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opencc-by-4.0Jul 2025View details →

A development-specific POLX essential for programmed genome rearrangement in Paramecium tetraurelia

During the sexual cycle, programmed genome rearrangement (PGR) in Paramecium tetraurelia involves the non-homologous end joining (NHEJ) DNA repair pathway to eliminate specific germinal Internal Eliminated Sequences (IESs) from the newly developing somatic nucleus. In addition to the core NHEJ factors Ku70/80 and Xrcc4/Lig4, additional enzymes are required to process the 4-base 5’-protruding ends generated following DNA cleavage at IES boundaries, prior to their ligation. Here, we report that PolX (a,b,c,d), four P. tetraurelia distant orthologs of the human Poll DNA polymerase, are involved in repair of IES excision junctions. During rearrangements, PolX-depleted cells accumulate genome-wide errors, such as unrepaired double-strand breaks, 1-nucleotide deletions and IES retention. Although all PolX paralogs can process DNA ends, two of them (PolXa&b) are induced during PGR and have acquired tight nuclear anchoring properties through their N-terminal region, which contains a predicted BRCT domain. Finally, we show that PolXa accumulates in nuclear foci together with other NHEJ proteins and the Dicer-like enzyme Dcl5, which is involved in the biogenesis of IES-specific small RNAs. We propose that these “DNA repair foci” correspond to the sites where IES concatemers, a by-product of IES excision, are ligated together to produce the precursors of iesRNAs.

Fig. 6 in Worldwide sampling reveals low genetic variability in populations of the freshwater ciliate Paramecium biaurelia (P. aurelia species complex, Ciliophora, Protozoa)

Fig. 6 Map of sampling sit_s of Paramecium biaurelia strains coll_ct_d during fi_ld r_s_arch in th_ Kraków ar_a. a Kraków, "At th_ brickyard" pond, 1 sampling point. b Kraków, Zaczarowana Dorożka Park (pond), 2 sampling points. c Pi_skowa Skała (pond), 1 sampling point. d Kraków,

Fig. 5 in Worldwide sampling reveals low genetic variability in populations of the freshwater ciliate Paramecium biaurelia (P. aurelia species complex, Ciliophora, Protozoa)

Fig. 5 Haplotyp_ n_twork of Paramecium biaurelia construct_d using th_ 123 s_qu_nc_s of ribosomal ITS1- 5.8S-ITS2-5'LSU fragm_nts (a) and 139 of mitochondrial COI fragm_nts (b). Th_ n_twork pr_s_nts a comparison of haplotyp_s obtain_d in th_ Kraków ar_a vs. th_ oth_r localiti_s, wh_r_ mol_cular data for P. biaurelia is availabl_. Black dash_s on particular branch_s r_pr_s_nt nucl_otid_ substitutions b_tw__n particular haplotyp_s. Analys_s w_r_ conduct_d using th_ M_dian Joining m_thod in PopART softwar_ v. 1.7

Fig. 4 in Worldwide sampling reveals low genetic variability in populations of the freshwater ciliate Paramecium biaurelia (P. aurelia species complex, Ciliophora, Protozoa)

Fig. 4 Haplotyp_ n_twork of Paramecium biaurelia construct_d using th_ 123 s_qu_nc_s of ribosomal ITS1- 5.8S-ITS2-5'LSU fragm_nts (a) and 139 of mitochondrial COI fragm_nts (b). Th_ n_twork pr_s_nts r_ciprocal r_lationships b_tw__n, and th_ origin of P. biaurelia haplotyp_s id_ntifi_d in curr_nt study. Black dash_s on particular branch_s r_pr_s_nt nucl_otid_ substitutions b_tw__n particular haplotyp_s. Analys_s w_r_ conduct_d using th_ M_dian Joining m_thod in PopART softwar_ v. 1.7

Fig. 2 in Worldwide sampling reveals low genetic variability in populations of the freshwater ciliate Paramecium biaurelia (P. aurelia species complex, Ciliophora, Protozoa)

Fig. 2 Phylog_n_tic tr__ construct_d for Paramecium aurelia compl_x, P. jenningsi compl_x and P. schewiakoffi (two sp_ci_s: P. caudatum and P. multimicronucleatum w_r_ us_d as an outgroup). Th_ tr__ was construct_d on th_ basis of a comparison of s_qu_nc_s from th_ ribosomal ITS1-5.8S-ITS2-5'LSU fragm_nt using th_ maximum lik_lihood m_thod. Bootstrap valu_s for n_ighbor joining, maximum parsimony, maximum lik_lihood, and post_rior probabiliti_s for

Fig. 3 in Worldwide sampling reveals low genetic variability in populations of the freshwater ciliate Paramecium biaurelia (P. aurelia species complex, Ciliophora, Protozoa)

Fig. 3 Phylog_n_tic tr__ construct_d for Paramecium aurelia compl_x, P. jenningsi compl_x and P. schewiakoffi (two sp_ci_s: P. caudatum and P. multimicronucleatum w_r_ us_d as an outgroup). Th_ tr__ was construct_d on th_ basis of a comparison of s_qu_nc_s from th_ mitochondrial COI fragm_nt using th_ maximum lik_lihood m_thod. Bootstrap valu_s for n_ighbor joining, maximum parsimony, maximum lik_lihood, and post_rior probabiliti_s for Bay_sian inf_r_nc_ ar_

opencc-by-4.0Apr 2023View details →

Inter-generational nuclear crosstalk links the control of gene expression to programmed genome rearrangements during the Paramecium sexual cycle

Multinucleate cells are found in many eukaryotes, but how multiple nuclei coordinate their functions is still poorly understood. In the cytoplasm of the ciliate Paramecium tetraurelia, two micronuclei (MIC) serving sexual reproduction coexist with a somatic macronucleus (MAC) dedicated to gene expression. During sexual processes, the MAC is progressively destroyed while still ensuring transcription and new MACs develop from copies of the zygotic MIC. Several gene clusters are successively induced and switched off before vegetative growth resumes. Concomitantly, programmed genome rearrangements (PGR) remove transposons and their relics from the new MACs. Development of the new MACs is controlled by the old MAC, since the latter expresses genes involved in PGR, including the PGM gene encoding the essential PiggyMac endonuclease that cleaves the ends of eliminated sequences. Using RNA deep sequencing and transcriptome analysis, we show that impairing PGR up-deregulates key PGR genes, together with ~600 other genes possibly also involved in PGR. Among these genes, 42% are no longer induced when no new MACs are formed, including 180 genes that are co-expressed with PGM under all tested conditions. We propose that bi-directional crosstalk between the two coexisting generations of MACs links gene expression to the progression of MAC development.

opencc-by-4.0Oct 2023View details →

Uncoupling of programmed DNA cleavage and repair jeopardizes the assembly of the Paramecium somatic genome

In the ciliate Paramecium, the precise excision of numerous Internal Eliminated Sequences (IESs) from the somatic genome is essential at each sexual cycle. DNA double strands breaks (DSBs) are introduced by the PiggyMac endonuclease, and repaired in a highly concerted manner by the Non-Homologous End Joining pathway (NHEJ), as illustrated by the complete inhibition of DNA cleavage when Ku70/80 proteins are missing. We show here that expression of a DNA binding-deficient Ku70 mutant (Ku70-6E) permits DNA cleavage but not DSB repair, leading to accumulation of unrepaired DSBs. When wildtype and mutant Ku are co-expressed, the DSBs induced by Ku70-6E can be repaired by wildtype Ku, which uncouples DNA repair from the cleavage step. High-throughput sequencing of the developing MAC genome in these conditions reveals the presence of extremities healed by de novo telomere addition and numerous translocations between IES-flanking sequences. We conclude that coupling the two steps of IES excision ensures that both extremities are maintained together throughout the process, and propose that Ku assists PiggyMac during assembly of the synaptic pre-cleavage complex.

opencc-by-4.0Feb 2022View details →

Electrophysiological recordings of Paramecium with particle image velocimetry

These files contain electrophysiological data as well as simultaneous video recordings and some analyses for the following paper: An electrophysiological and kinematic model of Paramecium, the “swimming neuron” Irene Elices, Anirudh Kulkarni, Nicolas Escoubet, Léa-Laetitia Pontani, Alexis Prevost, Romain Brette

Fig. 1 in Worldwide sampling reveals low genetic variability in populations of the freshwater ciliate Paramecium biaurelia (P. aurelia species complex, Ciliophora, Protozoa)

Fig. 1 Th_ origin (N = 92) of Paramecium biaurelia strains us_d in pr_s_nt studi_s

opencc-by-4.0Oct 2024View details →

The PIWI-interacting protein Gtsf1 controls the selective degradation of small RNAs in Paramecium

Ciliates undergo developmentally programmed genome elimination, in which small RNAs direct the removal of transposable elements during the development of the somatic nucleus. 25-nt scnRNAs are produced from the entire germline genome and transported to the maternal somatic nucleus, where selection of scnRNAs corresponding to germline-specific sequences is thought to take place. Selected scnRNAs then guide the elimination of transposable elements in the developing somatic nucleus. How germline-specific scnRNAs are selected remains to be determined. Here, we provide important mechanistic insights into the scnRNA selection pathway by identifying a Paramecium homolog of Gtsf1 as essential for the selective degradation of scnRNAs corresponding to retained somatic sequences. Consistently, we also show that Gtsf1 is localized in the maternal somatic nucleus where it associates with the scnRNA-binding protein Ptiwi09. Furthermore, we demonstrate that the scnRNA selection process is critical for genome elimination. We propose that Gtsf1 is required for the coordinated degradation of Ptiwi09-scnRNA complexes that pair with target RNA via the ubiquitin pathway, similarly to the mechanism suggested for microRNA target-directed degradation in metazoans.

dryad36/100

Resource gradients create energy trade-offs in the inducible defense response of Paramecium aurelia

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publicNov 2025View details →

opencc-by-4.0Sep 2023View details →

A histone methyltransferase-independent function of PRC2 controls small RNA dynamics during programmed DNA elimination in Paramecium

To limit transposable element (TE) mobilization, most eukaryotes have evolved small RNAs to silence TE activity via homology-dependent mechanisms. Small RNAs, 20-30 nucleotides in length, bind to PIWI proteins and guide them to nascent transcripts by sequence complementarity, triggering the recruitment of histone methyltransferase enzymes on chromatin to repress the transcriptional activity of TEs and other repeats. In the ciliate Paramecium tetraurelia, 25-nt scnRNAs corresponding to TEs recruit Polycomb Repressive Complex 2 (PRC2), and trigger their elimination during the formation of the somatic nucleus. Here, we sequenced sRNAs during the entire sexual cycle with unprecedented resolution. Our data confirmed that scnRNAs are produced from the entire germline genome, from TEs and non-TE sequences, during meiosis. Non-TE scnRNAs are selectively degraded, which results in the specific selection of TE-scnRNAs. We demonstrate that PRC2 is essential for the selective degradation of non-TE-scnRNAs, independently of its histone methyltransferase activity. We further show that the PRC2 cofactor Rf4 mediates the physical interaction between the scnRNA-binding protein Ptiwi09 and the zinc finger protein Gtsf1, pointing to an architectural role of PRC2 in scnRNA degradation.

opencc-by-4.0Jan 2019View details →

Population genetics of Paramecium mitochondrial genomes; genome assemblies and annotation files

Because of issues arising during submission to GenBank of Paramecium mitochondrial genomes due to the highly unconventional nature of the genetic code used in these genomes, we are initially making the genomes publicly available here (while we are still working on a submission to official databases).

Fig. 38 in New Paramecium (Ciliophora, Oligohymenophorea) congeners shape our view on its biodiversity

Fig. 38 Schematical images of major Parameciom morphospecies made according to its morphometric data (reprint of Fig. 1 from Fokin 2010/11). a P. moltimicronocleatom, b P. caodatom, c P. jenningsi, d P. schewiakoffi, e P. boetschlii sp. nov., f P. woodroffi, g P. aorelia, h

Fig. 35 in New Paramecium (Ciliophora, Oligohymenophorea) congeners shape our view on its biodiversity

Fig. 35 Dendrogram (a) for hierarchical clustering (UPGMA) and topogram (b) for non-metric multidimensional scaling (MDS) of morphometric and morphobiological characteristics of 16 Parameciom species. AU P. aorelia, BU P. borsaria, CA P. caodatom, CL P. calkinsi, DU P. doboscqoi, JE P. jenningsi, MU P. moltimicronocleatom, NR P. nephridiatom, PB BEocandidatos P. brazilianum^, PG BEocandidatos P. germanicum^, PH BEocandidatos P. hungarianum^, PN P. boetschlii sp. nov., PO P. polycaryom, PU P. potrinom, SH P. schewiakoffi, WO P. woodroffi

Fig. 27–29 in New Paramecium (Ciliophora, Oligohymenophorea) congeners shape our view on its biodiversity

Fig. 27–29 Morphology of BEocandidatos P. brazilianum^. 27 Ventral view of silver nitrate-impregnated cell. 28 General view of the Feulgenstained cell with macronucleus (MA) fragmentation and two surrounding micronuclei (indicated by arrows). 29 Nuclear apparatus of the cell with fragmented MA and surrounding micronuclei (arrows) in higher magnification. Bars 30 μm (27, 28), 15 μm (29)

Fig. 30–34 in New Paramecium (Ciliophora, Oligohymenophorea) congeners shape our view on its biodiversity

Fig. 30–34 Morphology of BEocandidatos P. hungarianum^. 30 Living ciliate with visible contractile vacuoles (CV) and macronucleus (MA). DIC contrast. 31–32 Ventral (31) and dorsal (32) views with indications for cytoproct (C) and pores of the contractile vacuoles (PCV). Silver nitrate impregnation. 33 Nuclear apparatus of the ciliate with MA and surrounding micronuclei (indicated by arrows). Feulgen-stained cell. 34 Nuclear apparatus with MA and MI (arrows). Large magnification. Bars 15 μm (30), 13 μm (31, 32), 20 μm (33), 10 μm (34)