TY - JOUR
T1 - A genome-wide genetic screen identifies a novel kDNA replication protein in trypanosomes
AU - Miskinyte, Migla
AU - Jetishi, Clirim
AU - Kalichava, Ana
AU - Ivens, Alasdair
AU - Waterfall, Martin
AU - Gould, Matthew K.
AU - Glover, Lucy
AU - Horn, David
AU - Ochsenreiter, Torsten
AU - Schnaufer, Achim
N1 - Publisher Copyright:
© The Author(s) 2026. Published by Oxford University Press. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (https://creativecommons.org/licenses/by/4.0/), which permits unrestricted reuse, distribution, and reproduction in any medium, provided the original work is properly cited.
PY - 2026/6/10
Y1 - 2026/6/10
N2 - Mitochondrial DNA of trypanosomatid parasites is organized into a topologically complex structure, named kinetoplast [kinetoplast DNA (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 further experimental characterization. Ultrastructure expansion microscopy using a tagged version of the protein reveals a dynamic localization during the cell cycle. RNA interference-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.
AB - Mitochondrial DNA of trypanosomatid parasites is organized into a topologically complex structure, named kinetoplast [kinetoplast DNA (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 further experimental characterization. Ultrastructure expansion microscopy using a tagged version of the protein reveals a dynamic localization during the cell cycle. RNA interference-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.
UR - https://www.scopus.com/pages/publications/105039434211
U2 - 10.1093/nar/gkag493
DO - 10.1093/nar/gkag493
M3 - Article
C2 - 42165136
AN - SCOPUS:105039434211
SN - 0305-1048
VL - 54
JO - Nucleic Acids Research
JF - Nucleic Acids Research
IS - 10
M1 - gkag493
ER -