FAP20 is required for flagellum assembly in Trypanosoma brucei

Abstract

Trypanosoma brucei is a human and animal pathogen that depends on flagellar motility for transmission and infection. The trypanosome flagellum is built around a canonical "9+2" axoneme, containing nine doublet microtubules (DMTs) surrounding two singlet microtubules. Each DMT contains a 13-protofilament A-tubule and a 10-protofilament B-tubule, connected to the A-tubule by a conserved, non-tubulin inner junction (IJ) filament made up of alternating PACRG and FAP20 subunits. Here we investigate FAP20 in procyclic form T. brucei. A FAP20-NeonGreen fusion protein localized to the axoneme as expected. Surprisingly, FAP20 knockdown led to a catastrophic failure in flagellum assembly and concomitant lethal cell division defect. This differs from other organisms, where FAP20 is required for normal flagellum motility, but generally dispensable for flagellum assembly and viability. Transmission electron microscopy demonstrates failed flagellum assembly in FAP20 mutants is associated with a range of DMT defects and defective assembly of the paraflagellar rod, a lineage-specific flagellum filament that attaches to DMT 4-7 in trypanosomes. Our studies reveal a lineage-specific requirement for FAP20 in trypanosomes, offering insight into adaptations for flagellum stability and motility in these parasites and highlighting pathogen versus host differences that might be considered for therapeutic intervention in trypanosome diseases. SIGNIFICANCE STATEMENTO_LIDiverse eukaryotic organisms rely on a generally conserved axoneme architecture and dynein-dependent beating mechanism to drive motility, but mechanisms conferring lineage-specific motility needs are largely unknown. C_LIO_LIFAP20 is a conserved flagellar protein that impacts flagellum motility in multiple organisms. C_LIO_LIThe current work demonstrates FAP20 is particularly important in the pathogen, T. brucei, providing insight into pathogen adaptations for moving through host environments and illuminating targets to consider for therapeutic intervention in trypanosome diseases. C_LI