Characterization of the novel mitochondrial genome replication factor MiRF172 inTrypanosoma brucei

Simona Amodeo; Martin Jakob; Torsten Ochsenreiter

doi:10.1242/jcs.211730

Characterization of the Novel Mitochondrial Genome Replication Factor MiRF172 in Trypanosoma brucei

10.1101/194811 ◽

2017 ◽

Author(s):

Simona Amodeo ◽

Martin Jakob ◽

Torsten Ochsenreiter

Keyword(s):

Mitochondrial Genome ◽

Trypanosoma Brucei ◽

Super Resolution ◽

Genome Replication ◽

The Novel ◽

Replication Factor ◽

Summary Statement ◽

Super Resolution Microscopy ◽

Novel Protein

AbstractThe unicellular parasite Trypanosoma brucei harbors one individual mitochondrial organelle with a singular genome the kinetoplast DNA or kDNA. The kDNA largely consists of concatenated minicircles and a few maxicircles that are also interlocked into the kDNA disc. More than 30 proteins involved in kDNA replication have been described, however several mechanistic questions are only poorly understood. Here, we describe and characterize MiRF172, a novel mitochondrial genome replication factor, which is essential for proper cell growth and kDNA maintenance. Using super-resolution microscopy, we localize MiRF172 to the antipodal sites of the kDNA. We demonstrate that depletion of MiRF172 leads to continuous loss of mini- and maxicircles during the cell division cycle. Detailed analysis suggests that MiRF172 is likely involved in the reattachment of replicated minicircles to the kDNA disc. Furthermore, we provide evidence that the localization of the replication factor MiRF172 not only depends on the kDNA itself, but also on the mitochondrial genome segregation machinery suggesting a tight interaction between the two essential entities.Summary StatementMiRF172 is a novel protein involved in the reattachment of replicated minicircles in Trypanosoma brucei, which requires the mitochondrial segregation machinery for proper localization.

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Characterization of the Novel Mitochondrial Genome Segregation Factor TAP110 in Trypanosoma brucei

Journal of Cell Science ◽

10.1242/jcs.254300 ◽

2021 ◽

pp. jcs.254300

Author(s):

Simona Amodeo ◽

Ana Kalichava ◽

Albert Fradera-Sola ◽

Eloïse Bertiaux-Lequoy ◽

Paul Guichard ◽

...

Keyword(s):

Mitochondrial Genome ◽

Trypanosoma Brucei ◽

Proteome Analysis ◽

Protozoan Parasite ◽

Eukaryotic Cell ◽

The Novel ◽

Precise Position ◽

Associated Proteins ◽

First Time

Proper mitochondrial genome inheritance is important for eukaryotic cell survival. Trypanosoma brucei, a protozoan parasite, contains a singular mitochondrial genome, the kDNA. The kDNA is anchored to the basal body via the tripartite attachment complex (TAC) to ensure proper segregation. Several components of the TAC have been described. However, the connection of the TAC to the kDNA remains elusive. Here, we characterize the TAC associated protein TAP110. Depletion as well as overexpression of TAP110 leads to a delay in the separation of the replicated kDNA networks. Proteome analysis after TAP110 overexpression identified several kDNA associated proteins including a TEX-like protein that dually localizes to the nucleus and the kDNA potentially linking replication/segregation in the two compartments. The assembly of TAP110 into the TAC region seems to require the TAC but not the kDNA itself, however once TAP110 has been assembled it also interacts with the kDNA. Finally, for the first time we use ultrastructure expansion microscopy in trypanosomes to reveal the precise position of TAP110 between TAC102 and the kDNA, showcasing the potential of this approach.

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Characterization of the Novel Mitochondrial Genome Segregation Factor TAP110 in Trypanosoma brucei

10.1101/2020.06.25.171090 ◽

2020 ◽

Cited By ~ 1

Author(s):

Simona Amodeo ◽

Ana Kalichava ◽

Albert Fradera-Sola ◽

Eloïse Bertiaux-Lequoy ◽

Paul Guichard ◽

...

Keyword(s):

Mitochondrial Genome ◽

Trypanosoma Brucei ◽

Protozoan Parasite ◽

Eukaryotic Cell ◽

The Novel ◽

Detergent Treatment ◽

Summary Statement ◽

First Time

AbstractProper mitochondrial genome inheritance is key for eukaryotic cell survival, however little is known about the molecular mechanism controlling this process. Trypanosoma brucei, a protozoan parasite, contains a singular mitochondrial genome aka kinetoplast DNA (kDNA). kDNA segregation requires anchoring of the genome to the basal body via the tripartite attachment complex (TAC). Several components of the TAC as well as their assembly have been described, it however remains elusive how the TAC connects to the kDNA. Here, we characterize the TAC associated protein TAP110 and for the first time use ultrastructure expansion microscopy in trypanosomes to reveal that TAP110 is the currently most proximal kDNA segregation factor. The kDNA proximal positioning is also supported by RNAi depletion of TAC102, which leads to loss of TAP110 at the TAC. Overexpression of TAP110 leads to expression level changes of several mitochondrial proteins and a delay in the separation of the replicated kDNA networks. In contrast to other kDNA segregation factors TAP110 remains only partially attached to the flagellum after DNAse and detergent treatment and can only be solubilized in dyskinetoplastic cells, suggesting that interaction with the kDNA might be important for stability of the TAC association. Furthermore, we demonstrate that the TAC, but not the kDNA, is required for correct TAP110 localization in vivo and suggest that TAP110 might interact with other proteins to form a >669 kDa complex.Summary StatementTAP110 is a novel mitochondrial genome segregation factor in Trypanosoma brucei that associates with the previously described TAC component TAC102. Ultrastructure expansion microscopy reveals its proximal position to the kDNA.

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