. Multidrug resistance has recently been shown to be caused by overexpression of the membrane-associated p-glycoprotein, most often caused by amplification of the mdr gene. Several types of experiments were conducted to determine whether the observed drug resistance in our cell lines could be due to changes at the mdr locus. The following results were obtained: (a) the mdr locus was not amplified; (b) transcription of the mdr gene and p-glycoprotein synthesis were not increased; (c) multidrug resistance cell lines, which carry an amplified mdr locus, were not cross-resistant to methylglyoxal bis(guanylhydrazone); (d) verapamil did not reverse the resistance of G cells or mdr cells to methylglyoxal bis(guanylhydrazone), nor that of G cells to vincristine; and (e) methylglyoxal bis(guanylhydrazone) resistance was recessive and depended on a block to drug uptake, as opposed to mdr cells which are dominant and express increased drug efflux. The results obtained suggest that the drug resistance in the G2, G4, and G5 cells was atypical and may be due to a mechanism distinct from that mediated by the mdr locus.

ATP Binding Cassette Transporter, Subfamily B, Mem↗
PubMed1988

A randomized trial of vitamins C and E in the prevention of recurrence of colorectal polyps.

Because supplements of vitamins C and E had been associated with reduction of fecal mutagen levels, a double-blind randomized trial was designed to examine the effects of these vitamins on the rate of recurrence of colorectal polyps, presumed precursors for colorectal cancer. Two hundred patients believed to be free of polyps after removal of at least one colorectal polyp were randomized to receive a supplement of 400 mg each of ascorbic acid and alpha-tocopherol, or a placebo. Fifteen patients had to be excluded because a review of pathology indicated that their polyps were not adenomatous. A second colonoscopic examination was planned after 2 yr of supplementation. One hundred thirty-seven people (75% of eligible subjects) completed the study; polyps were observed in the second colonoscopy in 41.4% of 70 subjects on vitamin supplements and in 50.7% of 67 subjects on placebos. After adjustment for differences between groups in demographic and dietary factors before study entry, the relative risk of polyp occurrence was 0.86, with 95% confidence limits from 0.51 to 1.45, in an analysis of 129 subjects with complete information on demographic and dietary factors who had completed the trial. Of the 48 patients who had not completed the study, 7 had died, 33 had not returned to their physician for an examination, and 8 had had a follow-up colonoscopy or sigmoidoscopy. Inclusion of the three polyps found in these eight examinations led to an estimate of relative risk of 0.86 (95% confidence limits, 0.51 to 1.43). The findings of this investigation suggest that any reduction in the rate of polyp recurrence associated with vitamin supplementation is small, and a larger study would be required to ensure that an effect of this size was not a chance finding.

Ascorbic Acid↗
PubMed1988

Construction of a Tn5 derivative encoding bioluminescence and its introduction in Pseudomonas, Agrobacterium and Rhizobium.

A simple method based upon the use of a Tn5 derivative, Tn5-Lux, has been devised for the introduction and stable expression of the character of bioluminescence in a variety of gram-negative bacteria. In Tn5-Lux, the luxAB genes of Vibrio harveyi encoding luciferase are inserted on a SalI--BglII fragment between the kanamycin resistance (Kmr) gene and the right insertion sequence. The transposon derivative was placed on a transposition suicide vehicle by in situ recombination with the Tn5 suicide vector pGS9, to yield pDB30. Mating between Escherichia coli WA803 (pDB30) and a strain from our laboratory, Pseudomonas sp. RB100C, gave a Kmr transfer frequency of 10(-6) per recipient, a value 10 times lower than that obtained with the original suicide vehicle pGS9. Tn5-Lux was also introduced by insertion mutagenesis in other strains of gram-negative soil bacteria. The bioluminescence marker was expressed in the presence of n-decanal, and was monitored as chemiluminescence in a liquid scintillation counter. The recorded light intensities were fairly comparable among the strains, and ranged between 0.2 to 1.8 x 10(6) cpm for a cell density of 10(3) colony forming units/ml. Nodules initiated by bioluminescent strains of Rhizobium leguminosarum on two different hosts were compared for intensity of the bioluminescence they produced.

DNA Transposable Elements↗
PubMed1983

Mutagenicity of different fractions of extracts of human feces.

An extraction-fractionation scheme for the isolation of non-volatile fecal mutagens is described. Extraction of feces was with acetone, and a 2-step fractionation scheme employing silica gel SepPak cartridges and normal-phase high-pressure liquid chromatography was used. Assay of mutagens was with the standard plate mutagenicity assay with Salmonella typhimurium tester strains TA98 and TA100, with and without the Aroclor induced S9 microsomal activation system. Single feces samples from 24 donors from a wide socioeconomic spectrum were tested. It was found that most fecal mutagenicity extracted by aqueous acetone could be attributed to a lipid soluble mutagen active on both TA98 and TA100 that has been previously reported.

Chromatography↗
PubMed1980

Germination properties as marker events characterizing later stages of Bacillus subtilis spore formation.

At various stages during spore formation sporangia were shocked by cold treatment or with toluene, and the germination requirements of the prespores were examined. Up to 5 h after induction of sporulation (t5) germination was spontaneous; i.e., it occurred without any added germinants. After t5, during stages V and VI, the capacity for spontaneous germination diminished progressively, and the spores acquired a need for externally added germinants. At t6 this need was satisfied by either L-alanine or a mixture of KCl, glucose, and fructose. By t8, the latter response had disappeared. The spores germinated only with L-alanine, and the response was much slower. Experiments with chloramphenicol showed that the germination properties of the spores appearing between t6 and t8 were the expression of events in protein synthesis that had occurred before t5. Although the germination requirements developed at about the same time as heat resistance, they could be dissociated from heat resistance in wild-type and mutant cells. The germination properties of the developing spores are additional marker events characterizing the later stages of sporulation, as follows: (i) spontaneous germination (up to the end of stage IV); (ii) germination requirements that are satisfied by KCl-glucose-fructose or L-alanine (stage V); and (iii) slow germination response with L-alanine only (stage VI).

Alanine↗