Journal articles: 'Bacteriophages – Molecular genetics; Genetic transcription'

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Collin, Alejandro, Araceli González-Jiménez, María del Carmen González-Jiménez, Manuel J. Alfonso, and Olga Calvo. "The Role of S. cerevisiae Sub1/PC4 in Transcription Elongation Depends on the C-Terminal Region and Is Independent of the ssDNA Binding Domain." Cells 11, no. 20 (October 21, 2022): 3320. http://dx.doi.org/10.3390/cells11203320.

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Saccharomyces cerevisiae Sub1 (ScSub1) has been defined as a transcriptional stimulatory protein due to its homology to the ssDNA binding domain (ssDBD) of human PC4 (hPC4). Recently, PC4/Sub1 orthologues have been elucidated in eukaryotes, prokaryotes, and bacteriophages with functions related to DNA metabolism. Additionally, ScSub1 contains a unique carboxyl–terminal region (CT) of unknown function up to date. Specifically, it has been shown that Sub1 is required for transcription activation, as well as other processes, throughout the transcription cycle. Despite the progress that has been made in understanding the mechanism underlying Sub1′s functions, some questions remain unanswered. As a case in point: whether Sub1’s roles in initiation and elongation are differentially predicated on distinct regions of the protein or how Sub1′s functions are regulated. Here, we uncover some residues that are key for DNA–ScSub1 interaction in vivo, localized in the ssDBD, and required for Sub1 recruitment to promoters. Furthermore, using an array of genetic and molecular techniques, we demonstrate that the CT region is required for transcription elongation by RNA polymerase II (RNAPII). Altogether, our data indicate that Sub1 plays a dual role during transcription—in initiation through the ssDBD and in elongation through the CT region.

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Tyler, Jessica S., Melissa J. Mills, and David I. Friedman. "The Operator and Early Promoter Region of the Shiga Toxin Type 2-Encoding Bacteriophage 933W and Control of Toxin Expression." Journal of Bacteriology 186, no. 22 (November 15, 2004): 7670–79. http://dx.doi.org/10.1128/jb.186.22.7670-7679.2004.

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ABSTRACT The genes encoding Shiga toxin (Stx), the major virulence factor of Shiga toxin-producing Escherichia coli, are carried in the genomes of bacteriophages that belong to the lambdoid family of phages. Previous studies demonstrated that induction of prophages encoding stx significantly enhances the production and/or release of Stx from the bacterium. Therefore, factors that regulate the switch between lysogeny and lytic growth, e.g., repressor, operator sites, and associated phage promoters, play important roles in regulating the production and/or release of Stx. We report the results of genetic and biochemical studies characterizing these elements of the Stx-encoding bacteriophage 933W. Like λ, 933W has three operator repeats in the right operator region (O R), but unlike λ and all other studied lambdoid phages, which have three operator repeats in the left operator region (O L), 933W only has two operator repeats in O L. As was observed with λ, the 933W O R and O L regions regulate transcription from the early P R and P L promoters, respectively. A lysogen carrying a 933W derivative encoding a noncleavable repressor fails to produce Stx, unlike a lysogen carrying a 933W derivative encoding a cleavable repressor. This finding provides direct evidence that measurable expression of the stx genes encoded by a 933W prophage requires induction of that prophage with the concomitant initiation of phage gene expression.

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Kropinski, Andrew M. "Phage Therapy -- Everything Old Is New again." Canadian Journal of Infectious Diseases and Medical Microbiology 17, no. 5 (2006): 297–306. http://dx.doi.org/10.1155/2006/329465.

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The study of bacterial viruses (bacteriophages or phages) proved pivotal in the nascence of the disciplines of molecular biology and microbial genetics, providing important information on the central processes of the bacterial cell (DNA replication, transcription and translation) and on how DNA can be transferred from one cell to another. As a result of the pioneering genetics studies and modern genomics, it is now known that phages have contributed to the evolution of the microbial cell and to its pathogenic potential. Because of their ability to transmit genes, phages have been exploited to develop cloning vector systems. They also provide a plethora of enzymes for the modern molecular biologist. Until the introduction of antibiotics, phages were used to treat bacterial infections (with variable success). Western science is now having to re-evaluate the application of phage therapy -- a therapeutic modality that never went out of vogue in Eastern Europe -- because of the emergence of an alarming number of antibiotic-resistant bacteria. The present article introduces the reader to phage biology, and the benefits and pitfalls of phage therapy in humans and animals.

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Niu, Guoqing, Toshinori Okinaga, Fengxia Qi, and Justin Merritt. "The Streptococcus mutans IrvR Repressor Is a CI-Like Regulator That Functions through Autocleavage and Clp-Dependent Proteolysis." Journal of Bacteriology 192, no. 6 (December 28, 2009): 1586–95. http://dx.doi.org/10.1128/jb.01261-09.

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ABSTRACT Previous work has shown that irvR is required for the proper regulation of genetic competence and dextran-dependent aggregation due to its ability to repress the transcription regulator irvA. In this study, we determined the mechanism used to relieve the repression of irvA. We demonstrate that IrvR is a “LexA-like” protein with four conserved amino acid residues likely required for IrvR autocleavage activity. Furthermore, recombinant IrvR protein purified from Escherichia coli was competent to undergo autocleavage in vitro. Using several truncated IrvR constructs, we show that the amino acids adjacent to the autocleavage site are essential for relieving irvA repression and engaging the irvA-dependent regulatory pathway primarily through the ClpXP and ClpCP proteases. By extending the IrvR C terminus with an epitope derived from the autocleavage site, we were also able to create a constitutive Clp-dependent degradation of the full-length IrvR protein. This suggests that the derepression of irvA occurs through a two-step mechanism involving the initial autocleavage of IrvR and exposure of a proteolytic degradation sequence followed by Clp-dependent degradation of the IrvR DNA binding domain. Thus, irvA derepression is highly analogous to the genetic switch mechanism used to regulate lysogeny in bacteriophages.

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Chatterjee, Anushila, Julia L. E. Willett, Gary M. Dunny, and Breck A. Duerkop. "Phage infection and sub-lethal antibiotic exposure mediate Enterococcus faecalis type VII secretion system dependent inhibition of bystander bacteria." PLOS Genetics 17, no. 1 (January 7, 2021): e1009204. http://dx.doi.org/10.1371/journal.pgen.1009204.

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Bacteriophages (phages) are being considered as alternative therapeutics for the treatment of multidrug resistant bacterial infections. Considering phages have narrow host-ranges, it is generally accepted that therapeutic phages will have a marginal impact on non-target bacteria. We have discovered that lytic phage infection induces transcription of type VIIb secretion system (T7SS) genes in the pathobiont Enterococcus faecalis. Membrane damage during phage infection induces T7SS gene expression resulting in cell contact dependent antagonism of different Gram positive bystander bacteria. Deletion of essB, a T7SS structural component, abrogates phage-mediated killing of bystanders. A predicted immunity gene confers protection against T7SS mediated inhibition, and disruption of its upstream LXG toxin gene rescues growth of E. faecalis and Staphylococcus aureus bystanders. Phage induction of T7SS gene expression and bystander inhibition requires IreK, a serine/threonine kinase, and OG1RF_11099, a predicted GntR-family transcription factor. Additionally, sub-lethal doses of membrane targeting and DNA damaging antibiotics activated T7SS expression independent of phage infection, triggering T7SS antibacterial activity against bystander bacteria. Our findings highlight how phage infection and antibiotic exposure of a target bacterium can affect non-target bystander bacteria and implies that therapies beyond antibiotics, such as phage therapy, could impose collateral damage to polymicrobial communities.

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Zaworski, Julie, Colleen McClung, Cristian Ruse, Peter R. Weigele, Roger W. Hendrix, Ching-Chung Ko, Robert Edgar, Graham F. Hatfull, Sherwood R. Casjens, and Elisabeth A. Raleigh. "Genome analysis of Salmonella enterica serovar Typhimurium bacteriophage L, indicator for StySA (StyLT2III) restriction-modification system action." G3 Genes|Genomes|Genetics 11, no. 1 (December 22, 2020): 1–10. http://dx.doi.org/10.1093/g3journal/jkaa037.

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Abstract Bacteriophage L, a P22-like phage of Salmonella enterica sv Typhimurium LT2, was important for definition of mosaic organization of the lambdoid phage family and for characterization of restriction-modification systems of Salmonella. We report the complete genome sequences of bacteriophage L cI–40 13–am43 and L cII–101; the deduced sequence of wildtype L is 40,633 bp long with a 47.5% GC content. We compare this sequence with those of P22 and ST64T, and predict 72 Coding Sequences, 2 tRNA genes and 14 intergenic rho-independent transcription terminators. The overall genome organization of L agrees with earlier genetic and physical evidence; for example, no secondary immunity region (immI: ant, arc) or known genes for superinfection exclusion (sieA and sieB) are present. Proteomic analysis confirmed identification of virion proteins, along with low levels of assembly intermediates and host cell envelope proteins. The genome of L is 99.9% identical at the nucleotide level to that reported for phage ST64T, despite isolation on different continents ∼35 years apart. DNA modification by the epigenetic regulator Dam is generally incomplete. Dam modification is also selectively missing in one location, corresponding to the P22 phase-variation-sensitive promoter region of the serotype-converting gtrABC operon. The number of sites for SenLTIII (StySA) action may account for stronger restriction of L (13 sites) than of P22 (3 sites).

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Borghetti, Ivo Alberto, Miriam Ribas Zambenedetti, Luciana Requião, Deusilene Souza Vieira, Marco Aurélio Krieger, and Rita de Cássia Pontello Rampazzo. "External Control Viral-Like Particle Construction for Detection of Emergent Arboviruses by Real-Time Reverse-Transcription PCR." BioMed Research International 2019 (October 7, 2019): 1–4. http://dx.doi.org/10.1155/2019/2560401.

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Arboviruses have been emerging and reemerging worldwide, predominantly in tropical and subtropical areas. As many arbovirus infections, including dengue (DENV), Zika (ZIKV), and chikungunya (CHIKV), have similar signs and symptoms, clinical diagnosis of arbovirus infections is challenging. Therefore, reliable laboratory tests are necessary to improve the clinical management of patients with suspected arbovirus infections. Real-time reverse-transcription PCR (RT-qPCR) is among the more effective methods to distinguish these viruses. The aim of this study was to construct a unique positive external control derived from a unique plasmid using genetic engineering for specific use in RT-qPCR assays to detect Zika, dengue (1–4), and chikungunya. An external control derived from the MS2 bacteriophage was constructed using sequences from arbovirus and human genomes. Laboratories were asked to test the control in the ZDC Biomol kit, a RT-qPCR kit which is able to detect Zika, dengue serotypes 1–4, chikungunya, and an internal human control. RNA extracted from the external control was able to be amplified and detected in RT-qPCR assays for each virus detected by using the ZDC Biomol kit. The external control, samples from viral culture, and infected patient samples display similar amplification using this assay. The pET47b(+)MS2-ZDC vector is a viable expression system for the production of external control viral-like particles (MS2-ZDC). The RNA from the recombinant particles can be easily extracted and can function as a tool to validate all steps of process from the extraction to the amplification of all targets in specific reaction. Thus, the MS2-ZDC particles are laboratory-safe in order to avoid risk for operators, and the phages are effective as positive control for use in the ZDC Biomol kit amplifying all kit targets making them effective for commercial profile.

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Godinho, Lia M., Mehdi El Sadek Fadel, Céline Monniot, Lina Jakutyte, Isabelle Auzat, Audrey Labarde, Karima Djacem, et al. "The Revisited Genome of Bacillus subtilis Bacteriophage SPP1." Viruses 10, no. 12 (December 11, 2018): 705. http://dx.doi.org/10.3390/v10120705.

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Bacillus subtilis bacteriophage SPP1 is a lytic siphovirus first described 50 years ago [1]. Its complete DNA sequence was reported in 1997 [2]. Here we present an updated annotation of the 44,016 bp SPP1 genome and its correlation to different steps of the viral multiplication process. Five early polycistronic transcriptional units encode phage DNA replication proteins and lysis functions together with less characterized, mostly non-essential, functions. Late transcription drives synthesis of proteins necessary for SPP1 viral particles assembly and for cell lysis, together with a short set of proteins of unknown function. The extensive genetic, biochemical and structural biology studies on the molecular mechanisms of SPP1 DNA replication and phage particle assembly rendered it a model system for tailed phages research. We propose SPP1 as the reference species for a new SPP1-like viruses genus of the Siphoviridae family.

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Wang, Pauline W., Linda Chu, and David S. Guttman. "Complete Sequence and Evolutionary Genomic Analysis of the Pseudomonas aeruginosa Transposable Bacteriophage D3112." Journal of Bacteriology 186, no. 2 (January 15, 2004): 400–410. http://dx.doi.org/10.1128/jb.186.2.400-410.2004.

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ABSTRACT Bacteriophage D3112 represents one of two distinct groups of transposable phage found in the clinically relevant, opportunistic pathogen Pseudomonas aeruginosa. To further our understanding of transposable phage in P. aeruginosa, we have sequenced the complete genome of D3112. The genome is 37,611 bp, with an overall G+C content of 65%. We have identified 53 potential open reading frames, including three genes (the c repressor gene and early genes A and B) that have been previously characterized and sequenced. The organization of the putative coding regions corresponds to published genetic and transcriptional maps and is very similar to that of enterobacteriophage Mu. In contrast, the International Committee on Taxonomy of Viruses has classified D3112 as a λ-like phage on the basis of its morphology. Similarity-based analyses identified 27 open reading frames with significant matches to proteins in the NCBI databases. Forty-eight percent of these were similar to Mu-like phage and prophage sequences, including proteins responsible for transposition, transcriptional regulation, virion morphogenesis, and capsid formation. The tail proteins were highly similar to prophage sequences in Escherichia coli and phage Phi12 from Staphylococcus aureus, while proteins at the right end were highly similar to proteins in Xylella fastidiosa. We performed phylogenetic analyses to understand the evolutionary relationships of D3112 with respect to Mu-like versus λ-like bacteriophages. Different results were obtained from similarity-based versus phylogenetic analyses in some instances. Overall, our findings reveal a highly mosaic structure and suggest that extensive horizontal exchange of genetic material played an important role in the evolution of D3112.

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Cicero, Marco P., Meghan M. Sharp, Carol A. Gross, and Kenneth N. Kreuzer. "Substitutions in Bacteriophage T4 AsiA andEscherichia coli ς70 That Suppress T4motA Activation Mutations." Journal of Bacteriology 183, no. 7 (April 1, 2001): 2289–97. http://dx.doi.org/10.1128/jb.183.7.2289-2297.2001.

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ABSTRACT Bacteriophage T4 middle-mode transcription requires two phage-encoded proteins, the MotA transcription factor and AsiA coactivator, along with Escherichia coli RNA polymerase holoenzyme containing the ς70 subunit. AmotA positive control (pc) mutant, motA-pc1, was used to select for suppressor mutations that alter other proteins in the transcription complex. Separate genetic selections isolated two AsiA mutants (S22F and Q51E) and five ς70 mutants (Y571C, Y571H, D570N, L595P, and S604P). All seven suppressor mutants gave partial suppressor phenotypes in vivo as judged by plaque morphology and burst size measurements. The S22F mutant AsiA protein and glutathione S-transferase fusions of the five mutant ς70 proteins were purified. All of these mutant proteins allowed normal levels of in vitro transcription when tested with wild-type MotA protein, but they failed to suppress the mutant MotA-pc1 protein in the same assay. The ς70 substitutions affected the 4.2 region, which binds the −35 sequence of E. coli promoters. In the presence of E. coli RNA polymerase without T4 proteins, the L595P and S604P substitutions greatly decreased transcription from standard E. colipromoters. This defect could not be explained solely by a disruption in −35 recognition since similar results were obtained with extended −10 promoters. The generalized transcriptional defect of these two mutants correlated with a defect in binding to core RNA polymerase, as judged by immunoprecipitation analysis. The L595P mutant, which was the most defective for in vitro transcription, failed to support E. coli growth.

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Lang, Andrew S., and J. Thomas Beatty. "A Bacterial Signal Transduction System Controls Genetic Exchange and Motility." Journal of Bacteriology 184, no. 4 (February 15, 2002): 913–18. http://dx.doi.org/10.1128/jb.184.4.913-918.2002.

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ABSTRACT The bacterium Rhodobacter capsulatus is capable of an unusual form of genetic exchange, mediated by a transducing bacteriophage-like particle called the gene transfer agent (GTA). GTA production by R. capsulatus is controlled at the level of transcription by a cellular two-component signal transduction system that includes a putative histidine kinase (CckA) and response regulator (CtrA). We found that, in addition to regulating genetic exchange by R. capsulatus, this signal transduction system controls motility. As with the regulation of GTA production, the control of motility by CckA and CtrA occurs through modulation of gene transcription. Disruptions of the cckA and ctrA genes resulted in a loss of class II, class III, and class IV flagellar gene transcripts, suggesting that cckA and ctrA function in motility as class I flagellar genes. We also found that, analogous to the GTA genes, transcription of R. capsulatus flagellar genes appears to be growth phase dependent: class II flagellar gene transcripts are maximal in the mid-log phase of the culture growth cycle, whereas class III gene transcripts are maximal in the late-log phase of growth. We speculate that coordinate regulation of motility and GTA-mediated genetic exchange in R. capsulatus exists because these two processes are complementary mechanisms for cells to cope with unfavorable conditions in natural environments.

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Szalewska-Pałasz, A., and G. Wegrzyn. "Inhibition of transcription starting from bacteriophage lambda pR promoter during the stringent response in Escherichia coli: implications for lambda DNA replication." Acta Biochimica Polonica 42, no. 2 (June 30, 1995): 233–39. http://dx.doi.org/10.18388/abp.1995_4615.

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Replication of lambda plasmid DNA is halted in amino acid-starved wild type (stringent) strains whereas it proceeds in relA (relaxed) mutants. The only transcription which could be important in lambda plasmid DNA replication in amino acid-starved Escherichia coli cells is that starting from the pR promoter. Using a fusion which consists of the lacZ gene under the control of bacteriophage lambda pR promoter we found that transcription starting from this promoter was inhibited during the stringent, but not the relaxed, response in E. coli. We confirmed our conclusion by estimating the relative level of the pR transcript by RNA-DNA hybridization. We propose that decreased transcription from the pR promoter which serves as transcriptional activation of ori lambda is responsible for inhibition of lambda plasmid replication during the stringent response. The results presented in this paper, combined with our recent findings (published elsewhere), indicate that the transcriptional activation of ori lambda may be a main regulatory process controlling lambda DNA replication not only during the relaxed response but also in normal growth conditions.

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Winkelman, J. W., G. A. Kassavetis, and E. P. Geiduschek. "Molecular genetic analysis of a prokaryotic transcriptional coactivator: functional domains of the bacteriophage T4 gene 33 protein." Journal of Bacteriology 176, no. 4 (1994): 1164–71. http://dx.doi.org/10.1128/jb.176.4.1164-1171.1994.

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Levint, Judith R., and Michael J. Chamberlin. "Mapping and characterization of transcriptional pause sites in the early genetic region of bacteriophage T7." Journal of Molecular Biology 196, no. 1 (July 1987): 61–84. http://dx.doi.org/10.1016/0022-2836(87)90511-0.

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Carpousis, Agamemnon J., Elisabeth A. Mudd, and Henry M. Krisch. "Transcription and messenger RNA processing upstream of bacteriophage T4 gene 32." Molecular and General Genetics MGG 219, no. 1-2 (October 1989): 39–48. http://dx.doi.org/10.1007/bf00261155.

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Kuchinski, Kevin S., Cedric A. Brimacombe, Alexander B. Westbye, Hao Ding, and J. Thomas Beatty. "The SOS Response Master Regulator LexA Regulates the Gene Transfer Agent of Rhodobacter capsulatus and Represses Transcription of the Signal Transduction Protein CckA." Journal of Bacteriology 198, no. 7 (February 1, 2016): 1137–48. http://dx.doi.org/10.1128/jb.00839-15.

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ABSTRACTThe gene transfer agent ofRhodobacter capsulatus(RcGTA) is a genetic exchange element that combines central aspects of bacteriophage-mediated transduction and natural transformation. RcGTA particles resemble a small double-stranded DNA bacteriophage, package random ∼4-kb fragments of the producing cell genome, and are released from a subpopulation (<1%) of cells in a stationary-phase culture. RcGTA particles deliver this DNA to surroundingR. capsulatuscells, and the DNA is integrated into the recipient genome though a process that requires homologs of natural transformation genes and RecA-mediated homologous recombination. Here, we report the identification of the LexA repressor, the master regulator of the SOS response in many bacteria, as a regulator of RcGTA activity. Deletion of thelexAgene resulted in the abolition of detectable RcGTA production and an ∼10-fold reduction in recipient capability. A search for SOS box sequences in theR. capsulatusgenome sequence identified a number of putative binding sites located 5′ of typical SOS response coding sequences and also 5′ of the RcGTA regulatory genecckA, which encodes a hybrid histidine kinase homolog. Expression ofcckAwas increased >5-fold in thelexAmutant, and alexA cckAdouble mutant was found to have the same phenotype as a ΔcckAsingle mutant in terms of RcGTA production. The data indicate that LexA is required for RcGTA production and maximal recipient capability and that the RcGTA-deficient phenotype of thelexAmutant is largely due to the overexpression ofcckA.IMPORTANCEThis work describes an unusual phenotype of alexAmutant of the alphaproteobacteriumRhodobacter capsulatusin respect to the phage transduction-like genetic exchange carried out by theR. capsulatusgene transfer agent (RcGTA). Instead of the expected SOS response characteristic of prophage induction, thislexAmutation not only abolishes the production of RcGTA particles but also impairs the ability of cells to receive RcGTA-borne genes. The data show that, despite an apparent evolutionary relationship to lambdoid phages, the regulation of RcGTA gene expression differs radically.

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Marsh, Jane W., and Ronald K. Taylor. "Genetic and Transcriptional Analyses of theVibrio cholerae Mannose-Sensitive Hemagglutinin Type 4 Pilus Gene Locus." Journal of Bacteriology 181, no. 4 (February 15, 1999): 1110–17. http://dx.doi.org/10.1128/jb.181.4.1110-1117.1999.

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ABSTRACT The mannose-sensitive hemagglutinin (MSHA) of the Vibrio cholerae O1 El Tor biotype is a member of the family of type 4 pili. Type 4 pili are found on the surface of a variety of gram-negative bacteria and have demonstrated importance as host colonization factors, bacteriophage receptors, and mediators of DNA transfer. The gene locus required for the assembly and secretion of the MSHA pilus has been localized to a 16.7-kb region of the V. cholerae chromosome. Sixteen genes required for hemagglutination, including five that encode prepilin or prepilin-like proteins, have been identified. Examination of MSHA-specific cDNAs has localized two promoters that drive expression of these genes. This evidence indicates that the MSHA gene locus is transcriptionally organized into two operons, one encoding the secretory components and the other encoding the structural subunits, an arrangement unique among previously characterized type 4 pilus loci. The genes flanking the MSHA locus encode proteins that show homology to YhdA and MreB ofEscherichia coli. In E. coli, theyhdA and mreB genes are adjacent to each other on the chromosome. The finding that the MSHA locus lies between these two E. coli homologs and that it is flanked by a 7-bp direct repeat suggests that the MSHA locus may have been acquired as a mobile genetic element.

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Swarts, Daan C. "Making the cut(s): how Cas12a cleaves target and non-target DNA." Biochemical Society Transactions 47, no. 5 (October 11, 2019): 1499–510. http://dx.doi.org/10.1042/bst20190564.

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Abstract CRISPR–Cas12a (previously named Cpf1) is a prokaryotic deoxyribonuclease that can be programmed with an RNA guide to target complementary DNA sequences. Upon binding of the target DNA, Cas12a induces a nick in each of the target DNA strands, yielding a double-stranded DNA break. In addition to inducing cis-cleavage of the targeted DNA, target DNA binding induces trans-cleavage of non-target DNA. As such, Cas12a–RNA guide complexes can provide sequence-specific immunity against invading nucleic acids such as bacteriophages and plasmids. Akin to CRISPR–Cas9, Cas12a has been repurposed as a genetic tool for programmable genome editing and transcriptional control in both prokaryotic and eukaryotic cells. In addition, its trans-cleavage activity has been applied for high-sensitivity nucleic acid detection. Despite the demonstrated value of Cas12a for these applications, the exact molecular mechanisms of both cis- and trans-cleavage of DNA were not completely understood. Recent studies have revealed mechanistic details of Cas12a-mediates DNA cleavage: base pairing of the RNA guide and the target DNA induces major conformational changes in Cas12a. These conformational changes render Cas12a in a catalytically activated state in which it acts as deoxyribonuclease. This deoxyribonuclease activity mediates cis-cleavage of the displaced target DNA strand first, and the RNA guide-bound target DNA strand second. As Cas12a remains in the catalytically activated state after cis-cleavage, it subsequently demonstrates trans-cleavage of non-target DNA. Here, I review the mechanistic details of Cas12a-mediated cis- and trans-cleavage of DNA. In addition, I discuss how bacteriophage-derived anti-CRISPR proteins can inhibit Cas12a activity.

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Miller, Eric S., Elizabeth Kutter, Gisela Mosig, Fumio Arisaka, Takashi Kunisawa, and Wolfgang Rüger. "Bacteriophage T4 Genome." Microbiology and Molecular Biology Reviews 67, no. 1 (March 2003): 86–156. http://dx.doi.org/10.1128/mmbr.67.1.86-156.2003.

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SUMMARY Phage T4 has provided countless contributions to the paradigms of genetics and biochemistry. Its complete genome sequence of 168,903 bp encodes about 300 gene products. T4 biology and its genomic sequence provide the best-understood model for modern functional genomics and proteomics. Variations on gene expression, including overlapping genes, internal translation initiation, spliced genes, translational bypassing, and RNA processing, alert us to the caveats of purely computational methods. The T4 transcriptional pattern reflects its dependence on the host RNA polymerase and the use of phage-encoded proteins that sequentially modify RNA polymerase; transcriptional activator proteins, a phage sigma factor, anti-sigma, and sigma decoy proteins also act to specify early, middle, and late promoter recognition. Posttranscriptional controls by T4 provide excellent systems for the study of RNA-dependent processes, particularly at the structural level. The redundancy of DNA replication and recombination systems of T4 reveals how phage and other genomes are stably replicated and repaired in different environments, providing insight into genome evolution and adaptations to new hosts and growth environments. Moreover, genomic sequence analysis has provided new insights into tail fiber variation, lysis, gene duplications, and membrane localization of proteins, while high-resolution structural determination of the “cell-puncturing device,” combined with the three-dimensional image reconstruction of the baseplate, has revealed the mechanism of penetration during infection. Despite these advances, nearly 130 potential T4 genes remain uncharacterized. Current phage-sequencing initiatives are now revealing the similarities and differences among members of the T4 family, including those that infect bacteria other than Escherichia coli. T4 functional genomics will aid in the interpretation of these newly sequenced T4-related genomes and in broadening our understanding of the complex evolution and ecology of phages—the most abundant and among the most ancient biological entities on Earth.

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Luo, Yun, and John D. Helmann. "Extracytoplasmic Function σ Factors with Overlapping Promoter Specificity Regulate Sublancin Production in Bacillus subtilis." Journal of Bacteriology 191, no. 15 (May 22, 2009): 4951–58. http://dx.doi.org/10.1128/jb.00549-09.

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ABSTRACT Bacillus subtilis harbors seven extracytoplasmic function (ECF) σ factors. At least three ECF σ factors (σM, σW, and σX) are induced by, and provide resistance to, antibiotics and other agents eliciting cell envelope stress. Here, we report that ECF σ factors also contribute to antibiotic production. B. subtilis 168 strains that are lysogenic for the SPβ bacteriophage produce sublancin, which inhibits the growth of other, nonlysogenic strains. Genetic studies demonstrate that synthesis of sublancin is largely dependent on σX, with a smaller contribution from σM. A sigM sigX double mutant is unable to produce sublancin. This defect is primarily due to the fact that the sublancin biosynthesis is positively activated by the transition state regulator and AbrB paralog Abh, which counteracts transcriptional repression of the sublancin biosynthesis operon by AbrB. Ectopic expression of abh bypasses the requirement for σM or σX in sublancin synthesis, as does an abrB mutation. In addition to their major role in regulating sublancin expression by activating abh transcription, σX and σM also have a second role as positive regulators of sublancin expression that is independent of AbrB and Abh. Since sublancin resistance in nonlysogens is largely dependent on σW, ECF σ factors control both sublancin production and resistance.

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Kumaraswami, Muthiah, Lakshmi Avanigadda, Rajendra Rai, Hee-Won Park, and Martha M. Howe. "Genetic Analysis of Phage Mu Mor Protein Amino Acids Involved in DNA Minor Groove Binding and Conformational Changes." Journal of Biological Chemistry 286, no. 41 (August 22, 2011): 35852–62. http://dx.doi.org/10.1074/jbc.m111.269860.

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Gene expression during lytic development of bacteriophage Mu occurs in three phases: early, middle, and late. Transcription from the middle promoter, Pm, requires the phage-encoded activator protein Mor and the bacterial RNA polymerase. The middle promoter has a −10 hexamer, but no −35 hexamer. Instead Pm has a hyphenated inverted repeat that serves as the Mor binding site overlapping the position of the missing −35 element. Mor binds to this site as a dimer and activates transcription by recruiting RNA polymerase. The crystal structure of the His-Mor dimer revealed three structural elements: an N-terminal dimerization domain, a C-terminal helix-turn-helix DNA-binding domain, and a β-strand linker between the two domains. We predicted that the highly conserved residues in and flanking the β-strand would be essential for the conformational flexibility and DNA minor groove binding by Mor. To test this hypothesis, we carried out single codon-specific mutagenesis with degenerate oligonucleotides. The amino acid substitutions were identified by DNA sequencing. The mutant proteins were characterized for their overexpression, solubility, DNA binding, and transcription activation. This analysis revealed that the Gly-Gly motif formed by Gly-65 and Gly-66 and the β-strand side chain of Tyr-70 are crucial for DNA binding by His-tagged Mor. Mutant proteins with substitutions at Gly-74 retained partial activity. Treatment with the minor groove- and GC-specific chemical chromomycin A3 demonstrated that chromomycin prevented His-Mor binding but could not disrupt a pre-formed His-Mor·DNA complex, consistent with the prediction that Mor interacts with the minor groove of the GC-rich spacer in the Mor binding site.

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Madsen, Peter Lynge, Annette H. Johansen, Karin Hammer, and Lone Brøndsted. "The Genetic Switch Regulating Activity of Early Promoters of the Temperate Lactococcal Bacteriophage TP901-1." Journal of Bacteriology 181, no. 24 (December 15, 1999): 7430–38. http://dx.doi.org/10.1128/jb.181.24.7430-7438.1999.

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ABSTRACT A functional analysis of open reading frame 4 (ORF4) and ORF5 from the temperate lactococcal phage TP901-1 was performed by mutant and deletion analysis combined with transcriptional studies of the early phage promoters p R andp L. ORF4 (180 amino acids) was identified as a phage repressor necessary for repression of both promoters. Furthermore, the presence of ORF4 confers immunity of the host strain to TP901-1. ORF5 (72 amino acids) was found to be able to inhibit repression of the lytic promoter p L by ORF4. Upon transformation with a plasmid containing both ORF4 and ORF5 and their cognate promoters, clonal variation is observed: in each transformant, either p L is open andp R is closed or vice versa. The repression is still dependent on ORF4, and the presence of ORF5 is needed for the clonal variation. Induction of a repressed p Lfusion containing orf4 and orf5 was obtained by addition of mitomycin C, and the induction was also shown to be dependent on the presence of the RecA protein, even though ORF4 does not contain a recognizable autocleavage site. Our results suggest that the relative amounts of the two proteins ORF4 and ORF5 determine the decision between lytic or lysogenic life cycle after phage infection and that a protein complex consisting of ORF4 and ORF5 may constitute a new type of genetic switch in bacteriophages.

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Gansz, Andreas, Ulrich Kruse, and Wolfgang Rüger. "Gene product dsbA of bacteriophage T4 binds to late promoters and enhances late transcription." Molecular and General Genetics MGG 225, no. 3 (March 1991): 427–34. http://dx.doi.org/10.1007/bf00261683.

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GREENBLATT, J., T. F. MAH, P. LEGAULT, J. MOGRIDGE, J. LI, and L. E. KAY. "Structure and Mechanism in Transcriptional Antitermination by the Bacteriophage N Protein." Cold Spring Harbor Symposia on Quantitative Biology 63 (January 1, 1998): 327–36. http://dx.doi.org/10.1101/sqb.1998.63.327.

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25

Sullivan, Christopher S., and James M. Pipas. "T Antigens of Simian Virus 40: Molecular Chaperones for Viral Replication and Tumorigenesis." Microbiology and Molecular Biology Reviews 66, no. 2 (June 2002): 179–202. http://dx.doi.org/10.1128/mmbr.66.2.179-202.2002.

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SUMMARY Simian virus 40 (SV40) is a small DNA tumor virus that has been extensively characterized due to its relatively simple genetic organization and the ease with which its genome is manipulated. The large and small tumor antigens (T antigens) are the major regulatory proteins encoded by SV40. Large T antigen is responsible for both viral and cellular transcriptional regulation, virion assembly, viral DNA replication, and alteration of the cell cycle. Deciphering how a single protein can perform such numerous and diverse functions has remained elusive. Recently it was established that the SV40 T antigens, including large T antigen, are molecular chaperones, each with a functioning DnaJ domain. The molecular chaperones were originally identified as bacterial genes essential for bacteriophage growth and have since been shown to be conserved in eukaryotes, participating in an array of both viral and cellular processes. This review discusses the mechanisms of DnaJ/Hsc70 interactions and how they are used by T antigen to control viral replication and tumorigenesis. The use of the DnaJ/Hsc70 system by SV40 and other viruses suggests an important role for these molecular chaperones in the regulation of the mammalian cell cycle and sheds light on the enigmatic SV40 T antigen—a most amazing molecule.

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Wang, R., E. Halper-Stromberg, M. Szymanski-Pierce, S. S. Bassett, and D. Avramopoulos. "Genetic determinants of neuroglobin transcription." neurogenetics 15, no. 1 (December 24, 2013): 65–75. http://dx.doi.org/10.1007/s10048-013-0388-3.

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Kashlev, Mikhail, Evgeny Nudler, Alex Goldfarb, Terry White, and Elizabeth Kutter. "Bacteriophage T4 Alc protein: A transcription termination factor sensing local modification of DNA." Cell 75, no. 1 (October 1993): 147–54. http://dx.doi.org/10.1016/s0092-8674(05)80091-1.

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Cech, Grzegorz M., Anna Kloska, Klaudyna Krause, Katarzyna Potrykus, Michael Cashel, and Agnieszka Szalewska-Pałasz. "Virus–Host Interaction Gets Curiouser and Curiouser. PART I: Phage P1vir Enhanced Development in an E. coli DksA-Deficient Cell." International Journal of Molecular Sciences 22, no. 11 (May 31, 2021): 5890. http://dx.doi.org/10.3390/ijms22115890.

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Bacteriophage P1 is among the best described bacterial viruses used in molecular biology. Here, we report that deficiency in the host cell DksA protein, an E. coli global transcription regulator, improves P1 lytic development. Using genetic and microbiological approaches, we investigated several aspects of P1vir biology in an attempt to understand the basis of this phenomenon. We found several minor improvements in phage development in the dksA mutant host, including more efficient adsorption to bacterial cell and phage DNA replication. In addition, gene expression of the main repressor of lysogeny C1, the late promoter activator Lpa, and lysozyme are downregulated in the dksA mutant. We also found nucleotide substitutions located in the phage immunity region immI, which may be responsible for permanent virulence of phage P1vir. We suggest that downregulation of C1 may lead to a less effective repression of lysogeny maintaining genes and that P1vir may be balancing between lysis and lysogeny, although finally it is able to enter the lytic pathway only. The mentioned improvements, such as more efficient replication and more “gentle” cell lysis, while considered minor individually, together may account for the phenomenon of a more efficient P1 phage development in a DksA-deficient host.

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Nakamura, Yoshikazu, Saeko Mizusawa, Akiko Tsugawa, and Mutsuo Imai. "Conditionally lethal nusAts mutation of Escherichia coli reduces transcription termination but does not affect antitermination of bacteriophage lambda." Molecular and General Genetics MGG 204, no. 1 (July 1986): 24–28. http://dx.doi.org/10.1007/bf00330182.

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Matsushita, Kenshi, Jumpei Uchiyama, Shin-ichiro Kato, Takako Ujihara, Hiroshi Hoshiba, Shigeyoshi Sugihara, Asako Muraoka, Hiroshi Wakiguchi, and Shigenobu Matsuzaki. "Morphological and genetic analysis of three bacteriophages ofSerratia marcescensisolated from environmental water." FEMS Microbiology Letters 291, no. 2 (February 2009): 201–8. http://dx.doi.org/10.1111/j.1574-6968.2008.01455.x.

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Domelier, Anne-Sophie, Nathalie van der Mee-Marquet, Pierre-Yves Sizaret, Geneviève Héry-Arnaud, Marie-Frédérique Lartigue, Laurent Mereghetti, and Roland Quentin. "Molecular Characterization and Lytic Activities of Streptococcus agalactiae Bacteriophages and Determination of Lysogenic-Strain Features." Journal of Bacteriology 191, no. 15 (May 22, 2009): 4776–85. http://dx.doi.org/10.1128/jb.00426-09.

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ABSTRACT The application of mitomycin C induction to 114 genetically diverse Streptococcus agalactiae strains generated 36 phage suspensions. On electron microscopy of the phage suspensions, it was possible to assign the phages to the Siphoviridae family, with three different morphotypes (A, B, and C). Phage genetic diversity was evaluated by a PCR-based multilocus typing method targeting key modules located in the packaging, structural, host lysis, lysogeny, replication, and transcriptional regulation clusters and in the integrase genes and by DNA digestion with EcoRI, HindIII, and ClaI. Thirty-three phages clustering in six distantly related molecular phage groups (I to VI) were identified. Each molecular group was morphotype specific except for morphotype A phages, which were found in five of the six phage groups. The various phage groups defined on the basis of molecular group and morphotype had specific lytic activities, suggesting that each recognized particular host cell targets and had particular lytic mechanisms. Comparison of the characteristics of lysogenic and propagating strains showed no difference in the serotype or clonal complex (CC) identified by multilocus sequence typing. However, all the lysogenic CC17 and CC19 strains presented catabolic losses due to a lack of catabolic decay of dl-alpha-glycerol-phosphate substrates (CC17) and of alpha-d-glucose-1-phosphate (CC19). Moreover, the phages from CC17 lysogenic strains displayed lytic replication in bacterial hosts from all S. agalactiae phylogenetic lineages other than CC23, whereas phages obtained from non-CC17 lysogenic strains lysed bacteria of similar evolutionary origin. Our findings suggest that the adaptive evolution of S. agalactiae exposed the bacteria of this species to various phage-mediated horizontal gene transfers, which may have affected the fitness of the more virulent clones.

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Mondal, Avisek, Rajagopal Chattopadhyaya, Ajit Bikram Datta, and Pradeep Parrack. "Crystallization and X-ray analysis of the transcription-activator protein C1 of bacteriophage P22 in complex with the PREpromoter element." Acta Crystallographica Section F Structural Biology Communications 71, no. 10 (September 23, 2015): 1286–91. http://dx.doi.org/10.1107/s2053230x15015708.

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The transcription-activator protein C1 of the temperate phage P22 ofSalmonella typhimuriumplays a key role in the lyticversuslysogenic switch of the phage. A homotetramer of 92-residue polypeptides, C1 binds to an approximate direct repeat similar to the transcription activator CII of coliphage λ. Despite this and several other similarities, including 57% sequence identity to coliphage CII, many biochemical observations on P22 C1 cannot be explained based on the structure of CII. To understand the molecular basis of these differences, C1 was overexpressed and purified and subjected to crystallization trials. Although no successful hits were obtained for the apoprotein, crystals could be obtained when the protein was subjected to crystallization trials in complex with a 23-mer promoter DNA fragment (PRE). These crystals diffracted very well at the home source, allowing the collection of a 2.2 Å resolution data set. The C1–DNA crystals belonged to space groupP21, with unit-cell parametersa= 87.27,b= 93.58,c= 111.16 Å, β = 94.51°. Solvent-content analysis suggests that the asymmetric unit contains three tetramer–DNA complexes. The three-dimensional structure is expected to shed light on the mechanism of activation by C1 and the molecular basis of its specificity.

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Chen, Wei, Stanley Tabor, and Kevin Struhl. "Distinguishing between mechanisms of eukaryotic transcriptional activation with bacteriophage T7 RNA polymerase." Cell 50, no. 7 (September 1987): 1047–55. http://dx.doi.org/10.1016/0092-8674(87)90171-1.

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Jeng, Shih-Tong, Sheue-Hwey Lay, and Hsi-Mei Lai. "Transcription termination by bacteriophage T3 and SP6 RNA polymerases at Rho-independent terminators." Canadian Journal of Microbiology 43, no. 12 (December 1, 1997): 1147–56. http://dx.doi.org/10.1139/m97-163.

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Transcription termination of T3 and SP6 DNA-dependent RNA polymerases have been studied on the DNA templates containing the threonine (thr) attenuator and its variants. The thr attenuator is from the regulatory region of the thr operon of Escherichia coli. The DNA template, encoding the thr attenuator, contains specific features of the rho-independent terminators. It comprises a dG + dC rich dyad symmetry, encoding a stem-and-loop RNA, which is followed by a poly(U) region at the 3′-end. Thirteen attenuator variants have been analyzed for their ability to terminate transcription and the results indicated that the structure as well as the sequence in the G + C rich region of RNA hairpin affect termination of both RNA polymerases. Also, a single base change in the A residues of the hairpin failed to influence termination, whereas changes in the poly(U) region significantly reduced the termination of both T3 and SP6 RNA polymerases. The requirement of a poly(U) region for termination by T3 and SP6 RNA polymerases was studied with nested deletion mutants in this region. The minimum number of U residues required for termination of SP6 and T3 RNA polymerases was five and three, respectively. However, both RNA polymerases needed at least eight U residues to reach a termination efficiency close to that achieved by wild-type thr attenuator encoding nine U residues. In addition, the orientation of the loop sequences of the RNA hairpin did not affect the transcription termination of either of the bacteriophage RNA polymerases.Key words: transcription termination, bacteriophage RNA polymerase.

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García, Pilar, Juan Evaristo Suárez, Victoria Bascarán, and Ana Rodríguez. "Isolation and characterization of promoters from the Lactobacillus casei temperate bacteriophage A2." Canadian Journal of Microbiology 43, no. 11 (November 1, 1997): 1063–68. http://dx.doi.org/10.1139/m97-151.

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Random Sau3AI DNA fragments from the temperate Lactobacillus bacteriophage A2 were cloned into the promoter-probe plasmid pGKV210. Seven DNA fragments with promoter activity were selected, after transformation of Escherichia coli and Lactococcus lactis, subsp. lactis, through the chloramphenicol resistance they conferred to the corresponding clones. The seven promoters were functional in Lactobacillus casei. Their strength was analysed by measuring the levels of chloramphenicol resistance and chloramphenicol acetyltransferase activity induced in each host. The nucleotide sequences of these fragments were determined and primer extension analysis was used to locate the initiation site of transcription from each promoter in E. coli. The promoters contained −10 and −35 regions similar to the consensus sequences of E. coli and Lactobacillus promoters.Key words: bacteriophage, Lactobacillus, promoter.

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36

Tadini, Luca, Nicolaj Jeran, Carlotta Peracchio, Simona Masiero, Monica Colombo, and Paolo Pesaresi. "The plastid transcription machinery and its coordination with the expression of nuclear genome: Plastid-Encoded Polymerase, Nuclear-Encoded Polymerase and the Genomes Uncoupled 1-mediated retrograde communication." Philosophical Transactions of the Royal Society B: Biological Sciences 375, no. 1801 (May 4, 2020): 20190399. http://dx.doi.org/10.1098/rstb.2019.0399.

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Plastid genes in higher plants are transcribed by at least two different RNA polymerases, the plastid-encoded RNA polymerase (PEP), a bacteria-like core enzyme whose subunits are encoded by plastid genes ( rpoA , rpoB , rpoC1 and rpoC2 ), and the nuclear-encoded plastid RNA polymerase (NEP), a monomeric bacteriophage-type RNA polymerase. Both PEP and NEP enzymes are active in non-green plastids and in chloroplasts at all developmental stages. Their transcriptional activity is affected by endogenous and exogenous factors and requires a strict coordination within the plastid and with the nuclear gene expression machinery. This review focuses on the different molecular mechanisms underlying chloroplast transcription regulation and its coordination with the photosynthesis-associated nuclear genes ( PhANGs ) expression. Particular attention is given to the link between NEP and PEP activity and the GUN1- (Genomes Uncoupled 1) mediated chloroplast-to-nucleus retrograde communication with respect to the Δrpo adaptive response, i.e. the increased accumulation of NEP-dependent transcripts upon depletion of PEP activity, and the editing-level changes observed in NEP-dependent transcripts, including rpoB and rpoC1 , in gun1 cotyledons after norflurazon or lincomycin treatment. The role of cytosolic preproteins and HSP90 chaperone as components of the GUN1-retrograde signalling pathway, when chloroplast biogenesis is inhibited in Arabidopsis cotyledons, is also discussed. This article is part of the theme issue ‘Retrograde signalling from endosymbiotic organelles’.

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37

Pan, Meichen, Matthew A. Nethery, Claudio Hidalgo-Cantabrana, and Rodolphe Barrangou. "Comprehensive Mining and Characterization of CRISPR-Cas Systems in Bifidobacterium." Microorganisms 8, no. 5 (May 12, 2020): 720. http://dx.doi.org/10.3390/microorganisms8050720.

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The clustered regularly interspaced short palindromic repeats (CRISPR)-Cas (CRISPR-associated cas) systems constitute the adaptive immune system in prokaryotes, which provides resistance against bacteriophages and invasive genetic elements. The landscape of applications in bacteria and eukaryotes relies on a few Cas effector proteins that have been characterized in detail. However, there is a lack of comprehensive studies on naturally occurring CRISPR-Cas systems in beneficial bacteria, such as human gut commensal Bifidobacterium species. In this study, we mined 954 publicly available Bifidobacterium genomes and identified CRIPSR-Cas systems in 57% of these strains. A total of five CRISPR-Cas subtypes were identified as follows: Type I-E, I-C, I-G, II-A, and II-C. Among the subtypes, Type I-C was the most abundant (23%). We further characterized the CRISPR RNA (crRNA), tracrRNA, and PAM sequences to provide a molecular basis for the development of new genome editing tools for a variety of applications. Moreover, we investigated the evolutionary history of certain Bifidobacterium strains through visualization of acquired spacer sequences and demonstrated how these hypervariable CRISPR regions can be used as genotyping markers. This extensive characterization will enable the repurposing of endogenous CRISPR-Cas systems in Bifidobacteria for genome engineering, transcriptional regulation, genotyping, and screening of rare variants.

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COTSAPAS, C., E. CHAN, M. KIRK, M. TANAKA, and P. LITTLE. "Genetic Variation and the Control of Transcription." Cold Spring Harbor Symposia on Quantitative Biology 68 (January 1, 2003): 109–14. http://dx.doi.org/10.1101/sqb.2003.68.109.

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39

Hayes, Sidney, and Roderick A. Slavcev. "Polarity withinpMandpEpromoted phage lambdacI-rexA-rexBtranscription and its suppression." Canadian Journal of Microbiology 51, no. 1 (January 1, 2005): 37–49. http://dx.doi.org/10.1139/w04-115.

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The cI-rexA-rexB operon of bacteriophage λ confers 2 phenotypes, Imm and Rex, to lysogenic cells. Immunity to homoimmune infecting λ phage depends upon the CI repressor. Rex exclusion of T4rII mutants requires RexA and RexB proteins. Both Imm and Rex share temperature-sensitive conditional phenotypes when expressed from cI[Ts]857 but not from cI+λ prophage. Plasmids were made in which cI-rexA-rexB was transcribed from a non-lambda promoter, pTet. The cI857-rexA-rexB plasmid exhibited Ts conditional Rex and CI phenotypes; the cI+-rexA-rexB plasmid did not. Polarity was observed within cI-rexA-rexB transcription at sites in cI and rexA when CI was nonfunctional. Renaturation of the Ts CI857 repressor, allowing it to regain functionality, suppressed the polar effect on downstream transcription from the site in cI. The second strong polar effect near the distal end of rexA was observed for transcription initiated from pE. The introduction of a rho Ts mutation into the host genome suppressed both polar effects, as measured by its suppression of the conditional Rex phenotype. Strong suppression of the conditional Rex[Ts] phenotype was imparted by ssrA and clpP (polar for clpX) null mutations, suggesting that RexA or RexB proteins made under conditions of polarity are subject to 10Sa RNA tagging and ClpXP degradation.Key words: bacteriophage lambda (λ), Rex exclusion, CI and Cro repressors, Rho, ClpXP, 10Sa RNA tagging.

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Gutiérrez, Diana, Antonio M. Martín-Platero, Ana Rodríguez, Manuel Martínez-Bueno, Pilar García, and Beatriz Martínez. "Typing of bacteriophages by randomly amplified polymorphic DNA (RAPD)-PCR to assess genetic diversity." FEMS Microbiology Letters 322, no. 1 (July 13, 2011): 90–97. http://dx.doi.org/10.1111/j.1574-6968.2011.02342.x.

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41

Szalewska-PaÅasz, Agnieszka, Alicja WÃ§grzyn, MichaÅ Obuchowski, Ryszard PawÅowski, Krzysztof Bielawski, Mark S. Thomas, and Grzegorz WeÄgrzyn. "Drastically decreased transcription from CII-activated promoters is responsible for impaired lysogenization of theEscherichia coli rpoA341mutant by bacteriophage Î»." FEMS Microbiology Letters 144, no. 1 (October 1996): 21–27. http://dx.doi.org/10.1111/j.1574-6968.1996.tb08503.x.

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WOLFFE, A. "A bacteriophage RNA polymerase transcribes through a Xenopus 5S RNA gene transcription complex without disrupting it." Cell 44, no. 3 (February 1986): 381–89. http://dx.doi.org/10.1016/0092-8674(86)90459-9.

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43

Attwood, Graeme T., Felicitas Herrera, Lee A. Weissenstein, and Bryan A. White. "An endo-β-1,4-glucanase gene (celA) from the rumen anaerobe Ruminococcus albus 8: cloning, sequencing, and transcriptional analysis." Canadian Journal of Microbiology 42, no. 3 (March 1, 1996): 267–78. http://dx.doi.org/10.1139/m96-039.

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A genomic library of Ruminococcus albus 8 DNA was constructed in Escherichia coli using bacteriophage λZapII. This library was screened for cellulase components and several Ostazin brilliant red/carboxymethyl cellulose positive clones were isolated. All of these clones contained a common 3.4-kb insert, which was recovered as a plasmid by helper phage excision. The carboxymethyl cellulase coding region was localized to a 1.4-kb region of DNA by nested deletions, and a clone containing the entire celA gene was sequenced. Analysis of the sequence revealed a 1231-bp open reading frame, coding for a protein of 411 amino acids with a predicted molecular weight of 45 747. This protein, designated CelA, showed extensive homology with family 5 endoglucanases by both primary amino acid sequence alignment and hydrophobic cluster analysis. Cell-free extracts of E. coli containing the celA clone demonstrated activity against carboxymethyl cellulose and acid swollen cellulose but not against any of the p-nitrophenol glycosides tested, indicating an endo-β-1,4-glucanase type of activity. In vitro transcription–translation experiments showed that three proteins of 48 000, 44 000, and 23 000 molecular weight were produced by clones containing the celA gene. Northern analysis of RNA extracted from R. albus 8 grown on cellulose indicated a celA transcript of approximately 2700 bases, whereas when R. albus 8 was grown on cellobiose, celA transcripts of approximately 3000 and 600 bases were detected. Primer extension analysis of these RNAs revealed different transcription initiation sites for the celA gene when cells were grown with cellulose or cellobiose as the carbon source. These two sites differed by 370 bases in distance. A model, based on transcription and sequence data, is proposed for celA regulation.Key words: Ruminococcus albus, endoglucanase, transcription, hydrophobic cluster analysis.

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Viswanathan, C., and Jian-Kang Zhu. "Molecular genetic analysis of cold–regulated gene transcription." Philosophical Transactions of the Royal Society of London. Series B: Biological Sciences 357, no. 1423 (July 29, 2002): 877–86. http://dx.doi.org/10.1098/rstb.2002.1076.

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Chilling and freezing temperatures adversely affect the productivity and quality of crops. Hence improving the cold hardiness of crop plants is an important goal in agriculture, which demands a clear understanding of cold stress signal perception and transduction. Pharmacological and biochemical evidence shows that membrane rigidification followed by cytoskeleton rearrangement, Ca 2+ influx and Ca 2+ –dependent phosphorylation are involved in cold stress signal transduction. Cold–responsive genes are regulated through C–repeat/dehydration–responsive elements (CRT/DRE) and abscisic acid (ABA)–responsive element cis elements by transacting factors C–repeat binding factors/dehydration–responsive element binding proteins (CBFs/DREBs) and basic leucine zippers (bZIPs) (SGBF1), respectively. We have carried out a forward genetic analysis using chemically mutagenized Arabidopsis plants expressing cold–responsive RD29A promoter–driven luciferase to dissect cold signal transduction. We have isolated the fiery1 ( fry1 ) mutant and cloned the FRY1 gene, which encodes an inositol polyphosphate 1–phosphatase. The fry1 plants showed enhanced induction of stress genes in response to cold, ABA, salt and dehydration due to higher accumulation of the second messenger, inositol (1,4,5)– triphosphate (IP 3 ). Thus our study provides genetic evidence suggesting that cold signal is transduced through changes in IP 3 levels. We have also identified the hos1 mutation, which showed super induction of cold–responsive genes and their transcriptional activators. Molecular cloning and characterization revealed that HOS1 encodes a ring finger protein, which has been implicated as an E3 ubiquitin conjugating enzyme. HOS1 is present in the cytoplasm at normal growth temperatures but accumulates in the nucleus upon cold stress. HOS1 appears to regulate temperature sensing by the cell as cold–responsive gene expression occurs in the hos1 mutant at relatively warm temperatures. Thus HOS1 is a negative regulator, which may be functionally linked to cellular thermosensors to modulate cold–responsive gene transcription.

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Horwitz, Robert J., Joyce Li, and Jack Greenblatt. "An elongation control particle containing the N gene transcriptional antitermination protein of bacteriophage lambda." Cell 51, no. 4 (November 1987): 631–41. http://dx.doi.org/10.1016/0092-8674(87)90132-2.

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46

Zaworski, Julie, Oyut Dagva, Julius Brandt, Chloé Baum, Laurence Ettwiller, Alexey Fomenkov, and Elisabeth A. Raleigh. "Reassembling a cannon in the DNA defense arsenal: Genetics of StySA, a BREX phage exclusion system in Salmonella lab strains." PLOS Genetics 18, no. 4 (April 4, 2022): e1009943. http://dx.doi.org/10.1371/journal.pgen.1009943.

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Understanding mechanisms that shape horizontal exchange in prokaryotes is a key problem in biology. A major limit on DNA entry is imposed by restriction-modification (RM) processes that depend on the pattern of DNA modification at host-specified sites. In classical RM, endonucleolytic DNA cleavage follows detection of unprotected sites on entering DNA. Recent investigation has uncovered BREX (BacteRiophage EXclusion) systems. These RM-like activities employ host protection by DNA modification, but immediate replication arrest occurs without evident of nuclease action on unmodified phage DNA. Here we show that the historical stySA RM locus of Salmonella enterica sv Typhimurium is a variant BREX system. A laboratory strain disabled for both the restriction and methylation activity of StySA nevertheless has wild type sequence in pglX, the modification gene homolog. Instead, flanking genes pglZ and brxC each carry multiple mutations (μ) in their C-terminal domains. We further investigate this system in situ, replacing the mutated pglZμ and brxCμ genes with the WT counterpart. PglZ-WT supports methylation in the presence of either BrxCμ or BrxC-WT but not in the presence of a deletion/insertion allele, ΔbrxC::cat. Restriction requires both BrxC-WT and PglZ-WT, implicating the BrxC C-terminus specifically in restriction activity. These results suggests that while BrxC, PglZ and PglX are principal components of the BREX modification activity, BrxL is required for restriction only. Furthermore, we show that a partial disruption of brxL disrupts transcription globally.

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Kotani, Hidehito, JoAnn M. Sekiguchi, Seema Dutta, and Eric B. Kmiec. "Genetic recombination of nucleosomal templates is mediated by transcription." Molecular and General Genetics MGG 244, no. 4 (July 1994): 410–19. http://dx.doi.org/10.1007/bf00286693.

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Zhao, Luyi, Amber D. Stancik, and Celeste J. Brown. "Differential Transcription of Bacteriophage φX174 Genes at 37°C and 42°C." PLoS ONE 7, no. 4 (April 23, 2012): e35909. http://dx.doi.org/10.1371/journal.pone.0035909.

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Gabig, M., M. Obuchowski, A. Ciesielska, B. Latała, A. Wegrzyn, M. S. Thomas, and G. Wegrzyn. "The Escherichia coli RNA polymerase alpha subunit and transcriptional activation by bacteriophage lambda CII protein." Acta Biochimica Polonica 45, no. 1 (March 31, 1998): 271–80. http://dx.doi.org/10.18388/abp.1998_4342.

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Bacteriophage lambda is not able to lysogenise the Escherichia coli rpoA341 mutant. This mutation causes a single amino acid substitution Lys271Glu in the C-terminal domain of the RNA polymerase alpha subunit (alphaCTD). Our previous studies indicated that the impaired lysogenisation of the rpoA341 host is due to a defect in transcriptional activation by the phage CII protein and suggested a role for alphaCTD in this process. Here we used a series of truncation and point mutants in the rpoA gene placed on a plasmid to investigate the process of transcriptional activation by the cII gene product. Our results indicate that amino-acid residues 265, 268 and 271 in the a subunit may play an important role in the CII-mediated activation of the pE promoter (most probably residue 271) or may be involved in putative interactions between alphaCTD and an UP-like element near pE (most probably residues 265 and 268). Measurement of the activity of pE-lacZ, pI-lacZ and p(aQ)-lacZ fusions in the rpoA+ and rpoA341 hosts demonstrated that the mechanism of activation of these CII-dependent promoters may be in each case different.

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Kereszt, Attila, Ernő Kiss, Bradley L. Reuhs, Russell W. Carlson, Ádám Kondorosi, and Péter Putnoky. "Novel rkp Gene Clusters ofSinorhizobium meliloti Involved in Capsular Polysaccharide Production and Invasion of the Symbiotic Nodule: the rkpKGene Encodes a UDP-Glucose Dehydrogenase." Journal of Bacteriology 180, no. 20 (October 15, 1998): 5426–31. http://dx.doi.org/10.1128/jb.180.20.5426-5431.1998.

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ABSTRACT The production of exopolysaccharide (EPS) was shown to be required for the infection process by rhizobia that induce the formation of indeterminate nodules on the roots of leguminous host plants. InSinorhizobium meliloti (also known as Rhizobium meliloti) Rm41, a capsular polysaccharide (KPS) analogous to the group II K antigens of Escherichia coli can replace EPS during symbiotic nodule development and serve as an attachment site for the strain-specific bacteriophage φ16-3. The rkpA to -J genes in the chromosomal rkp-1 region code for proteins that are involved in the synthesis, modification, and transfer of an as-yet-unknown lipophilic molecule which might function as a specific lipid carrier during KPS biosynthesis. Here we report that with a phage φ16-3-resistant population obtained after random Tn5 mutagenesis, we have identified novel mutants impaired in KPS production by genetic complementation and biochemical studies. The mutations represent two novel loci, designated therkp-2 and rkp-3 regions, which are required for the synthesis of rhizobial KPS. The rkp-2 region harbors two open reading frames (ORFs) organized in monocistronic transcription units. Although both genes are required for normal lipopolysaccharide production, only the second one, designated rkpK, is involved in the synthesis of KPS. We have demonstrated that RkpK possesses UDP-glucose dehydrogenase activity, while the protein product of ORF1 might function as a UDP-glucuronic acid epimerase.

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Journal articles on the topic 'Bacteriophages – Molecular genetics; Genetic transcription'

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