Opposing microtubule motors control motility, morphology and cargo segregation during ER-to-Golgi transport

Abstract

We recently demonstrated that dynein and kinesin motors drive multiple aspects of endosomal function in mammalian cells. These functions include driving motility, maintaining morphology (notably through providing longitudinal tension to support vesicle fission), and driving cargo sorting. Microtubule motors drive bidirectional motility during traffic between the endoplasmic reticulum (ER) and Golgi. Here, we have examined the role of microtubule motors in transport carrier motility, morphology, and domain organization during ER-to-Golgi transport. We show that, consistent with our findings for endosomal dynamics, microtubule motor function during ER-to-Golgi transport of secretory cargo is required for motility, morphology, and cargo sorting within vesicular tubular carriers en route to the Golgi. Our data are consistent with previous findings that defined roles for dynein-1, kinesin-1 (KIF5B) and kinesin-2 in this trafficking step. Our high resolution tracking data identify some intriguing aspects. Depletion of kinesin-1 reduces the number of motile structures seen, which is in line with other findings relating to the role of kinesin-1 in ER export. However, those transport carriers that were produced had a much greater run length suggesting that this motor can act as a brake on anterograde motility. Kinesin-2 depletion did not significantly reduce the number of motile transport carriers but did cause a similar increase in run length. These data suggest that kinesins act as negative regulators of ER-to-Golgi transport. Depletion of dynein not only reduced the number of motile carriers formed but also caused tubulation of carriers similar to that seen for sorting nexin-coated early endosomes. Our data indicated that the previously observed anterograde–retrograde polarity of transport carriers in transit to the Golgi from the ER is maintained by microtubule motor function.

Cytoplasmic dynein and its regulatory proteins in Golgi pathology in nervous system disorders

“…“LIC1-dynein” and “LIC2-dynein” have been implicated in different functions, but the extent to which they are functionally redundant is uncertain (Allan, 2011 ; Raaijmakers et al, 2013 ). A knockdown study suggested specific LIC1 functions in ER-Golgi trafficking (Palmer et al, 2009 ; Brown et al, 2014 ), but no Golgi abnormalities were found after LIC1 knockdown by others (Allan, 2011 ; Tan et al, 2011 ). Mice, derived from a mutagenesis screen, that were homozygous for a missense mutation (N215Y) in LIC1, and showed mild behavioral changes combined with changes in dendritic morphology also showed abnormalities in Golgi ribbon reassembly following exposure to nocodazole, pointing to a role of LIC1 in Golgi membrane trafficking (Banks et al, 2011 ).…”

“…How does dynein-Golgin160 activity cross-talk with other motors, i.e., kinesins and myosin (Barlan et al, 2013 )? Knock-down of the kinesin KIF5B results in longer run lengths of ER-to-Golgi membrane carriers, indicating a negative regulation of ER-to Golgi transport by plus-end motors, and the presence of motors of opposite polarity on ER-to-Golgi cargo (Brown et al, 2014 ). A variety of scaffolds such as huntingtin have been implicated in the coordination of minus- and plus-end motors for specific retrograde and anterograde axonal transport cargo's (Maday et al, 2014 ).…”

Assembly and activation of dynein–dynactin by the cargo adaptor protein Hook3

“…The mammalian LICs, encoded by two closely related gene products, LIC1 and LIC2 ( Hughes et al, 1995 ; Tynan et al, 2000 ), are involved in several different types of cargo interactions and dynein-based movements, including endosomal and lysosomal transport, ER export, Golgi transport, and axonal vesicle trafficking ( Koushika et al, 2004 ; Palmer et al, 2009 ; Horgan et al, 2010 ; Tan et al, 2011 ; Kong et al, 2013 ; Brown et al, 2014 ). The domain structure of the LIC allows it to interact with cargo adaptors while integrated into the dynein holoenzyme.…”

Intermediate compartment (IC): from pre-Golgi vacuoles to a semi-autonomous membrane system

“…Moreover, its dissociation from membranes during mitosis coincides with inhibition of the centralization of IC elements that still interact with spindle MTs (Marie et al 2012 ; Yadav et al 2012 ). Notably, the IC also contains the plus-end directed motor kinesin (Lippincott-Schwartz et al 1995 ; Tomás et al 2010 ), which could be involved in retrograde transport, the shaping of the vacuolar and tubular elements or their positioning at ERES and/or Golgi region (Brown et al 2014 ). A special class of MTs that are nucleated at cis -Golgi membranes (Rivero et al 2009 ) has been implicated in the formation of the Golgi ribbon.…”