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Molecular and Cellular Biology Nov 1981Previously we showed that the mitochondrial deoxyribonucleic acid (DNA) from Paramecium aurelia consists of a linear genome and that replication of this genome is...
Previously we showed that the mitochondrial deoxyribonucleic acid (DNA) from Paramecium aurelia consists of a linear genome and that replication of this genome is initiated at one terminus and proceeds unidirectionally to the other terminus. Analyses of mitochondria from four closely related species (1, 4, 5, and 7) indicated that the species 1, 5, and 7 DNAs are essentially completely homologous but that the species 4 mitochondrial DNA is only 40 to 50% homologous with that from species 1. The major regions of homology are those containing the genes for ribosomal ribonucleic acid (RNA). To understand the replication and organization of the linear mitochondrial genome better, we compared species 1 (Paramecium primaurelia) and 4 (Paramecium tetraaurelia) DNAs with regard to restriction fragment mapping and homology between initiation regions; we also identified the sites of the genes for ribosomal RNA. In general, the structures of the species 1 and 4 mitochondrial genomes were quite similar. Each ribosomal RNA gene was present in one copy per genome, with the large ribosomal RNA gene located near the terminal region of replication and the small ribosomal RNA gene located more centrally. These two genes were separated by about 10 kilobases in the species 1 genome and by about 12 kilobases in the species 4 genome. In contrast to our previous findings, by using nonstringent hybridization conditions we detected homology between the species 1 and 4 DNA fragments containing the initiation regions. We constructed recombinant DNA clones for many fragments, especially those containing the initiation region and the ribosomal RNA genes. We also constructed restriction enzyme maps for six enzymes for both P. primaurelia and P. tetraaurelia.
Topics: Animals; Base Sequence; Cloning, Molecular; DNA Restriction Enzymes; DNA, Mitochondrial; DNA, Recombinant; DNA, Ribosomal; Nucleic Acid Hybridization; Paramecium
PubMed: 6287214
DOI: 10.1128/mcb.1.11.972-982.1981 -
PLoS Biology Sep 2022In metazoa, cilia assembly is a cellular process that starts with centriole to basal body maturation, migration to the cell surface, and docking to the plasma membrane....
In metazoa, cilia assembly is a cellular process that starts with centriole to basal body maturation, migration to the cell surface, and docking to the plasma membrane. Basal body docking involves the interaction of both the distal end of the basal body and the transition fibers/distal appendages, with the plasma membrane. Mutations in numerous genes involved in basal body docking and transition zone assembly are associated with the most severe ciliopathies, highlighting the importance of these events in cilium biogenesis. In this context, the ciliate Paramecium has been widely used as a model system to study basal body and cilia assembly. However, despite the evolutionary conservation of cilia assembly events across phyla, whether the same molecular players are functionally conserved, is not fully known. Here, we demonstrated that CEP90, FOPNL, and OFD1 are evolutionary conserved proteins crucial for ciliogenesis. Using ultrastructure expansion microscopy, we unveiled that these proteins localize at the distal end of both centrioles/basal bodies in Paramecium and mammalian cells. Moreover, we found that these proteins are recruited early during centriole duplication on the external surface of the procentriole. Functional analysis performed both in Paramecium and mammalian cells demonstrate the requirement of these proteins for distal appendage assembly and basal body docking. Finally, we show that mammalian centrioles require another component, Moonraker (MNR), to recruit OFD1, FOPNL, and CEP90, which will then recruit the distal appendage proteins CEP83, CEP89, and CEP164. Altogether, we propose that this OFD1, FOPNL, and CEP90 functional module is required to determine in mammalian cells the future position of distal appendage proteins.
Topics: Animals; Cell Membrane; Centrioles; Cilia; Mammals; Paramecium
PubMed: 36070319
DOI: 10.1371/journal.pbio.3001782 -
PloS One 2014The epigenetic influence of maternal cells on the development of their progeny has long been studied in various eukaryotes. Multicellular organisms usually provide their...
The epigenetic influence of maternal cells on the development of their progeny has long been studied in various eukaryotes. Multicellular organisms usually provide their zygotes not only with nutrients but also with functional elements required for proper development, such as coding and non-coding RNAs. These maternally deposited RNAs exhibit a variety of functions, from regulating gene expression to assuring genome integrity. In ciliates, such as Paramecium these RNAs participate in the programming of large-scale genome reorganization during development, distinguishing germline-limited DNA, which is excised, from somatic-destined DNA. Only a handful of proteins playing roles in this process have been identified so far, including typical RNAi-derived factors such as Dicer-like and Piwi proteins. Here we report and characterize two novel proteins, Pdsg1 and Pdsg2 (Paramecium protein involved in Development of the Somatic Genome 1 and 2), involved in Paramecium genome reorganization. We show that these proteins are necessary for the excision of germline-limited DNA during development and the survival of sexual progeny. Knockdown of PDSG1 and PDSG2 genes affects the populations of small RNAs known to be involved in the programming of DNA elimination (scanRNAs and iesRNAs) and chromatin modification patterns during development. Our results suggest an association between RNA-mediated trans-generational epigenetic signal and chromatin modifications in the process of Paramecium genome reorganization.
Topics: Cell Nucleus; Chromatin; DNA, Protozoan; Epigenesis, Genetic; Genome, Protozoan; Histones; Methylation; Microscopy, Confocal; Paramecium; Protozoan Proteins; RNA Interference; RNA, Small Interfering; Recombinant Fusion Proteins
PubMed: 25397898
DOI: 10.1371/journal.pone.0112899 -
Genetica May 2011The multiple species concepts currently in use by the scientific community (e.g. Morphological, Biological, Phylogenetic) are united in that they all aim to capture the...
The multiple species concepts currently in use by the scientific community (e.g. Morphological, Biological, Phylogenetic) are united in that they all aim to capture the process of divergence between populations. For example, the Biological Species Concept defines a species as a natural group of organisms that is reproductively isolated from other such groups. Here we synthesize nearly a century of research on the ciliate genus Paramecium that highlights the shortcomings of our prevailing notions on the nature of species. In this lineage, there is discordance between morphology, mating behavior, and genetics, features assumed to be correlated, at least after sufficient time has passed, under all species concepts. Intriguingly, epigenetic phenomena are well documented in ciliates where they influence features such as germline/soma differentiation and mating type determination. Consequently, we hypothesize that divergence within ciliate populations is due to a dynamic interaction between genetic and epigenetic factors. The growing list of examples of epigenetic phenomena that potentially impact speciation (i.e. by influencing the dynamics of sex chromosomes, fate of hybrids, zygotic drive and genomic conflicts) suggests that interactions between genetics and epigenetics may also drive divergence in other eukaryotic lineages.
Topics: Epigenomics; Genetic Speciation; Isoenzymes; Paramecium; Phylogeny
PubMed: 21505762
DOI: 10.1007/s10709-011-9571-3 -
Medecine Sciences : M/S Apr 2005Since the middle of the last century, Paramecium has appeared as an intriguing genetic model, displaying a variety of heritable characters which do not follow the Mendel... (Review)
Review
Since the middle of the last century, Paramecium has appeared as an intriguing genetic model, displaying a variety of heritable characters which do not follow the Mendel laws but are cytoplasmically inherited. The analysis of the hereditary mechanisms at play in this eukaryotic unicellular organism has provided new insight into epigenetics mechanisms. Interestingly, the revealing phenomena concern two pecularities of Paramecium, its highly elaborate surface structure (with thousands of ciliary basal bodies as cytoskeleton organizers), and its nuclear dualism (coexistence of a diploid "germline" micronucleus and a highly polyploid somatic macronucleus devoted to transcription, which contains a rearranged version of the germline genome). Analysis of variant cortical organization has led to the concept of structural inheritance, implying that assembly of new organelles and supramolecular protein complexes is guided by pre-existing organization. Analysis of other cytoplasmically inherited characters revealed that the developing macronucleus is epigenetically programmed by the maternal macronucleus through RNA-mediated, homology-dependent effects, suggesting the transcriptome should be recognized as a third actor in cellular inheritance, along with the "structurome" and the genome.
Topics: Animals; Cell Nucleus; Epigenesis, Genetic; Paramecium
PubMed: 15811302
DOI: 10.1051/medsci/2005214377 -
BMC Biology Nov 2020Ciliates are an ancient and diverse eukaryotic group found in various environments. A unique feature of ciliates is their nuclear dimorphism, by which two types of...
BACKGROUND
Ciliates are an ancient and diverse eukaryotic group found in various environments. A unique feature of ciliates is their nuclear dimorphism, by which two types of nuclei, the diploid germline micronucleus (MIC) and polyploidy somatic macronucleus (MAC), are present in the same cytoplasm and serve different functions. During each sexual cycle, ciliates develop a new macronucleus in which newly fused genomes are extensively rearranged to generate functional minichromosomes. Interestingly, each ciliate species seems to have its way of processing genomes, providing a diversity of resources for studying genome plasticity and its regulation. Here, we sequenced and analyzed the macronuclear genome of different strains of Paramecium bursaria, a highly divergent species of the genus Paramecium which can stably establish endosymbioses with green algae.
RESULTS
We assembled a high-quality macronuclear genome of P. bursaria and further refined genome annotation by comparing population genomic data. We identified several species-specific expansions in protein families and gene lineages that are potentially associated with endosymbiosis. Moreover, we observed an intensive chromosome breakage pattern that occurred during or shortly after sexual reproduction and contributed to highly variable gene dosage throughout the genome. However, patterns of copy number variation were highly correlated among genetically divergent strains, suggesting that copy number is adjusted by some regulatory mechanisms or natural selection. Further analysis showed that genes with low copy number variation among populations tended to function in basic cellular pathways, whereas highly variable genes were enriched in environmental response pathways.
CONCLUSIONS
We report programmed DNA rearrangements in the P. bursaria macronuclear genome that allow cells to adjust gene copy number globally according to individual gene functions. Our results suggest that large-scale gene copy number variation may represent an ancient mechanism for cells to adapt to different environments.
Topics: Genome, Protozoan; Macronucleus; Metagenomics; Paramecium
PubMed: 33250052
DOI: 10.1186/s12915-020-00912-2 -
Biochimica Et Biophysica Acta.... Jan 2023Ever since the pioneering studies in the 1960s and 70s, the importance of order transitions for cell membrane functions has remained a matter of debate. Recently, it has...
Ever since the pioneering studies in the 1960s and 70s, the importance of order transitions for cell membrane functions has remained a matter of debate. Recently, it has been proposed that the nonlinear stimulus-response curve of excitable cells, which manifests in all-or-none pulses (action potentials (AP)), is due to a transition in the cell membrane. Indeed, evidence for transitions has accumulated in plant cells and neurons, but studies with other excitable cells are expedient in order to show if this finding is of a general nature. Herein, we investigated intact, motile specimens of the "swimming neuron" Paramecium. The cellular membranes were labelled with the solvatochromic fluorophores LAURDAN or Di-4-ANEPPDHQ. Subsequently, a cell was trapped in a microfluidic channel and investigated by fluorescence spectroscopy. The generalized polarization (GP) of the fluorescence emission from cell cortical membranes (probably plasma and alveolar membranes) was extracted by an edge-finding algorithm. The thermo-optical state diagram, i.e. the dependence of GP on temperature, exhibited clear indications for a reversible transition. This transition had a width of ~10-15 °C and a midpoint that was located ~4 °C below the growth temperature. The state diagrams with LAURDAN and Di-4-ANEPPDHQ had widely identical characteristics. These results suggested that the cortical membranes of Paramecium reside in an order transition regime under physiological growth conditions. Based on these findings, membrane potential fluctuations, spontaneous depolarizing spikes, and thermal excitation of Paramecium was interpreted.
Topics: Paramecium; Laurates; 2-Naphthylamine; Membranes
PubMed: 36243036
DOI: 10.1016/j.bbamem.2022.184073 -
Scientific Reports Apr 2016Legionella pneumophila, the causative agent of Legionnaires' disease, replicates within alveolar macrophages and free-living amoebae. However, the lifestyle of L....
Legionella pneumophila, the causative agent of Legionnaires' disease, replicates within alveolar macrophages and free-living amoebae. However, the lifestyle of L. pneumophila in the environment remains largely unknown. Here we established a novel natural host model of L. pneumophila endosymbiosis using the ciliate Paramecium caudatum. We also identified Legionella endosymbiosis-modulating factor A (LefA), which contributes to the change in life stage from endosymbiosis to host lysis, enabling escape to the environment. We isolated L. pneumophila strains from the environment, and they exhibited cytotoxicity toward P. caudatum and induced host lysis. Acidification of the Legionella-containing vacuole (LCV) was inhibited, and enlarged LCVs including numerous bacteria were observed in P. caudatum infected with L. pneumophila. An isogenic L. pneumophila lefA mutant exhibited decreased cytotoxicity toward P. caudatum and impaired the modification of LCVs, resulting in the establishment of endosymbiosis between them. Our results suggest that L. pneumophila may have a mechanism to switch their endosymbiosis in protistan hosts in the environment.
Topics: Cell Line; Disease Reservoirs; Gene Expression Regulation, Bacterial; Genes, Bacterial; Host-Pathogen Interactions; Humans; Legionella pneumophila; Legionnaires' Disease; Macrophages; Mutation; Paramecium; Symbiosis
PubMed: 27079173
DOI: 10.1038/srep24322 -
PLoS Biology Mar 2020Ciliary shedding occurs from unicellular organisms to metazoans. Although required during the cell cycle and during neurogenesis, the process remains poorly understood....
Ciliary shedding occurs from unicellular organisms to metazoans. Although required during the cell cycle and during neurogenesis, the process remains poorly understood. In all cellular models, this phenomenon occurs distal to the transition zone (TZ), suggesting conserved molecular mechanisms. The TZ module proteins (Meckel Gruber syndrome [MKS]/Nephronophtysis [NPHP]/Centrosomal protein of 290 kDa [CEP290]/Retinitis pigmentosa GTPase regulator-Interacting Protein 1-Like Protein [RPGRIP1L]) are known to cooperate to establish TZ formation and function. To determine whether they control deciliation, we studied the function of 5 of them (Transmembrane protein 107 [TMEM107], Transmembrane protein 216 [TMEM216], CEP290, RPGRIP1L, and NPHP4) in Paramecium. All proteins are recruited to the TZ of growing cilia and localize with 9-fold symmetry at the level of the most distal part of the TZ. We demonstrate that depletion of the MKS2/TMEM216 and TMEM107 proteins induces constant deciliation of some cilia, while depletion of either NPHP4, CEP290, or RPGRIP1L prevents Ca2+/EtOH deciliation. Our results constitute the first evidence for a role of conserved TZ proteins in deciliation and open new directions for understanding motile cilia physiology.
Topics: Cell Proliferation; Cilia; Cytoskeletal Proteins; Gene Expression; Membrane Fusion; Paramecium tetraurelia; Protein Domains; Protozoan Proteins; RNA Interference
PubMed: 32163404
DOI: 10.1371/journal.pbio.3000640 -
Genome Research Apr 2022The unicellular ciliate contains a large vegetative macronucleus with several unusual characteristics, including an extremely high coding density and high polyploidy....
The unicellular ciliate contains a large vegetative macronucleus with several unusual characteristics, including an extremely high coding density and high polyploidy. As macronculear chromatin is devoid of heterochromatin, our study characterizes the functional epigenomic organization necessary for gene regulation and proper Pol II activity. Histone marks (H3K4me3, H3K9ac, H3K27me3) reveal no narrow peaks but broad domains along gene bodies, whereas intergenic regions are devoid of nucleosomes. Our data implicate H3K4me3 levels inside ORFs to be the main factor associated with gene expression, and H3K27me3 appears in association with H3K4me3 in plastic genes. Silent and lowly expressed genes show low nucleosome occupancy, suggesting that gene inactivation does not involve increased nucleosome occupancy and chromatin condensation. Because of a high occupancy of Pol II along highly expressed ORFs, transcriptional elongation appears to be quite different from that of other species. This is supported by missing heptameric repeats in the C-terminal domain of Pol II and a divergent elongation system. Our data imply that unoccupied DNA is the default state, whereas gene activation requires nucleosome recruitment together with broad domains of H3K4me3. In summary, gene activation and silencing in run counter to the current understanding of chromatin biology.
Topics: Chromatin; Histone Code; Histones; Nucleosomes; Paramecium; RNA Polymerase II
PubMed: 35264449
DOI: 10.1101/gr.276126.121