Evolution of eukaryotic transcription : insights from the genome of Giardia lamblia

Author Posting. © Cold Spring Harbor Laboratory Press, 2004. This article is posted here by permission of Cold Spring Harbor Laboratory Press for personal use, not for redistribution. The definitive version was published in Genome Research 14 (2004): 1537-1547, doi:10.1101/gr.2256604.The Giardia lamblia genome sequencing project affords us a unique opportunity to conduct comparative analyses of core cellular systems between early and late-diverging eukaryotes on a genome-wide scale. We report a survey to identify canonical transcription components in Giardia, focusing on RNA polymerase (RNAP) subunits and transcription-initiation factors. Our survey revealed that Giardia contains homologs to 21 of the 28 polypeptides comprising eukaryal RNAPI, RNAPII, and RNAPIII; six of the seven RNAP subunits without giardial homologs are polymerase specific. Components of only four of the 12 general transcription initiation factors have giardial homologs. Surprisingly, giardial TATA-binding protein (TBP) is highly divergent with respect to archaeal and higher eukaryotic TBPs, and a giardial homolog of transcription factor IIB was not identified. We conclude that Giardia represents a transition during the evolution of eukaryal transcription systems, exhibiting a relatively complete set of RNAP subunits and a rudimentary basal initiation apparatus for each transcription system. Most class-specific RNAP subunits and basal initiation factors appear to have evolved after the divergence of Giardia from the main eukaryotic line of descent. Consequently, Giardia is predicted to be unique in many aspects of transcription initiation with respect to paradigms derived from studies in crown eukaryotes.This work was supported in part by NIH grant AI43273 to M.L.S., by NIH grant AI51089 to A.G.M, and DOE grant DE-FG02-01ER63201 to G.J.O. Additional support was provided by the G. Unger Vetlesen Foundation and LI-COR Biotechnology

Molecular evolution of the vesicle coat component βCOP in Toxoplasma gondii

Author Posting. © The Author(s), 2007. This is the author's version of the work. It is posted here by permission of Elsevier for personal use, not for redistribution. The definitive version was published in Molecular Phylogenetics and Evolution 44 (2007): 1284-1294, doi:10.1016/j.ympev.2007.02.031.Coatomer coated (COPI) vesicles play a pivotal role for multiple membrane trafficking steps throughout the eukaryotic cell. Our focus is on βCOP, one of the most well known components of the COPI multi-protein complex. Amino acid differences in βCOP may dictate functional divergence across species during the course of evolution, especially with regards to the evolutionary pressures on obligate intracellular parasites. A bioinformatic analysis of βCOP amino acid sequences was conducted for 49 eukaryotic species. Cloning and sequence analysis of the Toxoplasma gondii βCOP homologue revealed several amino acid insertions unique to T. gondii and one C-terminal insertion that is unique to apicomplexan parasites. These findings led us to investigate the possibility that βCOP experienced functional divergence during the course of its evolution. Bayesian phylogenetic analysis revealed a tree consistent with pan eukaryote distribution and long-branch lengths were observed among the apicomplexans. Further analysis revealed that kinetoplast βCOP underwent the most amount of change, leading to perhaps an overall change of function. In comparison, T. gondii exhibited subtle yet specific amino acid changes. The amino acid substitutions did not occur in the same places as other lineages, suggesting that TgβCOP has a role specific to the apicomplexans. Our work identifies forty-eight residues that are likely to be functionally important when comparing apicomplexan, kinetoplastid, and fungal βCOP.KMH is an Ellison Medical Foundation New Scholar in Infectious Disease; SLP was supported by an NIH training grant (NIH/NIAID/TMP T 32 7030); AGM was supported by the Marine Biological Laboratory’s Program in Global Infectious Diseases, also funded by the Ellison Medical Foundation

Understanding the α-crystallin cell membrane conjunction

PURPOSE. It is well established that levels of soluble α-crystallin in the lens cytoplasm fall steadily with age, accompanied by a corresponding increase in the amount of membrane-bound α-crystallin. Less well understood, is the mechanism driving this age-dependent membrane association. The aim of this study was to investigate the role of the membrane and its associated proteins and peptides in the binding of α-crystallin. METHODS. Fibre cell membranes from human and bovine lenses were separated from soluble proteins by centrifugation. Membranes were stripped of associated proteins with successive aqueous, urea and alkaline solutions. Protein constituents of the respective membrane isolates were examined by SDS-PAGE and Western immunoblotting. Recombinant αA- and αB-crystallins were fluorescently-labeled with Alexa350® dye and incubated with the membrane isolates and the binding capacity of membrane for α-crystallin was determined. RESULTS. The binding capacity of human membranes was consistently higher than that of bovine membranes. Urea- and alkali-treated membranes from the nucleus had similar binding capacities for αA-crystallin, which were significantly higher than both cortical membrane extracts. αB-Crystallin also had a higher affinity for nuclear membrane. However, urea-treated nuclear membrane had three times the binding capacity for αB-crystallin as compared to the alkali-treated nuclear membrane. Modulation of the membrane-crystallin interaction was achieved by the inclusion of an N-terminal peptide of αB-crystallin in the assays, which significantly increased the binding. Remarkably, following extraction with alkali, full length αA- and αB-crystallins were found to remain associated with both bovine and human lens membranes. CONCLUSIONS. Fiber cell membrane isolated from the lens has an inherent capacity to bind α-crystallin. For αB-crystallin, this binding was found to be proportional to the level of extrinsic membrane proteins in cells isolated from the lens nucleus, indicating these proteins may play a role in the recruitment of αB-crystallin. No such relationship was evident for αA-crystallin in the nucleus, or for cortical membrane binding. Intrinsic lens peptides, which increase in abundance with age, may also function to modulate the interaction between soluble α-crystallin and the membrane. In addition, the tight association between α-crystallin and the lens membrane suggests that the protein may be an intrinsic component of the membrane structure

Gene duplication and divergence produce divergent MHC genotypes without disassortative mating

Genes of the major histocompatibility complex (MHC) exhibit heterozygote advantage in immune defence, which in turn can select for MHC-disassortative mate choice. However, many species lack this expected pattern of MHC-disassortative mating. A possible explanation lies in evolutionary processes following gene duplication: if two duplicated MHC genes become functionally diverged from each other, offspring will inherit diverse multilocus genotypes even under random mating. We used locus-specific primers for high-throughput sequencing of two expressed MHC Class II B genes in Leach\u27s storm-petrels, Oceanodroma leucorhoa, and found that exon 2 alleles fall into two gene-specific monophyletic clades. We tested for disassortative vs. random mating at these two functionally diverged Class II B genes, using multiple metrics and different subsets of exon 2 sequence data. With good statistical power, we consistently found random assortment of mates at MHC. Despite random mating, birds had MHC genotypes with functionally diverged alleles, averaging 13 amino acid differences in pairwise comparisons of exon 2 alleles within individuals. To test whether this high MHC diversity in individuals is driven by evolutionary divergence of the two duplicated genes, we built a phylogenetic permutation model. The model showed that genotypic diversity was strongly impacted by sequence divergence between the most common allele of each gene, with a smaller additional impact of monophyly of the two genes. Divergence of allele sequences between genes may have reduced the benefits of actively seeking MHC-dissimilar mates, in which case the evolutionary history of duplicated genes is shaping the adaptive landscape of sexual selection

The cytochrome P450 (CYP) superfamily in cnidarians

© The Author(s), 2021. This article is distributed under the terms of the Creative Commons Attribution License. The definitive version was published in Pankov, K., McArthur, A. G., Gold, D. A., Nelson, D. R., Goldstone, J., & Wilson, J. Y. The cytochrome P450 (CYP) superfamily in cnidarians. Scientific Reports, 11(1), (2021): 9834, https://doi.org/10.1038/s41598-021-88700-y.The cytochrome P450 (CYP) superfamily is a diverse and important enzyme family, playing a central role in chemical defense and in synthesis and metabolism of major biological signaling molecules. The CYPomes of four cnidarian genomes (Hydra vulgaris, Acropora digitifera, Aurelia aurita, Nematostella vectensis) were annotated; phylogenetic analyses determined the evolutionary relationships amongst the sequences and with existing metazoan CYPs. 155 functional CYPs were identified and 90 fragments. Genes were from 24 new CYP families and several new subfamilies; genes were in 9 of the 12 established metazoan CYP clans. All species had large expansions of clan 2 diversity, with H. vulgaris having reduced diversity for both clan 3 and mitochondrial clan. We identified potential candidates for xenobiotic metabolism and steroidogenesis. That each genome contained multiple, novel CYP families may reflect the large evolutionary distance within the cnidarians, unique physiology in the cnidarian classes, and/or different ecology of the individual species.This study was supported by grants from the Natural Science Engineering Research Council of Canada in the form of a Discovery Grant (RGPIN-328204-2011 and RGPIN-05767-2016) to J.Y.W. A.G.M. held a Cisco Research Chair in Bioinformatics, supported by Cisco Systems Canada, Inc. K.V.P. was supported by a Natural Sciences and Engineering Research Council of Canada (NSERC) Undergraduate Student Research Award (USRA). Computer resources were supplied by the McMaster Service Lab and Repository computing cluster, funded in part by grants to A.G.M. from the Canadian Foundation for Innovation (34531 to A.G.M.)

Long serial analysis of gene expression for gene discovery and transcriptome profiling in the widespread marine coccolithophore Emiliania huxleyi

Author Posting. © American Society for Microbiology, 2006. This article is posted here by permission of American Society for Microbiology for personal use, not for redistribution. The definitive version was published in Applied and Environmental Microbiology 72 (2006): 252-260, doi:10.1128/AEM.72.1.252-260.2006.The abundant and widespread coccolithophore Emiliania huxleyi plays an important role in mediating CO2 exchange between the ocean and the atmosphere through its impact on marine photosynthesis and calcification. Here, we use long serial analysis of gene expression (SAGE) to identify E. huxleyi genes responsive to nitrogen (N) or phosphorus (P) starvation. Long SAGE is an elegant approach for examining quantitative and comprehensive gene expression patterns without a priori knowledge of gene sequences via the detection of 21-bp nucleotide sequence tags. E. huxleyi appears to have a robust transcriptional-level response to macronutrient deficiency, with 42 tags uniquely present or up-regulated twofold or greater in the N-starved library and 128 tags uniquely present or up-regulated twofold or greater in the P-starved library. The expression patterns of several tags were validated with reverse transcriptase PCR. Roughly 48% of these differentially expressed tags could be mapped to publicly available genomic or expressed sequence tag (EST) sequence data. For example, in the P-starved library a number of the tags mapped to genes with a role in P scavenging, including a putative phosphate-repressible permease and a putative polyphosphate synthetase. In short, the long SAGE analyses have (i) identified many new differentially regulated gene sequences, (ii) assigned regulation data to EST sequences with no database homology and unknown function, and (iii) highlighted previously uncharacterized aspects of E. huxleyi N and P physiology. To this end, our long SAGE libraries provide a new public resource for gene discovery and transcriptional analysis in this biogeochemically important marine organism.This work was supported by the Woods Hole Oceanographic Institution Ocean Life Institute, the J. Lamar Worzel Assistant Scientist Fund, and the Frank and Lisina Hoch Endowed Fund. A.G.M., S.R.B., and M.J.C. were supported in part by the Marine Biological Laboratory's Program in Global Infectious Diseases, funded by the Ellison Medical Foundation. Computational resources were provided by the Josephine Bay Paul Center for Comparative Molecular Biology and Evolution (Marine Biological Laboratory) through funds provided by the W. M. Keck Foundation and the G. Unger Vetlesen Foundation

Evidence for lateral transfer of genes encoding ferredoxins, nitroreductases, NADH oxidase, and alcohol dehydrogenase 3 from anaerobic prokaryotes to Giardia lamblia and Entamoeba histolytica

Author Posting. © American Society for Microbiology, 2002. This article is posted here by permission of American Society for Microbiology for personal use, not for redistribution. The definitive version was published in Eukaryotic Cell 1 (2002): 181-190, doi:10.1128/EC.1.2.181-190.2002.Giardia lamblia and Entamoeba histolytica are amitochondriate, microaerophilic protists which use fermentation enzymes like those of bacteria to survive anaerobic conditions within the intestinal lumen. Genes encoding fermentation enzymes and related electron transport peptides (e.g., ferredoxins) in giardia organisms and amebae are hypothesized to be derived from either an ancient anaerobic eukaryote (amitochondriate fossil hypothesis), a mitochondrial endosymbiont (hydrogen hypothesis), or anaerobic bacteria (lateral transfer hypothesis). The goals here were to complete the molecular characterization of giardial and amebic fermentation enzymes and to determine the origins of the genes encoding them, when possible. A putative giardia [2Fe-2S]ferredoxin which had a hypothetical organelle-targeting sequence at its N terminus showed similarity to mitochondrial ferredoxins and the hydrogenosomal ferredoxin of Trichomonas vaginalis (another luminal protist). However, phylogenetic trees were star shaped, with weak bootstrap support, so we were unable to confirm or rule out the endosymbiotic origin of the giardia [2Fe-2S]ferredoxin gene. Putative giardial and amebic 6-kDa ferredoxins, ferredoxin-nitroreductase fusion proteins, and oxygen-insensitive nitroreductases each tentatively supported the lateral transfer hypothesis. Although there were not enough sequences to perform meaningful phylogenetic analyses, the unique common occurrence of these peptides and enzymes in giardia organisms, amebae, and the few anaerobic prokaryotes suggests the possibility of lateral transfer. In contrast, there was more robust phylogenetic evidence for the lateral transfer of G. lamblia genes encoding an NADH oxidase from a gram-positive coccus and a microbial group 3 alcohol dehydrogenase from thermoanaerobic prokaryotes. In further support of lateral transfer, the G. lamblia NADH oxidase and adh3 genes appeared to have an evolutionary history distinct from those of E. histolytica.This work was supported by NIH grants (AI33492 to J.S., AI43273 to M.L.S., and AI46516 to B.J.L.). Additional support was provided by the G. Unger Vetlesen Foundation and LI-COR Biotechnology

Differential gene expression between fall- and spring-run Chinook salmon assessed by long serial analysis of gene expression

Author Posting. © American Fisheries Society, 2008. This article is posted here by permission of American Fisheries Society for personal use, not for redistribution. The definitive version was published in Transactions of the American Fisheries Society 137 (2008): 1378–1388, doi:10.1577/T07-222.1.Of all Pacific salmonids, Chinook salmon Oncorhynchus tshawytscha display the greatest variability in return times to freshwater. The molecular mechanisms of these differential return times have not been well described. Current methods, such as long serial analysis of gene expression (LongSAGE) and microarrays, allow gene expression to be analyzed for thousands of genes simultaneously. To investigate whether differential gene expression is observed between fall- and spring-run Chinook salmon from California's Central Valley, LongSAGE libraries were constructed. Three libraries containing between 25,512 and 29,372 sequenced tags (21 base pairs/tag) were generated using messenger RNA from the brains of adult Chinook salmon returning in fall and spring and from one ocean-caught Chinook salmon. Tags were annotated to genes using complementary DNA libraries from Atlantic salmon Salmo salar and rainbow trout O. mykiss. Differentially expressed genes, as estimated by differences in the number of sequence tags, were found in all pairwise comparisons of libraries (freshwater versus saltwater = 40 genes; fall versus spring = 11 genes; and spawning versus nonspawning = 51 genes). The gene for ependymin, an extracellular glycoprotein involved in behavioral plasticity in fish, exhibited the most differential expression among the three groupings. Reverse transcription polymerase chain reaction analysis verified the differential expression of ependymin between the fall- and spring-run samples. These LongSAGE libraries, the first reported for Chinook salmon, provide a window of the transcriptional changes during Chinook salmon return migration to freshwater and spawning and increase the amount of expressed sequence data.This work was supported with a grant from the California Department of Water Resources awarded to M.A.B.; J.C.B. received additional funding from the North Umpqua Foundation, Roseburg, Oregon

Differential gene expression between fall- and spring-run Chinook salmon assessed by long serial analysis of gene expression

Of all Pacific salmonids, Chinook salmon Oncorhynchus tshawytscha display the greatest variability in return times to freshwater. The molecular mechanisms of these differential return times have not been well described. Current methods, such as long serial analysis of gene expression (LongSAGE) and microarrays, allow gene expression to be analyzed for thousands of genes simultaneously. To investigate whether differential gene expression is observed between fall- and spring-run Chinook salmon from California's Central Valley, LongSAGE libraries were constructed. Three libraries containing between 25,512 and 29,372 sequenced tags (21 base pairs/tag) were generated using messenger RNA from the brains of adult Chinook salmon returning in fall and spring and from one ocean-caught Chinook salmon. Tags were annotated to genes using complementary DNA libraries from Atlantic salmon Salmo salar and rainbow trout O. mykiss. Differentially expressed genes, as estimated by differences in the number of sequence tags, were found in all pairwise comparisons of libraries (freshwater versus saltwater = 40 genes; fall versus spring = 11 genes; and spawning versus nonspawning = 51 genes). The gene for ependymin, an extracellular glycoprotein involved in behavioral plasticity in fish, exhibited the most differential expression among the three groupings. Reverse transcription polymerase chain reaction analysis verified the differential expression of ependymin between the fall- and spring-run samples. These LongSAGE libraries, the first reported for Chinook salmon, provide a window of the transcriptional changes during Chinook salmon return migration to freshwater and spawning and increase the amount of expressed sequence data.Keywords: Oncorhynchus tshawytscha, migration, serial analysis of gene expressionKeywords: Oncorhynchus tshawytscha, migration, serial analysis of gene expressio

The First Extrasolar Planet Discovered with a New Generation High Throughput Doppler Instrument

We report the detection of the first extrasolar planet, ET-1 (HD 102195b), using the Exoplanet Tracker (ET), a new generation Doppler instrument. The planet orbits HD 102195, a young star with solar metallicity that may be part of the local association. The planet imparts radial velocity variability to the star with a semiamplitude of $63.4\pm2.0$ m s$^{-1}$ and a period of 4.11 days. The planetary minimum mass ($m \sin i$) is $0.488\pm0.015$ $M_J$.Comment: 42 pages, 11 figures and 5 tables, Accepted for publication in Ap