Showing papers on "Lambda phage published in 2020"
TL;DR: It is found that different viral DNAs establish separate sub cellular compartments within cells, which sustains heterogeneous viral development in single cells, suggesting that phage decision-making transcripts are spatially organized in separate compartments to allow distinct subcellular decisions to develop.
Abstract: Spatial organization of biological processes allows for variability in molecular outcomes and coordinated development. Here, we investigate how organization underpins phage lambda development and decision-making by characterizing viral components and processes in subcellular space. We use live-cell and in situ fluorescence imaging at the single-molecule level to examine lambda DNA replication, transcription, virion assembly, and resource recruitment in single-cell infections, uniting key processes of the infection cycle into a coherent model of phage development encompassing space and time. We find that different viral DNAs establish separate subcellular compartments within cells, which sustains heterogeneous viral development in single cells. These individual phage compartments are physically separated by the E. coli nucleoid. Our results provide mechanistic details describing how separate viruses develop heterogeneously to resemble single-cell phenotypes. Here, the authors apply live-cell and in situ fluorescence imaging at the single-molecule level to examine lambda DNA replication in single cells, finding that individual phage DNAs sequester host factors to their own vicinity and confine their replicated DNAs into separate compartments, suggesting that phage decision-making transcripts are spatially organized in separate compartments to allow distinct subcellular decisions to develop.
13 citations
TL;DR: In this short review article, some of the in vitro studies that have led to the current understanding of the lambda recombinase system are highlighted and insights from more than half a century of research are synthesized, summarizing the state of current understanding.
Abstract: DNA recombination, replication, and repair are intrinsically interconnected processes. From viruses to humans, they are ubiquitous and essential to all life on Earth. Single-strand annealing homologous DNA recombination is a major mechanism for the repair of double-stranded DNA breaks. An exonuclease and an annealase work in tandem, forming a complex known as a two-component recombinase. Redβ annealase and λ-exonuclease from phage lambda form the archetypal two-component recombinase complex. In this short review article, we highlight some of the in vitro studies that have led to our current understanding of the lambda recombinase system. We synthesize insights from more than half a century of research, summarizing the state of our current understanding. From this foundation, we identify the gaps in our knowledge and cast an eye forward to consider what the next 50 years of research may uncover.
8 citations
TL;DR: LysECP26 was limited in that it could not pass the outer membrane, but could be effectively controlled by adding EDTA and citric acid within 30 min, and may serve as an effective biocontrol agent for gram-negative pathogens, including E. coli O157:H7.
Abstract: With an increase in the consumption of non-heated fresh food, foodborne shiga toxin-producing Escherichia coli (STEC) has emerged as one of the most problematic pathogens worldwide. Endolysin, a bacteriophage-derived lysis protein, is able to lyse the target bacteria without any special resistance, and thus has been garnering interest as a powerful antimicrobial agent. In this study, rV5-like phage endolysin targeting E. coli O157:H7, named as LysECP26, was identified and purified. This endolysin had a lysozyme-like catalytic domain, but differed markedly from the sequence of lambda phage endolysin. LysECP26 exhibited strong activity with a broad lytic spectrum against various gram-negative strains (29/29) and was relatively stable at a broad temperature range (4°C- 55°C). The optimum temperature and pH ranges of LysECP26 were identified at 37°C-42°C and pH 7- 8, respectively. NaCl supplementation did not affect the lytic activity. Although LysECP26 was limited in that it could not pass the outer membrane, E. coli O157: H7 could be effectively controlled by adding ethylenediaminetetraacetic acid (EDTA) and citric acid (1.44 and 1.14 log CFU/ml) within 30 min. Therefore, LysECP26 may serv as an effective biocontrol agent for gram-negative pathogens, including E. coli O157:H7.
8 citations
TL;DR: It is reported that λ phage productivity in the authors' experiments is maximized around an optimal lysis time of 63 min, which is the lysisTime of the λ “wildtype” strain.
Abstract: Optimality models have a checkered history in evolutionary biology. While optimality models have been successful in providing valuable insight into the evolution of a wide variety of biological traits, a common objection is that optimality models are overly simplistic and ignore organismal genetics. We revisit evolutionary optimization in the context of a major bacteriophage life history trait, lysis time. Lysis time refers to the period spanning phage infection of a host cell and its lysis, whereupon phage progeny are released. Lysis time, therefore, directly determines phage fecundity assuming progeny assembly rate is maximized. Noting that previous tests of lysis time optimality rely on batch culture, we implemented a quasi-steady state system to observe productivity of a panel of isogenic phage lambda mutants differing in lysis time. We report that lambda phage productivity in our experiments is maximized around an optimal lysis time of 65 min, which is the lysis time of the lambda wildtype strain. We discuss this finding in light of recent results that lysis time variation is also minimized in the lambda wildtype strain.
6 citations
TL;DR: Phage YA3 is a considerably novel lambda-like temperate phage, and a further study of its genome may deepen the understanding of the interaction between lysogenic phages and their bacterial hosts.
Abstract: It is well known that bacteriophages play crucial roles in many aspects, such as controlling the number and the diversity of bacteria and participating in horizontal gene transfer, which is a key process in the evolution of bacteria. However, so far, the number of temperate bacteriophages is still limited, and their life processes are severely unknown, except for members of the lambdoid family of coliphages. In this study, a novel temperate phage of Pseudomonas aeruginosa, YA3 (vB_PaeP_YA3), was isolated from waste water. The morphology of YA3 suggested that it is a Podoviridae. The YA3 genome is a circular double-stranded DNA of 45,253 bp, with an average G + C content of 57.2%. A total of 65 open reading frames (ORFs) were predicted according to the sequence of YA3's genome, of which only 32 (49.2%) ORFs were assigned with putative functions and 13 ORFs were confirmed by the structural proteome. Genome and proteome analyses confirmed the lysogenic nature of this phage, which encodes the typical lysogen-related proteins integrase, CI, Cro, and Q protein. The genome of YA3 is most closely related with that of temperate phage vB_PaeP_Tr60_Ab31, whereas the homology coverage is just 48%. There are many critical differences between their genomes, involving promoters, lysis pathways, and regulation patterns. YA3 is capable of stably lysogenizing its host P. aeruginosa PA14, targeting the integration site within the serine tRNA gene (PA14_RS20820), which is similar with phage vB_PaeP_Tr60_Ab31. The phylogenetic analysis is more complicated than we thought. Based on phage terminase large subunit (TerL) and CI proteins, phage YA3 is related with phage lambda, while their genome coverage is extremely low (<1%). Therefore, phage YA3 is a considerably novel lambda-like temperate phage, and a further study of its genome may deepen our understanding of the interaction between lysogenic phages and their bacterial hosts.
6 citations
TL;DR: The early steps of the infection cycle of coliphage P1 are studied, focusing on their correlation with spatial locations at the single-cell level, and it is found that P1 shows no spatial preference on cell surface adsorption.
6 citations
TL;DR: Differences between molecular processes occurring during formation of replication complexes in lambda and Stx phages may indicate the specialization of the latter phages and their adaptation to specific environmental conditions where quick genetic switches are crucial.
Abstract: Shiga toxin-converting bacteriophages (or Stx phages) are responsible for virulence of enterohemorrhagic Escherichia coli strains Although they belong to the group of lambdoid phages, which have served as models in studies on DNA replication mechanisms, details of regulation of replication of Stx phage genomes are poorly understood Despite high similarity of their replication regions to that of phage lambda, considerable differences occur between them Here, we present a comparison of origins of replication and O proteins of lambda and selected Stx phages (phages P27 and 933W) Stx initiator proteins, similarly to the lambda O protein, exist in the form of dimers Only 4 iteron sequences are strongly bound in vitro by the O proteins, despite the presence of 6 such fragments in the Stx ori, while the function of the other two iterons is still crucial for transformation of E coli wild-type strain by the P27-derived lambdoid plasmid As these sequences are found in the gene coding for Stx O proteins, the sequences of these proteins themselves are also extended compared to lambda phage Therefore, proteins O of Stx phages P27 and 933W have 13 additional amino acids They can act as a space barrier, thus affecting the lesser packing of the O-some Stx complex compared to the structure found in lambda Such structure of the DNA replication initiation complex may determine its lesser dependence on the processes occurring in the host cell, including transcriptional activation of the origin Differences between molecular processes occurring during formation of replication complexes in lambda and Stx phages may indicate the specialization of the latter phages and their adaptation to specific environmental conditions where quick genetic switches are crucial
6 citations
TL;DR: The I-Block assay, which can detect sequence-specific binding of proteins to DNA in Escherichia coli, was shown to work with standard liquid cultures, with cultures grown in microplate and with colonies on X-gal indicator plates.
Abstract: We have developed a simple method called I-Block assay, which can detect sequence-specific binding of proteins to DNA in Escherichia coli. The method works by detecting competition between the protein of interest and RNA polymerase for binding to overlapping target sites in a plasmid-borne lacI promoter variant. The assay utilizes two plasmids and an E. coli host strain, from which the gene of the Lac repressor (lacI) has been deleted. One of the plasmids carries the lacI gene with a unique NheI restriction site created in the lacI promoter. The potential recognition sequences of the tested protein are inserted into the NheI site. Introduction of the plasmids into the E. coliΔlacI host represses the constitutive β-galactosidase synthesis of the host bacterium. If the studied protein expressed from a compatible plasmid binds to its target site in the lacI promoter, it will interfere with lacI transcription and lead to increased β-galactosidase activity. The method was tested with two zinc finger proteins, with the lambda phage cI857 repressor, and with CRISPR-dCas9 targeted to the lacI promoter. The I-Block assay was shown to work with standard liquid cultures, with cultures grown in microplate and with colonies on X-gal indicator plates.
2 citations
TL;DR: The antirepressor Qtip of Vibrio phage VP882, which through CI sequestration triggers a lytic switch, is characterized by Silpe et al. (2020).
2 citations