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1.  Draft Genome Sequence of Rhodococcus rhodnii Strain LMG5362, a Symbiont of Rhodnius prolixus (Hemiptera, Reduviidae, Triatominae), the Principle Vector of Trypanosoma cruzi 
Genome Announcements  2013;1(3):e00329-13.
We report the 4,385,577-bp high-quality draft assembly of the bacterial symbiont Rhodococcus rhodnii strain LMG5362, isolated from the gut of Rhodnius prolixus (Hemiptera, Reduviidae, Triatominae), the principle vector of the protozoan Trypanosoma cruzi, the etiological agent of Chagas disease. This sequence might provide useful information for subsequent studies of the symbiotic relationship between Rd. prolixus and Rc. rhodnii, while also providing a starting point for the development of biotechnological applications for the control of Rd. prolixus.
doi:10.1128/genomeA.00329-13
PMCID: PMC3707589  PMID: 23788540
2.  A Laterally Acquired Galactose Oxidase-Like Gene Is Required for Aerial Development during Osmotic Stress in Streptomyces coelicolor 
PLoS ONE  2013;8(1):e54112.
Phylogenetic reconstruction revealed that most Actinobacterial orthologs of S. coelicolor SCO2837, encoding a metal-dependent galactose oxidase-like protein, are found within Streptomyces and were probably acquired by horizontal gene transfer from fungi. Disruption of SCO2837 (glxA) caused a conditional bld phenotype that could not be reversed by extracellular complementation. Studies aimed at characterising the regulation of expression of glxA showed that it is not a target for other bld genes. We provide evidence that glxA is required for osmotic adaptation, although independently from the known osmotic stress response element SigB. glxA has been predicted to be part of an operon with the transcription unit comprising the upstream cslA gene and glxA. However, both phenotypic and expression studies indicate that it is also expressed from an independent promoter region internal to cslA. GlxA displays an in situ localisation pattern similar to that one observed for CslA at hyphal tips, but localisation of the former is independent of the latter. The functional role of GlxA in relation to CslA is discussed.
doi:10.1371/journal.pone.0054112
PMCID: PMC3543389  PMID: 23326581
4.  Forkhead‐associated proteins genetically linked to the serine/threonine kinase PknB regulate carbon flux towards antibiotic biosynthesis in Streptomyces coelicolor 
Microbial Biotechnology  2011;4(2):263-274.
Summary
To date, the function of only two of the 34 predicted serine/threonine protein kinases (STPKs) of Streptomyces coelicolor has been described. Here we report functional analysis of pknB and two linked genes, fhaAB, encoding forkhead‐associated (FHA) domain proteins that are part of a highly conserved gene locus in actinobacteria. In contrast to the homologous gene of Mycobacterium tuberculosis, pknB in S. coelicolor is not essential and has no apparent role in defining cell shape. Phosphorylation of recombinant forms of both the full‐length protein and N‐terminal kinase domain suggest that PknB‐mediated signalling in S. coelicolor may be modulated by another factor(s). FhaAB are candidate interacting partners of PknB and loss of their function resulted in deregulation of central carbon metabolism, with carbon flux diverted to synthesis of the antibiotic actinorhodin. The substrate hyphae of the fhaAB mutant also exhibited an unusual cording morphology. The results indicate that inactivation of FHA ‘brake’ proteins can potentially amplify the function of STPKs and, in this case, provide a means to overproduce antibiotics.
doi:10.1111/j.1751-7915.2010.00237.x
PMCID: PMC3818866  PMID: 21342471
5.  FtsW Is a Dispensable Cell Division Protein Required for Z-Ring Stabilization during Sporulation Septation in Streptomyces coelicolor▿ † 
Journal of Bacteriology  2008;190(16):5555-5566.
The conserved rodA and ftsW genes encode polytopic membrane proteins that are essential for bacterial cell elongation and division, respectively, and each gene is invariably linked with a cognate class B high-molecular-weight penicillin-binding protein (HMW PBP) gene. Filamentous differentiating Streptomyces coelicolor possesses four such gene pairs. Whereas rodA, although not its cognate HMW PBP gene, is essential in these bacteria, mutation of SCO5302 or SCO2607 (sfr) caused no gross changes to growth and septation. In contrast, disruption of either ftsW or the cognate ftsI gene blocked the formation of sporulation septa in aerial hyphae. The inability of spiral polymers of FtsZ to reorganize into rings in aerial hyphae of these mutants indicates an early pivotal role of an FtsW-FtsI complex in cell division. Concerted assembly of the complete divisome was unnecessary for Z-ring stabilization in aerial hyphae as ftsQ mutants were found to be blocked at a later stage in cell division, during septum closure. Complete cross wall formation occurred in vegetative hyphae in all three fts mutants, indicating that the typical bacterial divisome functions specifically during nonessential sporulation septation, providing a unique opportunity to interrogate the function and dependencies of individual components of the divisome in vivo.
doi:10.1128/JB.00398-08
PMCID: PMC2519378  PMID: 18556789
6.  Characterization of Changes to the Cell Surface during the Life Cycle of Streptomyces coelicolor: Atomic Force Microscopy of Living Cells▿  
Journal of Bacteriology  2006;189(6):2219-2225.
Cell surface changes that accompany the complex life cycle of Streptomyces coelicolor were monitored by atomic force microscopy (AFM) of living cells. Images were obtained using tapping mode to reveal that young, branching vegetative hyphae have a relatively smooth surface and are attached to an inert silica surface by means of a secreted extracellular matrix. Older hyphae, representing a transition between substrate and aerial growth, are sparsely decorated with fibers. Previously, a well-organized stable mosaic of fibers, called the rodlet layer, coating the surface of spores has been observed using electron microscopy. AFM revealed that aerial hyphae, prior to sporulation, possess a relatively unstable dense heterogeneous fibrous layer. Material from this layer is shed as the hyphae mature, revealing a more tightly organized fibrous mosaic layer typical of spores. The aerial hyphae are also characterized by the absence of the secreted extracellular matrix. The formation of sporulation septa is accompanied by modification to the surface layer, which undergoes localized temporary disruption at the sites of cell division. The characteristics of the hyphal surfaces of mutants show how various chaplin and rodlin proteins contribute to the formation of fibrous layers of differing stabilities. Finally, older spores with a compact rodlet layer develop surface concavities that are attributed to a reduction of intracellular turgor pressure as metabolic activity slows.
doi:10.1128/JB.01470-06
PMCID: PMC1899363  PMID: 17194797
7.  Influence of CrgA on Assembly of the Cell Division Protein FtsZ during Development of Streptomyces coelicolor 
Journal of Bacteriology  2006;188(4):1540-1550.
The product of the crgA gene of Streptomyces coelicolor represents a novel family of small proteins. A single orthologous gene is located close to the origin of replication of all fully sequenced actinomycete genomes and borders a conserved gene cluster implicated in cell growth and division. In S. coelicolor, CrgA is important for coordinating growth and cell division in sporogenic hyphae. In this study, we demonstrate that CrgA is an integral membrane protein whose peak expression is coordinated with the onset of development of aerial hyphae. The protein localizes to discrete foci away from growing hyphal tips. Upon overexpression, CrgA localizes to apical syncytial cells of aerial hyphae and inhibits the formation of productive cytokinetic rings of the bacterial tubulin homolog FtsZ, leading to proteolytic turnover of this major cell division determinant. In the absence of known prokaryotic cell division inhibitors in actinomycetes, CrgA may have an important conserved function influencing Z-ring formation in these bacteria.
doi:10.1128/JB.188.4.1540-1550.2006
PMCID: PMC1367258  PMID: 16452438
8.  The Product of a Developmental Gene, crgA, That Coordinates Reproductive Growth in Streptomyces Belongs to a Novel Family of Small Actinomycete-Specific Proteins 
Journal of Bacteriology  2003;185(22):6678-6685.
On solid media, the reproductive growth of Streptomyces involves antibiotic biosynthesis coincident with the erection of filamentous aerial hyphae. Following cessation of growth of an aerial hypha, multiple septation occurs at the tip to form a chain of unigenomic spores. A gene, crgA, that coordinates several aspects of this reproductive growth is described. The gene product is representative of a well-conserved family of small actinomycete proteins with two C-terminal hydrophobic-potential membrane-spanning segments. In Streptomyces avermitilis, crgA is required for sporulation, and inactivation of the gene abolished most sporulation septation in aerial hyphae. Disruption of the orthologous gene in Streptomyces coelicolor indicates that whereas CrgA is not essential for sporulation in this species, during growth on glucose-containing media, it influences the timing of the onset of reproductive growth, with precocious erection of aerial hyphae and antibiotic production by the mutant. Moreover, CrgA subsequently acts to inhibit sporulation septation prior to growth arrest of aerial hyphae. Overexpression of CrgA in S. coelicolor, uncoupling any nutritional and growth phase-dependent regulation, results in growth of nonseptated aerial hyphae on all media tested, consistent with a role for the protein in inhibiting sporulation septation.
doi:10.1128/JB.185.22.6678-6685.2003
PMCID: PMC262101  PMID: 14594842
9.  VGJφ, a Novel Filamentous Phage of Vibrio cholerae, Integrates into the Same Chromosomal Site as CTXφ 
Journal of Bacteriology  2003;185(19):5685-5696.
We describe a novel filamentous phage, designated VGJφ, isolated from strain SG25-1 of Vibrio cholerae O139, which infects all O1 (classical and El Tor) and O139 strains tested. The sequence of the 7,542 nucleotides of the phage genome reveals that VGJφ has a distinctive region of 775 nucleotides and a conserved region with an overall genomic organization similar to that of previously characterized filamentous phages, such as CTXφ of V. cholerae and Ff phages of Escherichia coli. The conserved region carries 10 open reading frames (ORFs) coding for products homologous to previously reported peptides of other filamentous phages, and the distinctive region carries one ORF whose product is not homologous to any known peptide. VGJφ, like other filamentous phages, uses a type IV pilus to infect V. cholerae; in this case, the pilus is the mannose-sensitive hemagglutinin. VGJφ-infected V. cholerae overexpresses the product of one ORF of the phage (ORF112), which is similar to single-stranded DNA binding proteins of other filamentous phages. Once inside a cell, VGJφ is able to integrate its genome into the same chromosomal attB site as CTXφ, entering into a lysogenic state. Additionally, we found an attP structure in VGJφ, which is also conserved in several lysogenic filamentous phages from different bacterial hosts. Finally, since different filamentous phages seem to integrate into the bacterial dif locus by a general mechanism, we propose a model in which repeated integration events with different phages might have contributed to the evolution of the CTX chromosomal region in V. cholerae El Tor.
doi:10.1128/JB.185.19.5685-5696.2003
PMCID: PMC193952  PMID: 13129939
10.  The dpsA Gene of Streptomyces coelicolor: Induction of Expression from a Single Promoter in Response to Environmental Stress or during Development 
PLoS ONE  2011;6(9):e25593.
The DpsA protein plays a dual role in Streptomyces coelicolor, both as part of the stress response and contributing to nucleoid condensation during sporulation. Promoter mapping experiments indicated that dpsA is transcribed from a single, sigB-like dependent promoter. Expression studies implicate SigH and SigB as the sigma factors responsible for dpsA expression while the contribution of other SigB-like factors is indirect by means of controlling sigH expression. The promoter is massively induced in response to osmotic stress, in part due to its sensitivity to changes in DNA supercoiling. In addition, we determined that WhiB is required for dpsA expression, particularly during development. Gel retardation experiments revealed direct interaction between apoWhiB and the dpsA promoter region, providing the first evidence for a direct WhiB target in S. coelicolor.
doi:10.1371/journal.pone.0025593
PMCID: PMC3184153  PMID: 21984935

Results 1-10 (10)