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PreprintCited by 0Trypanosoma species research and implicationsMitochondrial Function and PathologyChromosomal and Genetic Variations

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A Genome-Wide Genetic Screen Identifies a Novel kDNA Replication Protein in Trypanosomes

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Miskinyte, Migla, Jetishi, Clirim, Kalichava, Ana, Ivens, Alasdair, Waterfall, Martin, Gould, Matt, Glover, Lucy, Horn, David, Ochsenreiter, Torsten, Schnaufer, Achim·2026·Nucleic Acids Research
DOI 10.64898/2026.01.05.697628PMC PMC13191291

Preprint — published as 10.1093/nar/gkag493

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Abstract

Mitochondrial DNA of trypanosomatid parasites is organized into a topologically complex structure, named kinetoplast (kDNA). Replication, segregation and expression of kDNA involve an estimated ∼300 proteins, only a fraction of which have been identified and characterized. Here, we report the development of a genetic screen in Trypanosoma brucei to identify novel kDNA maintenance factors. Of the 20 highest-ranked genes identified, six are known kDNA maintenance factors. We selected one hit, Tb927.8.4240 , a gene of previously unknown function, for experimental follow-up. Ultrastructure expansion microscopy using a tagged version of the protein reveals a dynamic localization during the cell cycle. RNAi-mediated ablation of Tb927.8.4240 results in the progressive but incomplete loss of kDNA, with only a minor effect on the tripartite attachment complex, suggesting the protein is involved in kDNA replication but not segregation. The growth phenotype of Tb927.8.4240 ablation is fully rescued in a kDNA-independent genetic background, confirming a specific role in kDNA replication. In summary, we describe a functional genetic screen for the identification of kDNA maintenance factors in trypanosomes, validate one hit as a novel kDNA replication factor, and provide a prioritized hit list as a promising starting point for the future identification of additional factors.

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Research works printed but not linked: 99

  1. 10.1172/JCI33945 · DOI
  2. 10.1146/annurev-micro-092611-150057 · DOI
  3. 2020, Mitochondrial DNAs provide insight into trypanosome phylogeny and molecular evolution, BMC Evolutionary Biology, 20, 161, 10.1186/s12862-020-01701-9 · DOI
  4. 10.1093/nar/25.19.3751 · DOI
  5. 10.1016/S0020-7519(02)00020-6 · DOI
  6. 10.1007/s00294-005-0027-0 · DOI
  7. 10.1091/mbc.E02-08-0525 · DOI
  8. 10.1074/jbc.X114.620666 · DOI
  9. 2023, Single-Molecule Structure and Topology of Kinetoplast DNA Networks, Phys. Rev. X, 13, 021010
  10. 10.1016/S0020-7519(01)00296-X · DOI
  11. 10.1016/j.pt.2020.01.006 · DOI
  12. 10.1093/nar/gkz928 · DOI
  13. 10.1128/AAC.00122-14 · DOI
  14. 10.1371/journal.ppat.1001226 · DOI
  15. 10.1242/jcs.221820 · DOI
  16. 10.1093/femsre/fuac047 · DOI
  17. 10.1083/jcb.153.4.735 · DOI
  18. 10.1111/j.1365-2958.2010.07061.x · DOI
  19. 10.1073/pnas.0706858104 · DOI
  20. 10.1093/emboj/20.17.4674 · DOI
  21. 2018, Characterization of the novel mitochondrial genome replication factor MiRF172 in Trypanosoma brucei, Journal of cell science, 131, pii: jcs211730, 10.1242/jcs.211730 · DOI
  22. 10.1128/EC.00308-10 · DOI
  23. 10.1016/j.molcel.2009.07.004 · DOI
  24. 2009, TbPIF5 is a Trypanosoma brucei mitochondrial DNA helicase involved in processing of minicircle Okazaki fragments, PLoS.Pathog, 5, e1000589, 10.1371/journal.ppat.1000589 · DOI
  25. 10.1074/jbc.M308565200 · DOI
  26. 10.1128/EC.4.4.765-774.2005 · DOI
  27. Amodeo, S. , Bregy, I. and Ochsenreiter, T . (2023) Mitochondrial genome maintenance-the kinetoplast story. FEMS Microbiol Rev, 47, fuac047. · DOI
  28. 10.1128/MCB.15.12.6794 · DOI
  29. 10.1016/S1097-2765(02)00571-3 · DOI
  30. 10.1128/EC.00199-08 · DOI
  31. 10.1073/pnas.2204294119 · DOI
  32. 2025, Connecting basal body and mitochondrial DNA: TAC53 and the tubular organization of the tripartite attachment complex, PLoS Pathog, 21, e1013521, 10.1371/journal.ppat.1013521 · DOI
  33. 10.1371/journal.ppat.1010207 · DOI
  34. 2020, A comparison of three approaches for the discovery of novel tripartite attachment complex proteins in Trypanosoma brucei, PLOS Neglected Tropical Diseases, 14, e0008568, 10.1371/journal.pntd.0008568 · DOI
  35. 10.1371/journal.ppat.1011486 · DOI
  36. 10.1016/j.celrep.2023.113083 · DOI
  37. 10.1016/j.pt.2021.09.002 · DOI
  38. 2015, Genome-scale RNAi screens for high-throughput phenotyping in bloodstream-form African trypanosomes, Nature protocols, 10, 106, 10.1038/nprot.2015.005 · DOI
  39. 10.2307/3282883 · DOI
  40. 10.1371/journal.ppat.1007195 · DOI
  41. 10.1128/AAC.00044-13 · DOI
  42. 2012, Fast gapped-read alignment with Bowtie 2, Nature Methods, 9, 357, 10.1038/nmeth.1923 · DOI · Paper page
  43. 10.1093/bioinformatics/btp352 · DOI · Paper page
  44. Quinlan, A.R . (2014) BEDTools: The Swiss-Army Tool for Genome Feature Analysis. Curr Protoc Bioinformatics, 47, 11.12.1-34. · DOI
  45. 10.1093/bioinformatics/btp616 · DOI · Paper page
  46. 10.1074/jbc.M412336200 · DOI
  47. 10.1016/j.molbiopara.2007.02.008 · DOI
  48. 10.1016/S0166-6851(99)00002-X · DOI
  49. 10.1006/meth.2001.1262 · DOI · Paper page
  50. 10.1016/j.molbiopara.2010.03.007 · DOI
  51. 2017, Quantitating the cell: turning images into numbers with ImageJ, Wiley Interdisciplinary Reviews: Developmental Biology, 6, e260, 10.1002/wdev.260 · DOI
  52. 10.1371/journal.ppat.1005586 · DOI
  53. 10.1073/pnas.1305404110 · DOI
  54. 10.1093/emboj/cdf474 · DOI
  55. 10.1101/gr.115089.110 · DOI
  56. 10.1093/emboj/cdf482 · DOI
  57. 10.1016/j.yexcr.2004.10.013 · DOI
  58. 2021, Characterization of the novel mitochondrial genome segregation factor TAP110 in Trypanosoma brucei, Journal of cell science, 134, jcs.254300, 10.1242/jcs.254300 · DOI
  59. 10.1093/nar/gkad1003 · DOI
  60. 2023, Genome-wide subcellular protein map for the flagellate parasite Trypanosoma brucei, Nat Microbiol, 8, 533, 10.1038/s41564-022-01295-6 · DOI
  61. 10.1371/journal.ppat.1006679 · DOI
  62. 10.1002/pmic.200800477 · DOI
  63. 2017, Charting organellar importomes by quantitative mass spectrometry, Nature Communications, 8, 15272, 10.1038/ncomms15272 · DOI
  64. 10.1371/journal.ppat.1006808 · DOI
  65. 10.1073/pnas.1605497113 · DOI
  66. 10.1038/sj.emboj.7601956 · DOI
  67. 10.1371/journal.pone.0234918 · DOI
  68. 10.1007/s13258-024-01506-w · DOI
  69. 10.1371/journal.ppat.1008129 · DOI
  70. 10.1016/j.molbiopara.2009.11.009 · DOI
  71. 10.1128/EC.00269-12 · DOI
  72. 10.1016/j.nbd.2018.09.020 · DOI
  73. 10.1371/journal.pntd.0002121 · DOI
  74. 2018, Branched late-steps of the cytosolic iron-sulphur cluster assembly machinery of Trypanosoma brucei, PLoS Pathog, 14, e1007326, 10.1371/journal.ppat.1007326 · DOI
  75. 10.1093/nar/gkae1082 · DOI
  76. 10.1371/journal.pntd.0011058 · DOI
  77. 10.1111/j.1365-2958.2007.05749.x · DOI
  78. 10.1101/2024.04.22.590512 · DOI
  79. 10.1091/mbc.E02-05-0266 · DOI
  80. 10.1128/EC.05008-11 · DOI
  81. 10.1016/S0166-6851(02)00018-X · DOI
  82. 10.1111/j.1365-2958.2007.05686.x · DOI
  83. 10.1111/j.1365-2958.2011.07938.x · DOI
  84. 10.1128/MCB.01176-10 · DOI
  85. 10.1371/journal.ppat.1006310 · DOI
  86. 2024, A novel nabelschnur protein regulates segregation of the kinetoplast DNA in Trypanosoma brucei, Curr Biol, 34, 4803, 10.1016/j.cub.2024.08.044 · DOI
  87. 10.1128/EC.05291-11 · DOI
  88. 10.1091/mbc.E18-02-0127 · DOI
  89. 10.1128/EC.5.1.54-61.2006 · DOI
  90. 10.1038/s41580-023-00673-0 · DOI
  91. 10.1128/EC.1.4.495-502.2002 · DOI
  92. 10.1017/S1355838200992021 · DOI
  93. 1991, Why kinetoplast DNA networks?, Trends in genetics : TIG, 7, 139, 10.1016/0168-9525(91)90091-4 · DOI
  94. 2023, Phylogenetic framework to explore trait evolution in Trypanosomatidae, Trends Parasitol, 40, 96
  95. 10.1074/jbc.M202347200 · DOI
  96. 2009, Prohibitin function within mitochondria: Essential roles for cell proliferation and cristae morphogenesis, Biochimica et Biophysica Acta (BBA) - Molecular Cell Research, 1793, 27, 10.1016/j.bbamcr.2008.05.013 · DOI
  97. 10.1093/nar/gkh340 · DOI · Paper page
  98. Waterhouse, A.M., Procter, J.B., Martin, D.M.A., Clamp, M. and Barton, G.J. (2009) Jalview Version 2--a multiple sequence alignment editor and analysis workbench. Bioinformatics, 25, 1189–1191. · DOI
  99. 10.1093/bioinformatics/9.6.745 · DOI

Metadata

Authors
Miskinyte, M., Jetishi, C., Kalichava, A., Ivens, A., Waterfall, M., Gould, M., Glover, L., Horn, D., Ochsenreiter, T., & Schnaufer, A.
Affiliations
  • University of Edinburgh (GB)
  • University of Bern (CH)
  • Institute of Immunology and Infection Research, University of Edinburgh, UK
  • University of Dundee (GB)
Corresponding author
Schnaufer, Achim
ORCID
Venue
Nucleic Acids Research
Publisher
openRxiv
Published
2026-01-05
Volume
54
Issue
10
Pages
gkag493
ISSN
0305-1048 (ISSN-L)
DOI
10.64898/2026.01.05.697628
PMC
PMC13191291
OpenAlex
W7161938064
Cited by
0
References
1
FWCI
0.00
Type
journal article
Language
en
Open access
Licence
Locations
  • Nucleic Acids Research (publishedVersion, cc-by)
  • bioRxiv (Cold Spring Harbor Laboratory) (acceptedVersion, cc-by)
  • PubMed (publishedVersion)
  • Discovery Research Portal (University of Dundee) (publishedVersion, cc-by)
  • PubMed Central (publishedVersion, cc-by)
  • HAL (submittedVersion, cc-by)
  • Edinburgh Research Explorer (University of Edinburgh) (submittedVersion, cc-by)
  • Open Access CRIS of the University of Bern
Funders
  • Medical Research Council (MR/L019701/1)
  • Swiss National Science Foundation (207525)
  • Wellcome Trust (217105/Z/19/Z)
Topics
  • Health Sciences › Medicine › Epidemiology › Trypanosoma species research and implications
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Mitochondrial Function and Pathology
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Genetics › Chromosomal and Genetic Variations
Keywords
kinetoplast DNA, Trypanosoma brucei, genome-wide screen, ultrastructure expansion microscopy, RNA interference, mitochondrial DNA segregation
MeSH terms
DNA Replication, Genetic Testing, Trypanosoma brucei brucei, Protozoan Proteins, DNA, Kinetoplast, Genome, Protozoan, RNA Interference

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