Genetic dissection of the Leishmania paraflagellar rod, a unique flagellarcytoskeleton structure

mutant revealed that: (1) fibers that attach the PFR to the axoneme did not contain PFR1 or PFR2, and assemble in the absence of a PFR. (2) PFR1 was s ynthesized and transported to the flagella in the absence of PFR2, where it formed a stable association with the axoneme attachment fibers, (3) PFR2 w as synthesized and transported to the flagella in the absence of PFR1, thou gh it was not found associated with the axoneme attachment fibers, (4) PFR1 and PFR2 were located throughout the subdomains of the PFR, These data sug gest that while PFR filaments contain both PFR1 and PFR2, the PFR is attach ed to the axoneme by interaction of PFR1 with the axoneme attachment fibers . The paraflagellar rod (PFR) is a unique network of cytoskeletal filaments that lies alongside the axoneme in the flagella of most trypanosomatids. W hile little is known about how two major Leishmania mexicana PFR protein co mponents, PFR1 and PFR2, assemble into this complex structure, previous ana lysis of PFR2 null mutants demonstrated that the PFR is essential for prope r cell motility. The structural roles of PFR1 and PFR2 are now examined thr ough comparison of PFR2 null mutants with new PFR1 null mutant and PFR1/PFR 2 double null mutant parasites. Both PFR1 and PFR2 were essential for PFR f ormation and cell motility. When elimination of one PFR gene prevented asse mbly of a native PFR structure, the other PFR protein accumulated at the di stal flagellar tip. Comparison of PFR substructures remaining in each mutan t revealed that: (1) fibers that attach the PFR to the axoneme did not cont ain PFR1 or PFR2, and assemble in the absence of a PFR. (2) PFR1 was synthe sized and transported to the flagella in the absence of PFR2, where it form ed a stable association with the axoneme attachment fibers, (3) PFR2 was sy nthesized and transported to the flagella in the absence of PFR1, though it was not found associated with the axoneme attachment fibers. (4) PFR1 and PFR2 were located throughout the subdomains of the PFR, These data suggest that while PFR filaments contain both PFR1 and PFR2, the PFR is attached to the axoneme by interaction of PFR1 with the axoneme attachment fibers.