Regulation of ER-Golgi Intermediate Compartment Tubulation and Mobility by COPI Coats, Motor Proteins and Microtubules

Abstract

Little is known about the formation and regulation of endoplasmic reticulum (ER)-Golgi transport intermediates, although previous studies suggest that cargo is the main regulator of their morphology. In this study, we analyze the role of coat protein I (COPI) and cytoskeleton in the formation of tubular ER-Golgi intermediate compartment (ERGIC) and also show that partial COPI detachment by means of low temperature (15 • C) or brefeldin A induces the formation of transient tubular ERGIC elements. Most of them moved from the cell periphery to the perinuclear area and were 2.5× slower than vesicles. Time-lapse analysis of living cells demonstrates that the ERGIC elements are able to shift very fast from tubular to vesicular forms and vice versa, suggesting that the amount of cargo is not the determining factor for ERGIC morphology. Both the partial microtubule depolymerization and the inhibition of uncoating of the membranes result in the formation of long tubules that grow from round ERGICs and form at complex network. Interestingly, both COPI detachment and microtubule depolymerization induce a redistribution of kinesin from peripheral ERGIC elements to the Golgi area, while dynein distribution is not affected. However, both kinesin and dynein downregulation by RNA interference induced ERGIC tubulation. The tubules induced by kinesin depletion were static, whereas those resulting from dynein depletion were highly mobile. Our results strongly suggest that the interaction of motor proteins with COPI-coated membranes and microtubules is a key regulator of ERGIC morphology and mobility.

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

“…Analysis of fluorescence intensity in the data sets described here indicates that the increased tubulation seen here is due to DHC1 suppression and not an increase in tsO45-G-GFP expression in these selected cases (not shown). These data are consistent with work showing that tubular transport intermediates are dependent on an intact microtubule network (Simpson et al, 2006), and that describing the formation of highly dynamic ERGIC tubules following motor depletion (Tomás et al, 2010). …”

Regulation of the Golgi complex by phospholipid remodeling enzymes

“…2, Step 5). ERGIC tubulation and mobility are also regulated by microtubules and their associated motor proteins and whether these tubule-inducing functions are independent or shared with PLA2G6/iPLA 2 -β remains unknown [138]. …”

Unconventional functions of microtubule motors

“…Finally, the microtubule motors participate, along with other specialized, coat-forming, and small GTP-binding proteins, in vesicle budding and membrane tubulation, including the generation of the early endocytic vesicles [105], or of the tubes of ER-to-Golgi intermediate compartment (ERGIC) [95, 106]. Of course, these activities rely on the presence of microtubules at the sites of membrane invagination.…”