A Novel, Highly Related Jumbo Family of Bacteriophages That Were Isolated Against Erwinia

Erwinia amylovora is a plant pathogen from the Erwiniaceae family and a causative agent of the devastating agricultural disease fire blight. Here we characterize eight lytic bacteriophages of E. amylovora that we isolated from the Wasatch front (Utah, United States) that are highly similar to vB_EamM_Ea35-70 which was isolated in Ontario, Canada. With the genome size ranging from 271 to 275 kb, this is a novel jumbo family of bacteriophages. These jumbo bacteriophages were further characterized through genomic and proteomic comparison, mass spectrometry, host range and burst size. Their proteomes are highly unstudied, with over 200 putative proteins with no known homologs. The production of 27 of these putative proteins was confirmed by mass spectrometry analysis. These bacteriophages appear to be most similar to bacteriophages that infect Pseudomonas and Ralstonia rather than Enterobacteriales bacteria by protein similarity, however, we were only able to detect infection of Erwinia and the closely related strains of Pantoea.United States Dairy Association; University of Arizona; Brigham Young University; Department of Microbiology and Molecular Biology; College of Life SciencesOpen access journalThis item from the UA Faculty Publications collection is made available by the University of Arizona with support from the University of Arizona Libraries. If you have questions, please contact us at [email protected]

Phage cluster relationships identified through single gene analysis

Abstract Background Phylogenetic comparison of bacteriophages requires whole genome approaches such as dotplot analysis, genome pairwise maps, and gene content analysis. Currently mycobacteriophages, a highly studied phage group, are categorized into related clusters based on the comparative analysis of whole genome sequences. With the recent explosion of phage isolation, a simple method for phage cluster prediction would facilitate analysis of crude or complex samples without whole genome isolation and sequencing. The hypothesis of this study was that mycobacteriophage-cluster prediction is possible using comparison of a single, ubiquitous, semi-conserved gene. Tape Measure Protein (TMP) was selected to test the hypothesis because it is typically the longest gene in mycobacteriophage genomes and because regions within the TMP gene are conserved. Results A single gene, TMP, identified the known Mycobacteriophage clusters and subclusters using a Gepard dotplot comparison or a phylogenetic tree constructed from global alignment and maximum likelihood comparisons. Gepard analysis of 247 mycobacteriophage TMP sequences appropriately recovered 98.8% of the subcluster assignments that were made by whole-genome comparison. Subcluster-specific primers within TMP allow for PCR determination of the mycobacteriophage subcluster from DNA samples. Using the single-gene comparison approach for siphovirus coliphages, phage groupings by TMP comparison reflected relationships observed in a whole genome dotplot comparison and confirm the potential utility of this approach to another widely studied group of phages. Conclusions TMP sequence comparison and PCR results support the hypothesis that a single gene can be used for distinguishing phage cluster and subcluster assignments. TMP single-gene analysis can quickly and accurately aid in mycobacteriophage classification

Snf1 cooperates with the CWI MAPK pathway to mediate the degradation of Med13 following oxidative stress

Eukaryotic cells, when faced with unfavorable environmental conditions, mount either pro-survival or pro-death programs. The conserved cyclin C-Cdk8 kinase plays a key role in this decision. Both are members of the Cdk8 kinase module that, along with Med12 and Med13, associate with the core Mediator complex of RNA polymerase II. In Saccharomyces cerevisiae, oxidative stress triggers Med13 destruction, which releases cyclin C into the cytoplasm to promote mitochondrial fission and programmed cell death. The SCFGrr1 ubiquitin ligase mediates Med13 degradation dependent on the cell wall integrity pathway, MAPK Slt2. Here we show that the AMP kinase Snf1 activates a second SCFGrr1 responsive degron in Med13. Deletion of Snf1 resulted in nuclear retention of cyclin C and failure to induce mitochondrial fragmentation. This degron was able to confer oxidative-stress-induced destruction when fused to a heterologous protein in a Snf1 dependent manner. Although snf1∆ mutants failed to destroy Med13, deleting the degron did not prevent destruction. These results indicate that the control of Med13 degradation following H2O2 stress is complex, being controlled simultaneously by CWI and MAPK pathways

Complete Genome Sequences of Paenibacillus Larvae Phages BN12, Dragolir, Kiel007, Leyra, Likha, Pagassa, PBL1c, and Tadhana

We present here the complete genomes of eight phages that infect Paenibacillus larvae, the causative agent of American foulbrood in honeybees. Phage PBL1c was originally isolated in 1984 from a P. larvae lysogen, while the remaining phages were isolated in 2014 from bee debris, honeycomb, and lysogens from three states in the USA

The Role of PAS Kinase in PASsing the Glucose Signal

PAS kinase is an evolutionarily conserved nutrient responsive protein kinase that regulates glucose homeostasis. Mammalian PAS kinase is activated by glucose in pancreatic beta cells, and knockout mice are protected from obesity, liver triglyceride accumulation, and insulin resistance when fed a high-fat diet. Yeast PAS kinase is regulated by both carbon source and cell integrity stress and stimulates the partitioning of glucose toward structural carbohydrate biosynthesis. In our current model for PAS kinase regulation, a small molecule metabolite binds the sensory PAS domain and activates the enzyme. Although bona fide PAS kinase substrates are scarce, in vitro substrate searches provide putative targets for exploration

Regulation of NAD Synthesis by the Trifunctional NadR Protein of Salmonella enterica

The three activities of NadR were demonstrated in purified protein and assigned to separate domains by missense mutations. The N-terminal domain represses transcription of genes for NAD synthesis and salvage. The C-terminal domain has nicotinamide ribose kinase (NmR-K; EC 2.7.1.22) activity, which is essential for assimilation of NmR, converting it internally to nicotinamide mononucleotide (NMN). The central domain has a weak adenylyltransferase (NMN-AT; EC 2.7.7.1) activity that converts NMN directly to NAD but is physiologically irrelevant. This central domain mediates regulatory effects of NAD on all NadR activities. In the absence of effectors, pure NadR protein binds operator DNA (the default state) and is released by ATP (expected to be present in vivo). NAD allows NadR to bind DNA in the presence of ATP and causes repression in vivo. A superrepressor mutation alters an ATP-binding residue in the central (NMN-AT) domain. This eliminates NMN-AT activity and places the enzyme in its default (DNA binding) state. The mutant protein shows full NmR kinase activity that is 10-fold more sensitive to NAD inhibition than the wild type. It is proposed that NAD and the superrepressor mutation exert their effects by preventing ATP from binding to the central domain