TFK1, a basal body transition fibre protein that is essential for cytokinesis in Trypanosoma brucei

Abstract

In Trypanosoma brucei, transition fibres (TF) form a nine-bladed pattern-like structure connecting the base of the flagellum to the flagellar pocket membrane. Despite the characterization of two TF proteins, CEP164C and TbRP2, little is known about the organization of these fibres. Here, we report the identification and characterization of the first kinetoplastid-specific TF protein named TFK1 (Tb927.6.1180). Bioinformatics and functional domain analysis identified three TFK1 distinct domains: an N-terminal domain of an unpredicted function, a coiled-coil domain involved in TFK1-TFK1 interaction and a C-terminal intrinsically disordered region potentially involved in protein interaction. Cellular immuno-localization showed that TFK1 is a newly identified basal body maturation marker. Further, using ultrastructure expansion and immuno-electron microscopies we localized CEP164C and TbRP2 at the TF and TFK1 on the distal appendage matrix of the TF. Importantly, RNAi knockdown of TFK1 in bloodstream form cells induced misplacement of basal bodies, a defect in the furrow or fold generation and eventually cell death. We hypothesize that TFK1 is a basal body positioning specific actor and a key regulator of cytokinesis in the bloodstream form Trypanosoma brucei.

Identification of 30 transition fibre proteins in Trypanosoma brucei reveals a complex and dynamic structure

“…Retinitis pigmentosa-2 (RP-2) (Stephan et al, 2007) and TFK1 to the matrix between the TF blades (Ramanantsalama et al, 2022). Our screen identified and confirmed that a further 30 proteins localise to the TFs in trypanosomes, bringing the total to 32.…”

“…Two TFPs have previously been experimentally confirmed in trypanosomes: retinitis pigmentosa-2 ( Stephan et al, 2007 ) and TFK1, which localises to the matrix between the transition fibre blades ( Ramanantsalama et al, 2022 ). Our screen identified and confirmed that a further 30 proteins localise to the transition fibres in trypanosomes, bringing the total number of TFPs to 32.…”

“…Scanning transmission electron tomography studies have confirmed the presence of the canonical nine blades of transition fibres docked to the flagellar pocket in trypanosomes ( Trépout et al, 2018 ). In addition, two widely conserved proteins – CEP164C (CEP164 in mammals) and retinitis pigmentosa-2 (RP-2, also known as XRP2 in mammals) – have been determined to be transition fibre components ( Stephan et al, 2007 ; Harmer et al, 2017 ; Atkins et al, 2021 ), and a kinetoplastid-specific protein, TFK1, has been found to localise to transition fibres with CEP164C and RP-2 ( Ramanantsalama et al, 2022 ). In overall molecular terms, transition fibres have broadly been considered a simple structure comprising only a handful of proteins.…”

Identification of 30 transition fibre proteins reveals a complex and dynamic structure with essential roles in ciliogenesis

“…Complexity of transition fibres in ciliated eukaryotes Our screen identified and confirmed 30 proteins that localise to the TFs in trypanosomes. Two further TFPs were previously experimentally confirmed in Trypanosomes -Retinitis pigmentosa-2 (RP-2) (Stephan et al, 2007) and TFK1 to the matrix between the TF blades (Ramanantsalama et al, 2022) bringing the current number to 32. This challenges the traditional view that TFs are simple structures comprising only a few proteins and demonstrates that trypanosome TFs are complex structures.…”

“…The copyright holder for this preprint (which this version posted May 29, 2023. ; https://doi.org/10.1101/2023.05.29.542710 doi: bioRxiv preprint (Ramanantsalama et al, 2022). Overall in molecular terms, transition fibres have broadly been considered a simple structure comprising only a handful of proteins.…”