Vampirovibrio chlorellavorus draft genome sequence, annotation, and preliminary characterization of pathogenicity determinants

Vampirovibrio chlorellavorus is recognized as a pathogen of commercially-relevant Chlorella species. Algal infection and total loss of productivity (biomass) often occurs when susceptible algal hosts are cultivated in outdoor open pond systems. The pathogenic life cycle of this bacterium has been inferred from laboratory and field observations, and corroborated in part by the genomic analyses for two Arizona isolates recovered from an open algal reactor. V. chlorellavorus predation has been reported to occur in geographically- and environmentally-diverse conditions. Genomic analyses of these and additional field isolates is expected to reveal new information about the extent of ecological diversity and genes involved in host-pathogen interactions. The draft genome sequences for two isolates of the predatory V. chlorellavorus (Cyanobacteria; Ca. Melainabacteria) from an outdoor cultivation system located in the Arizona Sonoran Desert were assembled and annotated. The genomes were sequenced and analyzed to identify genes (proteins) with predicted involvement in predation, infection, and cell death of Chlorella host species prioritized for biofuel production at sites identified as highly suitable for algal production in the southwestern USA. Genomic analyses identified several predicted genes encoding secreted proteins that are potentially involved in pathogenicity, and at least three apparently complete sets of virulence (Vir) genes, characteristic of the VirB-VirD type system encoding the canonical VirB1-11 and VirD4 proteins, respectively. Additional protein functions were predicted suggesting their involvement in quorum sensing and motility. The genomes of two previously uncharacterized V. chlorellavorus isolates reveal nucleotide and protein level divergence between each other, and a previously sequenced V. chlorellavorus genome. This new knowledge will enhance the fundamental understanding of trans-kingdom interactions between a unique cosmopolitan cyanobacterial pathogen and its green microalgal host, of broad interest as a source of harvestable biomass for biofuels or bioproducts.Bioenergy Technology Office within the US Department of Energy Office of Energy Efficiency and Renewable Energy [NL0029949 (WBS 1.3.1.600)]; US Department of Energy [DE-EE0006269]Open access articleThis 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]

Detection of Abrin-Like and Prepropulchellin-Like Toxin Genes and Transcripts Using Whole Genome Sequencing and Full-Length Transcript Sequencing of Abrus precatorius

The sequenced genome and the leaf transcriptome of a near relative of Abrus pulchellus and Abrus precatorius was analyzed to characterize the genetic basis of toxin gene expression. From the high-quality genome assembly, a total of 26 potential coding regions were identified that contain genes with abrin-like, pulchellin-like, and agglutinin-like homology, with full-length transcripts detected in leaf tissue for 9 of the 26 coding regions. All of the toxin-like genes were identified within only five isolated regions of the genome, with each region containing 1 to 16 gene variants within each genomic region (<1 Mbp). The Abrusprecatorius cultivar sequenced here contains genes which encode for proteins that are homologous to certain abrin and prepropulchellin genes previously identified, and we observed substantial diversity of genes and predicted gene products in Abrus precatorius and previously characterized toxins. This suggests diverse toxin repertoires within Abrus, potentially the results of rapid toxin evolution.Open 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]

Comprehensive homing endonuclease target site specificity profiling reveals evolutionary constraints and enables genome engineering applications

Homing endonucleases (HEs) promote the evolutionary persistence of selfish DNA elements by catalyzing element lateral transfer into new host organisms. The high site specificity of this lateral transfer reaction, termed homing, reflects both the length (14–40 bp) and the limited tolerance of target or homing sites for base pair changes. In order to better understand molecular determinants of homing, we systematically determined the binding and cleavage properties of all single base pair variant target sites of the canonical LAGLIDADG homing endonucleases I-CreI and I-MsoI. These Chlorophyta algal HEs have very similar three-dimensional folds and recognize nearly identical 22 bp target sites, but use substantially different sets of DNA-protein contacts to mediate site-specific recognition and cleavage. The site specificity differences between I-CreI and I-MsoI suggest different evolutionary strategies for HE persistence. These differences also provide practical guidance in target site finding, and in the generation of HE variants with high site specificity and cleavage activity, to enable genome engineering applications

Addressing the pervasive scarcity of structural annotation in eukaryotic algae

Abstract Despite a continuous increase in algal genome sequencing, structural annotations of most algal genome assemblies remain unavailable. This pervasive scarcity of genome annotation has restricted rigorous investigation of these genomic resources and may have precipitated misleading biological interpretations. However, the annotation process for eukaryotic algal species is often challenging as genomic resources and transcriptomic evidence are not always available. To address this challenge, we benchmark the cutting-edge gene prediction methods that can be generalized for a broad range of non-model eukaryotes. Using the most accurate methods selected based on high-quality algal genomes, we predict structural annotations for 135 unannotated algal genomes. Using previously available genomic data pooled together with new data obtained in this study, we identified the core orthologous genes and the multi-gene phylogeny of eukaryotic algae, including of previously unexplored algal species. This study not only provides a benchmark for the use of structural annotation methods on a variety of non-model eukaryotes, but also compensates for missing data in the current spectrum of algal genomic resources. These results bring us one step closer to the full potential of eukaryotic algal genomics

Transposon insertional mutagenesis in Saccharomyces uvarum reveals trans-acting effects influencing species-dependent essential genes

To understand how complex genetic networks perform and regulate diverse cellular processes, the function of each individual component must be defined. Comprehensive phenotypic studies of mutant alleles have been successful in model organisms in determining what processes depend on the normal function of a gene. These results are often ported to newly sequenced genomes by using sequence homology. However, sequence similarity does not always mean identical function or phenotype, suggesting that new methods are required to functionally annotate newly sequenced species. We have implemented comparative analysis by high-throughput experimental testing of gene dispensability in Saccharomyces uvarum, a sister species of Saccharomyces cerevisiae. We created haploid and heterozygous diploid Tn7 insertional mutagenesis libraries in S. uvarum to identify species-dependent essential genes, with the goal of detecting genes with divergent functions and/or different genetic interactions. Comprehensive gene dispensability comparisons with S. cerevisiae predicted diverged dispensability at 12% of conserved orthologs, and validation experiments confirmed 22 differentially essential genes. Despite their differences in essentiality, these genes were capable of cross-species complementation, demonstrating that trans-acting factors that are background-dependent contribute to differential gene essentiality. This study shows that direct experimental testing of gene disruption phenotypes across species can inform comparative genomic analyses and improve gene annotations. Our method can be widely applied in microorganisms to further our understanding of genome evolution

Genome Sequence and Transcriptome Analyses of <i>Chrysochromulina tobin</i>: Metabolic Tools for Enhanced Algal Fitness in the Prominent Order Prymnesiales (Haptophyceae)

<div><p>Haptophytes are recognized as seminal players in aquatic ecosystem function. These algae are important in global carbon sequestration, form destructive harmful blooms, and given their rich fatty acid content, serve as a highly nutritive food source to a broad range of eco-cohorts. Haptophyte dominance in both fresh and marine waters is supported by the mixotrophic nature of many taxa. Despite their importance the nuclear genome sequence of only one haptophyte, <i>Emiliania huxleyi</i> (Isochrysidales), is available. Here we report the draft genome sequence of <i>Chrysochromulina tobin</i> (Prymnesiales), and transcriptome data collected at seven time points over a 24-hour light/dark cycle. The nuclear genome of <i>C</i>. <i>tobin</i> is small (59 Mb), compact (∼40% of the genome is protein coding) and encodes approximately 16,777 genes. Genes important to fatty acid synthesis, modification, and catabolism show distinct patterns of expression when monitored over the circadian photoperiod. The <i>C</i>. <i>tobin</i> genome harbors the first hybrid polyketide synthase/non-ribosomal peptide synthase gene complex reported for an algal species, and encodes potential anti-microbial peptides and proteins involved in multidrug and toxic compound extrusion. A new haptophyte xanthorhodopsin was also identified, together with two “red” RuBisCO activases that are shared across many algal lineages. The <i>Chrysochromulina tobin</i> genome sequence provides new information on the evolutionary history, ecology and economic importance of haptophytes.</p></div