Assembly of the paraflagellar rod and the flagellum attachment zone complex during the Trypanosoma brucei cell cycle

Trypanosomes possess a single flagellum that is attached to their cell body via the flagellum attachment zone (FAZ). The FAZ is composed of two struct ures: a cytoplasmic filament complex and four microtubules situated next to it. There is a complex transmembrane crosslinking of this FAZ to the paraf lagellar rod (PFR) and axoneme within the flagellum. We have partially puri fied the FAZ complex and have produced monoclonal antibodies both against t he FAZ and the paraflagellar rod. The two antibodies against the FAZ (L3B2 and L6B3) recognise the cytoplasmic filament in immunofluorescence and in i mmunoelectron microscopy. On Western blot, they detect a doublet of high mo lecular weight (M-r 200,000). Two anti-PFR antibodies (L13D6 and L8C4) reco gnise the paraflagellar rod in immunofluorescence, but show a difference on Western blot: L13D6 recognises both major PFR proteins, whereas L8C4 is sp ecific for only one of them. Using these new antibodies we have shown that although the growth of both cytoplasmic FAZ filament and external PFR are r elated, their growth initiates at different time points during the cell cyc le and the two structures elongate at distinct rates.