Insights on UNC‐104‐dynein/dynactin interactions and their implications on axonal transport in Caenorhabditis elegans

Bidirectional cargo transport in neurons can be explained by two models: the “tug‐of‐war model” for short‐range transport, in which several kinesin and dynein motors are bound to the same cargo but travel in opposing directions, and by the “motor coordination model” for long‐range transport, in which small adaptors or the cargo itself activates or deactivates opposing motors. Direct interactions between the major axonal transporter kinesin‐3 UNC‐104(KIF1A) and the dynein/dynactin complex remains unknown. In this study, we dissected and evaluated the interaction sites between UNC‐104 and dynein as well as between UNC‐104 and dynactin using yeast two‐hybrid assays. We found that the DYLT‐1(Tctex) subunit of dynein binds near the coiled coil 3 (CC3) of UNC‐104, and that the DYRB‐1(Roadblock) subunit binds near the CC2 region of UNC‐104. Regarding dynactin, we specifically revealed strong interactions between DNC‐6(p27) and the FHA‐CC3 stretch of UNC‐104, as well as between the DNC‐5(p25) and the CC2‐CC3 region of UNC‐104. Motility analysis of motors and cargo in the nervous system of Caenorhabditis elegans revealed impaired transport of UNC‐104 and synaptic vesicles in dynein and dynactin mutants (or in RNAi knockdown animals). Further, in these mutants UNC‐104 clustering along axons was diminished. Interestingly, when dynamic UNC‐104 motors enter a stationary UNC‐104 cluster their dwelling times are increased in dynein mutants (suggesting that dynein may act as an UNC‐104 activator). In summary, we provide novel insights on how UNC‐104 interacts with the dynein/dynactin complex and how UNC‐104 driven axonal transport depends on dynein/dynactin in C. elegans neurons. We used yeast two‐hybrid assays to dissect the interaction domains between UNC‐104 and the dynein/dynactin complex. From our results, we propose two scenarios how synaptic vesicles may be transported in neurons by these two opposing motors. We also revealed how eliminating various domains affects axonal transport in C. elegans.