Inteins are self-splicing protein elements found in viruses and all three domains of life. How the DNA encoding these selfish elements spreads within and between genomes is poorly understood, particularly in eukaryotes where inteins are scarce. Here, we show that the nuclear genomes of three strains of Anaeramoeba encode between 45 and 103 inteins, in stark contrast to four found in the most intein-rich eukaryotic genome described previously. The Anaeramoeba inteins reside in a wide range of proteins, only some of which correspond to intein-containing proteins in other eukaryotes, prokaryotes, and viruses. Our data also suggest that viruses have contributed to the spread of inteins in Anaeramoeba and the colonization of new alleles. The persistence of Anaeramoeba inteins might be partly explained by intragenomic movement of intein-encoding regions from gene to gene. Our intein dataset greatly expands the spectrum of intein-containing proteins and provides insights into the evolution of inteins in eukaryotes.

Department of Biochemistry and Molecular Biology Dalhousie University Halifax Nova Scotia B3H 4R2 Canada

Department of Zoology Charles University Prague 128 00 Czech Republic

Institute for Comparative Genomics Dalhousie University Halifax Nova Scotia B3H 4R2 Canada

Microbiology and Immunology Department of Cell and Molecular Biology Biomedical Centre Uppsala University Uppsala 751 24 Sweden

Microbiology Group Department of Biology Lund University Lund 223 62 Sweden

Zobrazit více v PubMed

Lennon C. W., Belfort M., Inteins. Curr. Biol. 27, R204–R206 (2017). PubMed

Perler F. B., et al. , Protein splicing elements: Inteins and exteins–a definition of terms and recommended nomenclature. Nucleic Acids Res. 22, 1125–1127 (1994). PubMed PMC

Liu X. Q., Protein-splicing intein: Genetic mobility, origin, and evolution. Annu. Rev. Genet. 34, 61–76 (2000). PubMed

Belfort M., Reaban M. E., Coetzee T., Dalgaard J. Z., Prokaryotic introns and inteins: A panoply of form and function. J. Bacteriol. 177, 3897–3903 (1995). PubMed PMC

Gimble F. S., Thorner J., Homing of a DNA endonuclease gene by meiotic gene conversion in Saccharomyces cerevisiae. Nature 357, 301–306 (1992). PubMed

Gogarten J. P., Hilario E., Inteins, introns, and homing endonucleases: Recent revelations about the life cycle of parasitic genetic elements. BMC Evol. Biol. 6, 94 (2006). PubMed PMC

Soucy S. M., Fullmer M. S., Papke R. T., Gogarten J. P., Inteins as indicators of gene flow in the halobacteria. Front. Microbiol. 5, 299 (2014). PubMed PMC

Swithers K. S., Senejani A. G., Fournier G. P., Gogarten J. P., Conservation of intron and intein insertion sites: Implications for life histories of parasitic genetic elements. BMC Evol. Biol. 9, 303 (2009). PubMed PMC

Perler F. B., Olsen G. J., Adam E., Compilation and analysis of intein sequences. Nucleic Acids Res. 25, 1087–1093 (1997). PubMed PMC

Hirata R., et al. , Molecular structure of a gene, VMA1, encoding the catalytic subunit of H(+)-translocating adenosine triphosphatase from vacuolar membranes of Saccharomyces cerevisiae. J. Biol. Chem. 265, 6726–6733 (1990). PubMed

Kane P. M., et al. , Protein splicing converts the yeast TFP1 gene product to the 69-kD subunit of the vacuolar H(+)-adenosine triphosphatase. Science 250, 651–657 (1990). PubMed

Green C. M., Novikova O., Belfort M., The dynamic intein landscape of eukaryotes. Mob. DNA 9, 4 (2018). PubMed PMC

Novikova O., et al. , Intein clustering suggests functional importance in different domains of life. Mol. Biol. Evol. 33, 783–799 (2016). PubMed PMC

Novikova O., Topilina N., Belfort M., Enigmatic distribution, evolution, and function of inteins. J. Biol. Chem. 289, 14490–14497 (2014). PubMed PMC

Bernstein C., Deoxyribonucleic acid repair in bacteriophage. Microbiol. Rev. 45, 72–98 (1981). PubMed PMC

Lopes A., Amarir-Bouhram J., Faure G., Petit M.-A., Guerois R., Detection of novel recombinases in bacteriophage genomes unveils Rad52, Rad51 and Gp2.5 remote homologs. Nucleic Acids Res. 38, 3952–3962 (2010). PubMed PMC

Weigel C., Seitz H., Bacteriophage replication modules. FEMS Microbiol. Rev. 30, 321–381 (2006). PubMed

Gogarten J. P., Senejani A. G., Zhaxybayeva O., Olendzenski L., Hilario E., Inteins: Structure, function, and evolution. Annu. Rev. Microbiol. 56, 263–287 (2002). PubMed

Pavankumar T. L., Inteins: Localized distribution, gene regulation, and protein engineering for biological applications. Microorganisms 6, 19 (2018). PubMed PMC

Lennon C. W., Stanger M., Banavali N. K., Belfort M., Conditional protein splicing switch in hyperthermophiles through an intein-extein partnership. mBio 9, e02304–17 (2018). PubMed PMC

Stairs C. W., et al. , Anaeramoebae are a divergent lineage of eukaryotes that shed light on the transition from anaerobic mitochondria to hydrogenosomes. Curr. Biol. 31, 5605–5612.e5 (2021). PubMed

Táborský P., Pánek T., Čepička I., Anaeramoebidae fam. nov., a novel lineage of anaerobic amoebae and amoeboflagellates of uncertain phylogenetic positio n. Protist 168, 495–526 (2017). PubMed

Jerlström-Hultqvist J., et al. , A novel symbiosome in an anaerobic single-celled eukaryote. bioRxiv [Preprint] (2023). 10.1101/2023.03.03.530753 (Accessed 5 March 2023). DOI

Perler F. B., InBase: The intein database. Nucleic Acids Res. 30, 383–384 (2002). PubMed PMC

Green C. M., et al. , Spliceosomal Prp8 intein at the crossroads of protein and RNA splicing. PLoS Biol. 17, e3000104 (2019). PubMed PMC

Liu X.-Q., Yang J., Meng Q., Four inteins and three group II introns encoded in a bacterial ribonucleotide reductase gene. J. Biol. Chem. 278, 46826–46831 (2003). PubMed

Monier A., Sudek S., Fast N. M., Worden A. Z., Gene invasion in distant eukaryotic lineages: Discovery of mutually exclusive genetic elements reveals marine biodiversity. ISME J. 7, 1764–1774 (2013). PubMed PMC

Kelley D. S., Lennon C. W., Sea-Phages M., Belfort O. Novikova., Mycobacteriophages as incubators for intein dissemination and evolution. mBio 7, e01537-16 (2016). PubMed PMC

Tori K., Perler F. B., Expanding the definition of class 3 inteins and their proposed phage origin▿. J. Bacteriol. 193, 2035–2041 (2011). PubMed PMC

Gallot-Lavallee L., Blanc G., Claverie J.-M., Comparative genomics of Chrysochromulina Ericina Virus (CeV) and other microalgae-infecting large DNA viruses highlight their intricate evolutionary relationship with the established Mimiviridae family. J. Virol. 91, e00230-17 (2017), 10.1128/JVI.00230-17. PubMed DOI PMC

Southworth M. W., Benner J., Perler F. B., An alternative protein splicing mechanism for inteins lacking an N-terminal nucleophile. EMBO J. 19, 5019–5026 (2000). PubMed PMC

Tori K., et al. , Splicing of the mycobacteriophage Bethlehem DnaB intein: Identification of a new mechanistic class of inteins that contain an obligate block F nucleophile. J. Biol. Chem. 285, 2515–2526 (2010). PubMed PMC

Featherston J., et al. , The 4-Celled Tetrabaena socialis nuclear genome reveals the essential components for genetic control of cell number at the origin of multicellularity in the volvocine lineage. Mol. Biol. Evol. 35, 855–870 (2018). PubMed

Filée J., Multiple occurrences of giant virus core genes acquired by eukaryotic genomes: The visible part of the iceberg? Virology 466–467, 53–59 (2014). PubMed

Maumus F., Blanc G., Study of gene trafficking between acanthamoeba and giant viruses suggests an undiscovered family of amoeba-infecting viruses. Genome Biol. Evol. 8, 3351–3363 (2016). PubMed PMC

Maumus F., Epert A., Nogué F., Blanc G., Plant genomes enclose footprints of past infections by giant virus relatives. Nat. Commun. 5, 4268 (2014). PubMed PMC

Gallot-Lavallée L., Blanc G., A glimpse of nucleo-cytoplasmic large DNA virus biodiversity through the eukaryotic genomics window. Viruses 9, 17 (2017). PubMed PMC

Moniruzzaman M., Weinheimer A. R., Martinez-Gutierrez C. A., Aylward F. O., Widespread endogenization of giant viruses shapes genomes of green algae. Nature 588, 141–145 (2020). PubMed

Elleuche S., Pelikan C., Nolting N., Pöggeler S., Inteins and introns within the prp8 -gene of four Eupenicillium species. J. Basic Microbiol. 49, 52–57 (2009). PubMed

Edgar R. C., MUSCLE: Multiple sequence alignment with high accuracy and high throughput. Nucleic Acids Res. 32, 1792–1797 (2004). PubMed PMC

Goddard M. R., Burt A., Recurrent invasion and extinction of a selfish gene. Proc. Natl. Acad. Sci. U.S.A. 96, 13880–13885 (1999). PubMed PMC

Belfort M., Roberts R. J., Homing endonucleases: Keeping the house in order. Nucleic Acids Res. 25, 3379–3388 (1997). PubMed PMC

Liu X. Q., Hu Z., A DnaB intein in Rhodothermus marinus: Indication of recent intein homing across remotely related organisms. Proc. Natl. Acad. Sci. U.S.A. 94, 7851–7856 (1997). PubMed PMC

Swithers K. S., Soucy S. M., Lasek-Nesselquist E., Lapierre P., Gogarten J. P., Distribution and evolution of the mobile vma-1b intein. Mol. Biol. Evol. 30, 2676–2687 (2013). PubMed

Fischer M. G., Kelly I., Foster L. J., Suttle C. A., The virion of Cafeteria roenbergensis virus (CroV) contains a complex suite of proteins for transcription and DNA repair. Virology 466–467, 82–94 (2014). PubMed

Salas-Leiva D. E., et al. , Genomic analysis finds no evidence of canonical eukaryotic DNA processing complexes in a free-living protist. Nat. Commun. 12, 6003 (2021). PubMed PMC

Tornabene P., et al. , Intein-mediated protein trans-splicing expands adeno-associated virus transfer capacity in the retina. Sci. Transl. Med. 11, eaav4523 (2019). PubMed PMC

López-Igual R., Bernal-Bayard J., Rodríguez-Patón A., Ghigo J.-M., Mazel D., Engineered toxin-intein antimicrobials can selectively target and kill antibiotic-resistant bacteria in mixed populations. Nat. Biotechnol. 37, 755–760 (2019). PubMed

Sarmiento C., Camarero J. A., Biotechnological applications of protein splicing. Curr. Protein Pept. Sci. 20, 408–424 (2019). PubMed PMC

Altschul S. F., Gish W., Miller W., Myers E. W., Lipman D. J., Basic local alignment search tool. J. Mol. Biol. 215, 403–410 (1990). PubMed

Eddy S. R., Multiple alignment using hidden Markov models. Proc. Int. Conf. Intell. Syst. Mol. Biol. 3, 114–120 (1995). PubMed

Katoh K., Standley D. M., MAFFT multiple sequence alignment software version 7: Improvements in performance and usability. Mol. Biol. Evol. 30, 772–780 (2013). PubMed PMC

Criscuolo A., Gribaldo S., BMGE (Block Mapping and Gathering with Entropy): A new software for selection of phylogenetic informative regions from multiple sequence alignments. BMC Evol. Biol. 10, 210 (2010). PubMed PMC

Nguyen L.-T., Schmidt H. A., von Haeseler A., Minh B. Q., IQ-TREE: A fast and effective stochastic algorithm for estimating maximum-likelihood phylogenies. Mol. Biol. Evol. 32, 268–274 (2015). PubMed PMC

Hoang D. T., Chernomor O., von Haeseler A., Minh B. Q., Vinh L. S., UFboot2: Improving the ultrafast bootstrap approximation. Mol. Biol. Evol. 35, 518–522 (2018). PubMed PMC

Zallot R., Oberg N., Gerlt J. A., The EFI web resource for genomic enzymology tools: Leveraging protein, genome, and metagenome databases to discover novel enzymes and metabolic pathways. Biochemistry 58, 4169–4182 (2019). PubMed PMC

Shannon P., et al. , Cytoscape: A software environment for integrated models of biomolecular interaction networks. Genome Res. 13, 2498–2504 (2003). PubMed PMC

Mi H., et al. , Protocol update for large-scale genome and gene function analysis with the PANTHER classification system (v.14.0). Nat. Protoc. 14, 703–721 (2019). PubMed PMC