This study dissects the recruitment of dynein and dynactin to cargo by a conserved motor adaptor BICD2. It is shown that dynein, dynactin, and BICD2 form a triple complex in vitro and in vivo. Investigation of the properties of this complex by direct visualization of dynein in live cells shows that BICD2-induced dynein transport requires LIS1.

Rab6 is a conserved small GTPase that localizes to the Golgi apparatus and cytoplasmic vesicles and controls transport and fusion of secretory carriers [1]. Another Rab implicated in trafficking from the trans-Golgi to the plasma membrane is Rab8 [2-5]. Here we show that Rab8A stably associates with exocytotic vesicles in a Rab6-dependent manner. Rab8A function is not needed for budding or motility of exocytotic carriers but is required for their docking and fusion. These processes also depend on the Rab6-interacting cortical factor ELKS [1], suggesting that Rab8A and ELKS act in the same pathway. We show that Rab8A and ELKS can be linked by MICAL3, a member of the MICAL family of flavoprotein monooxygenases [6]. Expression of a MICAL3 mutant with an inactive monooxygenase domain resulted in a strong accumulation of secretory vesicles that were docked at the cell cortex but failed to fuse with the plasma membrane, an effect that correlated with the strongly reduced mobility of MICAL3. We propose that the monooxygenase activity of MICAL3 is required to regulate its own turnover and the concomitant remodeling of vesicle-docking protein complexes in which it is engaged. Taken together, the results of our study illustrate cooperation of two Rab proteins in constitutive exocytosis and implicates a redox enzyme in this process.

Cargo transport along microtubules is driven by the collective function of microtubule plus- and minus-end-directed motors (kinesins and dyneins). How the velocity of cargo transport is driven by opposing teams of motors is still poorly understood. Here, we combined inducible recruitment of motors and adaptors to Rab6 secretory vesicles with detailed tracking of vesicle movements to investigate how changes in the transport machinery affect vesicle motility. We find that the velocities of kinesin-based vesicle movements are slower and more homogeneous than those of dynein-based movements. We also find that Bicaudal D (BICD) adaptor proteins can regulate dynein-based vesicle motility. BICD-related protein 1 (BICDR-1) accelerates minus-end-directed vesicle movements and affects Rab6 vesicle distribution. These changes are accompanied by reduced axonal outgrowth in neurons, supporting their physiological importance. Our study suggests that adaptor proteins can modulate the velocity of dynein-based motility and thereby control the distribution of transport carriers.

Ciliary membrane composition is controlled by transition zone (TZ) proteins such as RPGRIP1, RPGRIPL and NPHP4, which are vital for balanced coordination of diverse signalling systems like the Sonic hedgehog (Shh) pathway. Activation of this pathway involves Shh-induced ciliary accumulation of Smoothened (SMO), which is disrupted by disease-causing mutations in TZ components. Here we identify kinesin-3 motor protein KIF13B as a novel member of the RPGRIP1N-C2 domain-containing protein family and show that KIF13B regulates TZ membrane composition and ciliary SMO accumulation. KIF13B is upregulated during ciliogenesis and is recruited to the ciliary base by NPHP4, which binds to two distinct sites in the KIF13B tail region, including an RPGRIP1N-C2 domain. KIF13B and NPHP4 are both essential for establishment of a CAV1 membrane microdomain at the TZ, which in turn is required for Shh-induced ciliary SMO accumulation. Thus KIF13B is a novel regulator of ciliary TZ configuration, membrane composition and Shh signalling.