In vivo imaging reveals independent intraflagellar transport of the nexin–dynein regulatory complex subunits DRC2 and DRC4

Many axonemal proteins enter cilia and flagella on intraflagellar transport (IFT) trains, which move bidirectionally along the axonemal microtubules. Certain axonemal substructures including the radial spokes and outer dynein arms are preassembled in the cell body and transported as multisubunit complexes into flagella by IFT. Here, we used in vivo imaging to analyze the transport and assembly of DRC2 and DRC4, two core subunits of the nexin–dynein regulatory complex (N-DRC). Tagged DRC2 moved by IFT in mutants lacking DRC4 and vice versa, showing that they do not depend on each other for IFT. Simultaneous imaging of tagged DRC2 and DRC4, expressed from transgenes that rescue a corresponding double mutant, mostly showed transport on separate IFT trains, but occasional cotransports were also observed. The results demonstrate that DRC2 and DRC4 are transported largely independently of each other into flagella. These studies suggest that the N-DRC assembles onto the axoneme by the stepwise addition of subunits.