Alternative Processing of Scrambled Genes Generates Protein Diversity in the Ciliate \u3ci\u3eChilodonella uncinata\u3c/i\u3e

In ciliates, chromosomal rearrangements occur during the development of the somatic macronuclear genome from the germline micronuclear genome. These rearrangements are extensive in three ciliate classes-Armophorea, Spirotrichea, and Phyllopharyngea-generating a macronucleus with up to 20,000,000 gene-sized chromosomes. Earlier, we have shown that these three classes also share elevated rates of protein evolution relative to other ciliates. To assess the evolution of germline-limited sequences in the class Phyllopharyngea, we used a combination of traditional and walking PCR to analyze micronuclear copies of multiple genes from two lines of the morphospecies Chilodonella uncinata for which we had previously characterized macronuclear sequences. Analyses of the resulting data yield three main results: (1) conserved macronuclear (somatic) regions are found within rapidly evolving micronuclear (germline) regions; (2) gene scrambling exists within this ciliate lineage; and (3) alternative processing of micronuclear regions yields diverse macronuclear β-tubulin paralogs. To our knowledge, this is the first study to demonstrate gene scrambling outside the nonsister class Spirotrichea, and to show that alternative processing of scrambled genes generates diversity in gene families. Intriguingly, the Spirotrichea and Phyllopharyngea are also united in having transient giant polytene chromosomes, gene-sized somatic chromosomes, and elevated rates of protein evolution. We hypothesize that this suite of characters enables these ciliates to enjoy the benefits of asexuality while still maintaining the ability to go through sexual cycles. The data presented here add to the growing evidence of the dynamic nature of eukaryotic genomes within diverse lineages across the tree of life

The Dynamic Nature of Genomes Across the Tree of Life

Genomes are dynamic in lineages across the tree of life. Among bacteria and archaea, for example, DNA content varies through out life cycles, and nonbinary cell division in diverse lineages indicates the need for coordination of the inheritance of genomes. These observations contrast with the textbook view that bacterial and archaeal genomes are monoploid (i.e., single copied) and fixed both within species and throughout an individual\u27s lifetime. Here, we synthesize information on three aspects of dynamic genomes from exemplars representing a diverse array of bacterial and archaeal lineages: 1) ploidy level variation, 2) epigenetic mechanisms, and 3) life cycle variation. For example, the Euryarchaeota analyzed to date are all polyploid, as is the bacterium Epulopiscium that contains up to tens of thousands of copies of its genome and reproduces by viviparity. The bacterium Deinococcus radiodurans and the archaeon Halobacterium sp. NRC-1 can repair a highly fragmented genome within a few hours. Moreover, bacterial genera such as Dermocarpella and Planctomyces reproduce by fission (i.e., generating many cells from one cell) and budding, respectively, highlighting the need for regulation of genome inheritance in these lineages. Combining these data with our previous work on widespread genome dynamics among eukaryotes, we hypothesize that dynamic genomes are a rule rather than the exception across the tree of life. Further, we speculate that all domains may have the ability to distinguish germline from somatic DNA and that this ability may have been present the last universal common ancestor

Epigenetic Influences of Mobile Genetic Elements on Ciliate Genome Architecture and Evolution

Mobile genetic elements (MGEs) are transient genetic material that can move either within a single organism\u27s genome or between individuals or species. While historically considered “junk” DNA (i.e., deleterious or at best neutral), more recent studies reveal the potential adaptive advantages MGEs provide in lineages across the tree of life. Ciliates, a group of single-celled microbial eukaryotes characterized by nuclear dimorphism, exemplify how epigenetic influences from MGEs shape genome architecture and patterns of molecular evolution. Ciliate nuclear dimorphism may have evolved as a response to transposon invasion and ciliates have since co-opted transposons to carry out programmed DNA deletion. Another example of the effect of MGEs is in providing mechanisms for lateral gene transfer (LGT) from bacteria, which introduces genetic diversity and, in several cases, may drive ecological specialization in ciliates. As a third example, the integration of viral DNA, likely through transduction, provides new genetic materials and can change the way host cells defend themselves against other viral pathogens. We argue that the acquisition of MGEs through non-Mendelian patterns of inheritance, coupled with their effects on ciliate genome architecture and persistence throughout evolutionary history, exemplify how the transmission of mobile elements should be considered a mechanism of transgenerational epigenetic inheritance

Are Microbes Fundamentally Different Than Macroorganisms? Convergence and a Possible Case for Neutral Phenotypic Evolution in Testate Amoeba (Amoebozoa: Arcellinida)

This study reveals extensive phenotypic convergence based on the non-monophyly of genera and morphospecies of testate (shelled) amoebae. Using two independent markers, small subunit ribosomal DNA (ssu-rDNA) and mitochondrial cytochrome oxidase I (COI), we demonstrate discordance between morphology and molecules for ‘core Nebela’ species (Arcellinida; Amoebozoa). Prior work using just a single locus, ssu-rDNA, also supported the non-monophyly of the genera Hyalosphenia and Nebela as well as for several morphospecies within these genera. Here, we obtained COI gene sequences of 59 specimens from seven morphospecies and ssu-rDNA gene sequences of 50 specimens from six morphospecies of hyalosphenids. Our analyses corroborate the prior ssu-rDNA findings of morphological convergence in test (shell) morphologies, as COI and ssu-rDNA phylogenies are concordant. Further, the monophyly of morphospecies is rejected using approximately unbiased tests. Given that testate amoebae are used as bioindicators in both palaeoecological and contemporary studies of threatened ecosystems such as bogs and fens, understanding the discordance between morphology and genetics in the hyalosphenids is essential for interpretation of indicator species. Further, while convergence is normally considered the result of natural selection, it is possible that neutrality underlies phenotypic evolution in these microorganisms

Interpreting the seasonal cycles of atmospheric oxygen and carbon dioxide concentrations at American Samoa Observatory

We present seven years of atmospheric O2/N2 ratio and CO2 concentration data measured from flask samples collected at American Samoa. These data are unusual, exhibiting higher short-term variability, and seasonal cycles not in phase with other sampling stations. The unique nature of atmospheric data from Samoa has been noted previously from measurements of CO2, methyl chloroform, and ozone. With our O2 data, we observe greater magnitude in the short-term variability, but, in contrast, no clear seasonal pattern to this variability. This we attribute to significant regional sources and sinks existing for O2 in both hemispheres, and a dependence on both the latitudinal and altitudinal origins of air masses. We also hypothesize that some samples exhibit a component of "older" air, demonstrating recirculation of air within the tropics. Our findings could be used to help constrain atmospheric transport models which are not well characterized in tropical regions

Nuclear Features of the Heterotrich Ciliate \u3ci\u3eBlepharisma americanum\u3c/i\u3e: Genomic Amplification, Life Cycle, and Nuclear Inclusion

Blepharisma americanum, a member of the understudied ciliate class Heterotrichea, has a moniliform somatic macronucleus that resembles beads on a string. Blepharisma americanum is distinguishable by its pink coloration derived from the autofluorescent pigment blepharismin and tends to have a single somatic macronucleus with 3–6 nodes and multiple germline micronuclei. We used fluorescence confocal microscopy to explore the DNA content and amplification between the somatic and germline nuclei of B. americanum through its life cycle. We estimate that the DNA content of the macronucleus and micronucleus are 43 ± 8 Gbp and 83 ± 16 Mbp respectively. This correlates with an approximate DNA content difference of 500-fold from micronucleus to macronucleus and a macronuclear ploidy of ~1,100 N as compared to the presumably diploid micronucleus. We also investigate a previously reported macronuclear inclusion, which is present sporadically across all life cycle stages; this inclusion looks as if it contains blepharismin based on its fluorescent properties, but its function remains unknown. We also provide additional detail to our understanding of life cycles changes in B. americanum by analyses of fluorescent images. Overall, the data analyzed here contribute to our understanding of the diversity of nuclear architecture in ciliates by providing details on the highly polyploid somatic macronucleus of B. americanum

Microbial Diversity in the Eukaryotic SAR Clade: Illuminating the Darkness Between Morphology and Molecular Data

Despite their diversity and ecological importance, many areas of the SAR—Stramenopila, Alveolata, and Rhizaria—clade are poorly understood as the majority (90%) of SAR species lack molecular data and only 5% of species are from well-sampled families. Here, we review and summarize the state of knowledge about the three major clades of SAR, describing the diversity within each clade and identifying synapomorphies when possible. We also assess the “dark area” of SAR: the morphologically described species that are missing molecular data. The majority of molecular data for SAR lineages are characterized from marine samples and vertebrate hosts, highlighting the need for additional research effort in areas such as freshwater and terrestrial habitats and “non-vertebrate” hosts. We also describe the paucity of data on the biogeography of SAR species, and point to opportunities to illuminate diversity in this major eukaryotic clade. See also the video abstract above

Phylogenomic Analyses of 2,786 Genes in 158 Lineages Support a Root of the Eukaryotic Tree of Life between Opisthokonts and All Other Lineages

Advances in phylogenomics and high-throughput sequencing have allowed the reconstruction of deep phylogenetic relationships in the evolution of eukaryotes. Yet, the root of the eukaryotic tree of life remains elusive. The most popular hypothesis in textbooks and reviews is a root between Unikonta (Opisthokonta + Amoebozoa) and Bikonta (all other eukaryotes), which emerged from analyses of a single-gene fusion. Subsequent, highly cited studies based on concatenation of genes supported this hypothesis with some variations or proposed a root within Excavata. However, concatenation of genes does not consider phylogenetically-informative events like gene duplications and losses. A recent study using gene tree parsimony (GTP) suggested the root lies between Opisthokonta and all other eukaryotes, but only including 59 taxa and 20 genes. Here we use GTP with a duplication-loss model in a gene-rich and taxon-rich dataset (i.e., 2,786 gene families from two sets of 155 and 158 diverse eukaryotic lineages) to assess the root, and we iterate each analysis 100 times to quantify tree space uncertainty. We also contrasted our results and discarded alternative hypotheses from the literature using GTP and the likelihood-based method SpeciesRax. Our estimates suggest a root between Fungi or Opisthokonta and all other eukaryotes; but based on further analysis of genome size, we propose that the root between Opisthokonta and all other eukaryotes is the most likely

Morphological Description of Telaepolella tubasferens n. g., n. sp., Isolate ATCC© 50593™, a Filose Amoeba in the Gracilipodida, Amoebozoa

We describe the amoeboid isolate ATCC© 50593™ as a new taxon, Telaepolella tubasferens n. gen. n. sp. This multinucleated amoeba has filose pseudopods and is superficially similar to members of the vampyrellids (Rhizaria) such as Arachnula impatiens Cienkowski, 1876, which was the original identification upon deposition. However, previous multigene analyses place this taxon within the Gracilipodida Lahr and Katz 2011 in the Amoebozoa. Here, we document the morphology of this organism at multiple life history stages and describe data underlying the description as a new taxon. We demonstrate that T. tubaspherens is distinct from Arachnula and other rhizarians (Theratromyxa, Leptophrys) in a suite of morphological characters such as general body shape, relative size of pseudopods, distinction of ecto- and endoplasmic regions, and visibility of nuclei in non-stained cells (an important diagnostic character). Although Amoebozoa taxa generally have lobose pseudopods, genera in Gracilipodida such as Flamella and Filamoeba as well as several organisms previously classified as protosteloid amoebae (e.g. schizoplasmodiis, cavosteliids and Stemonitales) present filose pseudopodia. Thus, classification of amoeboid organisms merely by filose-lobose distinction must be reconsidered