scholarly journals Importance of Proteins Controlling Initiation of DNA Replication in the Growth of the High-Pressure-Loving Bacterium Photobacterium profundum SS9

2009 ◽  
Vol 191 (20) ◽  
pp. 6383-6393 ◽  
Author(s):  
Ziad W. El-Hajj ◽  
Theodora Tryfona ◽  
David J. Allcock ◽  
Fariha Hasan ◽  
Federico M. Lauro ◽  
...  
Keyword(s):  
High Pressure ◽  
Dna Replication ◽  
Deep Sea ◽  
Negative Regulator ◽  
Cold Sensitivity ◽  
E Coli ◽  
Positive Regulator ◽  
Initiation Of Dna Replication ◽  
Photobacterium Profundum ◽  
Insight Into

ABSTRACT The molecular mechanism(s) by which deep-sea bacteria grow optimally under high hydrostatic pressure at low temperatures is poorly understood. To gain further insight into the mechanism(s), a previous study screened transposon mutant libraries of the deep-sea bacterium Photobacterium profundum SS9 and identified mutants which exhibited alterations in growth at high pressure relative to that of the parent strain. Two of these mutants, FL23 (PBPRA3229::mini-Tn10) and FL28 (PBPRA1039::mini-Tn10), were found to have high-pressure sensitivity and enhanced-growth phenotypes, respectively. The PBPRA3229 and PBPRA1039 genes encode proteins which are highly similar to Escherichia coli DiaA, a positive regulator, and SeqA, a negative regulator, respectively, of the initiation of DNA replication. In this study, we investigated the hypothesis that PBPRA3229 and PBPRA1039 encode DiaA and SeqA homologs, respectively. Consistent with this, we determined that the plasmid-carried PBPRA3229 and PBPRA1039 genes restored synchrony to the initiation of DNA replication in E. coli mutants lacking DiaA and SeqA, respectively. Additionally, PBPRA3229 restored the cold sensitivity phenotype of an E. coli dnaA(Cs) diaA double mutant whereas PBPRA1039 suppressed the cold sensitivity phenotype of an E. coli dnaA(Cs) single mutant. Taken together, these findings show that the genes disrupted in FL23 and FL28 encode DiaA and SeqA homologs, respectively. Consequently, our findings add support to a model whereby high pressure affects the initiation of DNA replication in P. profundum SS9 and either the presence of a positive regulator (DiaA) or the removal of a negative regulator (SeqA) promotes growth under these conditions.

scholarly journals RecD Function Is Required for High-Pressure Growth of a Deep-Sea Bacterium

1999 ◽  
Vol 181 (8) ◽  
pp. 2330-2337 ◽  
Author(s):  
Kelly A. Bidle ◽  
Douglas H. Bartlett
Keyword(s):  
High Pressure ◽  
Deep Sea ◽  
Genomic Library ◽  
Plasmid Stability ◽  
Stop Codon ◽  
Dna Recombination ◽  
Repair Gene ◽  
E Coli ◽  
Pressure Sensitive ◽  
Photobacterium Profundum

ABSTRACT A genomic library derived from the deep-sea bacteriumPhotobacterium profundum SS9 was conjugally delivered into a previously isolated pressure-sensitive SS9 mutant, designated EC1002 (E. Chi and D. H. Bartlett, J. Bacteriol. 175:7533–7540, 1993), and exconjugants were screened for the ability to grow at 280-atm hydrostatic pressure. Several clones were identified that had restored high-pressure growth. The complementing DNA was localized and in all cases found to possess strong homology torecD, a DNA recombination and repair gene. EC1002 was found to be deficient in plasmid stability, a phenotype also seen inEscherichia coli recD mutants. The defect in EC1002 was localized to a point mutation that created a stop codon within the recD gene. Two additional recDmutants were constructed by gene disruption and were both found to possess a pressure-sensitive growth phenotype, although the magnitude of the defect depended on the extent of 3′ truncation of the recD coding sequence. Surprisingly, the introduction of the SS9 recD gene into an E. coli recD mutant had two dramatic effects. At high pressure, SS9recD enabled growth in the E. coli mutant strain under conditions of plasmid antibiotic resistance selection and prevented cell filamentation. Both of these effects were recessive to wild-type E. coli recD. These results suggest that the SS9recD gene plays an essential role in SS9 growth at high pressure and that it may be possible to identify additional aspects of RecD function through the characterization of this activity.

scholarly journals FabF Is Required for Piezoregulation ofcis-Vaccenic Acid Levels and Piezophilic Growth of the Deep-Sea Bacterium Photobacterium profundum Strain SS9

2000 ◽  
Vol 182 (5) ◽  
pp. 1264-1271 ◽  
Author(s):  
Eric E. Allen ◽  
Douglas H. Bartlett
Keyword(s):  
High Pressure ◽  
Hydrostatic Pressure ◽  
Deep Sea ◽  
Unsaturated Fatty Acids ◽  
Acyl Carrier Protein ◽  
Vaccenic Acid ◽  
Bacterial Species ◽  
Elevated Pressure ◽  
Wild Type ◽  
Photobacterium Profundum

ABSTRACT To more fully explore the role of unsaturated fatty acids in high-pressure, low-temperature growth, the fabF gene from the psychrotolerant, piezophilic deep-sea bacteriumPhotobacterium profundum strain SS9 was characterized and its role and regulation were examined. An SS9 strain harboring a disruption in the fabF gene (strain EA40) displayed growth impairment at elevated hydrostatic pressure concomitant with diminishedcis-vaccenic acid (18:1) production. However, growth ability at elevated pressure could be restored to wild-type levels by the addition of exogenous 18:1 to the growth medium. Transcript analysis did not indicate that the SS9 fabF gene is transcriptionally regulated, suggesting that the elevated 18:1 levels produced in response to pressure increase result from posttranscriptional changes. Unlike many pressure-adapted bacterial species such as SS9, the mesophile Escherichia coli did not regulate its fatty acid composition in an adaptive manner in response to changes in hydrostatic pressure. Moreover, an E. coli fabF strain was as susceptible to elevated pressure as wild-type cells. It is proposed that the SS9 fabF product, β-ketoacyl–acyl carrier protein synthase II has evolved novel pressure-responsive characteristics which facilitate SS9 growth at high pressure.

scholarly journals Selective Inhibition by Pantoyl Lactone and Butyl Alcohol of the Initiation of DNA Replication in E. coli

1973 ◽  
Vol 37 (6) ◽  
pp. 1317-1320
Author(s):  
Yoshihiro YOSHIYAMA ◽  
Kazuo NAGAI ◽  
Hideaki SOME ◽  
Gakuzo TAMURA
Keyword(s):  
Dna Replication ◽  
Selective Inhibition ◽  
Butyl Alcohol ◽  
E Coli ◽  
Initiation Of Dna Replication

scholarly journals Molecular cloning of mtp70, a mitochondrial member of the hsp70 family.

1989 ◽  
Vol 9 (11) ◽  
pp. 5163-5168 ◽  
Author(s):  
D M Engman ◽  
L V Kirchhoff ◽  
J E Donelson
Keyword(s):  
Mitochondrial Dna ◽  
Dna Replication ◽  
Protozoan Parasite ◽  
Specific Region ◽  
Kinetoplast Dna ◽  
Intracellular Location ◽  
E Coli ◽  
Hsp70 Family ◽  
Mitochondrial Dna Replication ◽  
Initiation Of Dna Replication

We have isolated a gene from the protozoan parasite Trypanosoma cruzi that encodes a previously unidentified member of the 70-kilodalton heat shock protein (hsp70) family. Among all the eucaryotic hsp70 proteins described to date, this trypanosome protein, mtp70, is uniquely related in sequence and structure to the hsp70 of Escherichia coli, DnaK, which functions in the initiation of DNA replication. This relationship to DnaK is especially relevant in view of the intracellular location of the protein. Within the trypanosome, mtp70 is located in the mitochondrion, where it associates with kinetoplast DNA (kDNA), the unusual mitochondrial DNA that distinguishes this order of protozoa. Moreover, mtp70 is located in the specific region of the kinetoplast in which kDNA replication occurs. In view of the known functions of DnaK, the localization of mtp70 to the site of kDNA replication suggests that mtp70 may participate in eucaryotic mitochondrial DNA replication in a manner analogous to that of DnaK in E. coli.

Molecular cloning of mtp70, a mitochondrial member of the hsp70 family

1989 ◽  
Vol 9 (11) ◽  
pp. 5163-5168
Author(s):  
D M Engman ◽  
L V Kirchhoff ◽  
J E Donelson
Keyword(s):  
Mitochondrial Dna ◽  
Dna Replication ◽  
Protozoan Parasite ◽  
Specific Region ◽  
Kinetoplast Dna ◽  
Intracellular Location ◽  
E Coli ◽  
Hsp70 Family ◽  
Mitochondrial Dna Replication ◽  
Initiation Of Dna Replication

We have isolated a gene from the protozoan parasite Trypanosoma cruzi that encodes a previously unidentified member of the 70-kilodalton heat shock protein (hsp70) family. Among all the eucaryotic hsp70 proteins described to date, this trypanosome protein, mtp70, is uniquely related in sequence and structure to the hsp70 of Escherichia coli, DnaK, which functions in the initiation of DNA replication. This relationship to DnaK is especially relevant in view of the intracellular location of the protein. Within the trypanosome, mtp70 is located in the mitochondrion, where it associates with kinetoplast DNA (kDNA), the unusual mitochondrial DNA that distinguishes this order of protozoa. Moreover, mtp70 is located in the specific region of the kinetoplast in which kDNA replication occurs. In view of the known functions of DnaK, the localization of mtp70 to the site of kDNA replication suggests that mtp70 may participate in eucaryotic mitochondrial DNA replication in a manner analogous to that of DnaK in E. coli.

scholarly journals The Deep-Sea Bacterium Photobacterium profundum SS9 Utilizes Separate Flagellar Systems for Swimming and Swarming under High-Pressure Conditions

2008 ◽  
Vol 74 (20) ◽  
pp. 6298-6305 ◽  
Author(s):  
Emiley A. Eloe ◽  
Federico M. Lauro ◽  
Rudi F. Vogel ◽  
Douglas H. Bartlett
Keyword(s):  
High Pressure ◽  
Gene Cluster ◽  
Deep Sea ◽  
Gene Clusters ◽  
Maximum Pressure ◽  
Swimming Velocity ◽  
Flagellar Motor ◽  
Lateral Flagellum ◽  
Photobacterium Profundum ◽  
Flagellum Gene

ABSTRACT Motility is a critical function needed for nutrient acquisition, biofilm formation, and the avoidance of harmful chemicals and predators. Flagellar motility is one of the most pressure-sensitive cellular processes in mesophilic bacteria; therefore, it is ecologically relevant to determine how deep-sea microbes have adapted their motility systems for functionality at depth. In this study, the motility of the deep-sea piezophilic bacterium Photobacterium profundum SS9 was investigated and compared with that of the related shallow-water piezosensitive strain Photobacterium profundum 3TCK, as well as that of the well-studied piezosensitive bacterium Escherichia coli. The SS9 genome contains two flagellar gene clusters: a polar flagellum gene cluster (PF) and a putative lateral flagellum gene cluster (LF). In-frame deletions were constructed in the two flagellin genes located within the PF cluster (flaA and flaC), the one flagellin gene located within the LF cluster (flaB), a component of a putative sodium-driven flagellar motor (motA2), and a component of a putative proton-driven flagellar motor (motA1). SS9 PF flaA, flaC, and motA2 mutants were defective in motility under all conditions tested. In contrast, the flaB and motA1 mutants were defective only under conditions of high pressure and high viscosity. flaB and motA1 gene expression was strongly induced by elevated pressure plus increased viscosity. Direct swimming velocity measurements were obtained using a high-pressure microscopic chamber, where increases in pressure resulted in a striking decrease in swimming velocity for E. coli and a gradual reduction for 3TCK which proceeded up to 120 MPa, while SS9 increased swimming velocity at 30 MPa and maintained motility up to a maximum pressure of 150 MPa. Our results indicate that P. profundum SS9 possesses two distinct flagellar systems, both of which have acquired dramatic adaptations for optimal functionality under high-pressure conditions.

scholarly journals Large-Scale Transposon Mutagenesis of Photobacterium profundum SS9 Reveals New Genetic Loci Important for Growth at Low Temperature and High Pressure

2007 ◽  
Vol 190 (5) ◽  
pp. 1699-1709 ◽  
Author(s):  
Federico M. Lauro ◽  
Khiem Tran ◽  
Alessandro Vezzi ◽  
Nicola Vitulo ◽  
Giorgio Valle ◽  
...  
Keyword(s):  
High Pressure ◽  
Low Temperature ◽  
Deep Sea ◽  
Large Scale ◽  
High Pressures ◽  
Cell Envelope ◽  
Transposon Mutagenesis ◽  
Chromosomal Structure ◽  
And Function ◽  
Photobacterium Profundum

ABSTRACT Microorganisms adapted to piezopsychrophilic growth dominate the majority of the biosphere that is at relatively constant low temperatures and high pressures, but the genetic bases for the adaptations are largely unknown. Here we report the use of transposon mutagenesis with the deep-sea bacterium Photobacterium profundum strain SS9 to isolate dozens of mutant strains whose growth is impaired at low temperature and/or whose growth is altered as a function of hydrostatic pressure. In many cases the gene mutation-growth phenotype relationship was verified by complementation analysis. The largest fraction of loci associated with temperature sensitivity were involved in the biosynthesis of the cell envelope, in particular the biosynthesis of extracellular polysaccharide. The largest fraction of loci associated with pressure sensitivity were involved in chromosomal structure and function. Genes for ribosome assembly and function were found to be important for both low-temperature and high-pressure growth. Likewise, both adaptation to temperature and adaptation to pressure were affected by mutations in a number of sensory and regulatory loci, suggesting the importance of signal transduction mechanisms in adaptation to either physical parameter. These analyses were the first global analyses of genes conditionally required for low-temperature or high-pressure growth in a deep-sea microorganism.

scholarly journals Differential inhibition of the initiation of DNA replication in stringent and relaxed strains ofEscherichia coli

1988 ◽  
Vol 51 (3) ◽  
pp. 173-177 ◽  
Author(s):  
Elena C. Guzman ◽  
Francisco J. Carrillo ◽  
Alfonso Jimenez-Sanchez
Keyword(s):  
Amino Acids ◽  
Protein Synthesis ◽  
Dna Replication ◽  
Ofescherichia Coli ◽  
Differential Inhibition ◽  
E Coli ◽  
Pbr322 Dna ◽  
Initiation Of Dna Replication ◽  
Do So

SummaryStarvation for isoleucine inhibits chromosome, minichromosome and pBR322 DNA replication in a stringent strain ofE. coli, but does not do so in a relaxed mutant. Starvation for other amino acids inhibits either chromosome and minichromosome replication in both strains. From these results we conclude thatoriCand pBR322 replication are stringently regulated and that isoleucine seems not to be essential for the protein synthesis required at the initiation oforiCreplication. Deprivation of isoleucine in a Rel−strain gives rise to amplification of minichromosome and pBR322 with a better yield of the latter plasmid than that following treatment with chloramphenicol.

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