title: Roles of dynein and dynactin in early endosome dynamics revealed using automated tracking and global analysis.
abstract: Microtubule-dependent movement is crucial for the spatial organization of endosomes in most eukaryotes, but as yet there has been no systematic analysis of how a particular microtubule motor contributes to early endosome dynamics. Here we tracked early endosomes labeled with GFP-Rab5 on the nanometer scale, and combined this with global, first passage probability (FPP) analysis to provide an unbiased description of how the minus-end microtubule motor, cytoplasmic dynein, supports endosome motility. Dynein contributes to short-range endosome movement, but in particular drives 85-98% of long, inward translocations. For these, it requires an intact dynactin complex to allow membrane-bound p150(Glued) to activate dynein, since p50 over-expression, which disrupts the dynactin complex, inhibits inward movement even though dynein and p150(Glued) remain membrane-bound. Long dynein-dependent movements occur via bursts at up to ∼8 µms(-1) that are linked by changes in rate or pauses. These peak speeds during rapid inward endosome movement are still seen when cellular dynein levels are 50-fold reduced by RNAi knock-down of dynein heavy chain, while the number of movements is reduced 5-fold. Altogether, these findings identify how dynein helps define the dynamics of early endosomes.
authors.affiliation: Faculty of Life Sciences, University of Manchester, Manchester, United Kingdom.
authors.forename: Neftali
authors.initials: N
authors.lastname: Flores-Rodriguez
authors.forename: Salman S
authors.initials: SS
authors.lastname: Rogers
authors.forename: David A
authors.initials: DA
authors.lastname: Kenwright
authors.forename: Thomas A
authors.initials: TA
authors.lastname: Waigh
authors.forename: Philip G
authors.initials: PG
authors.lastname: Woodman
authors.forename: Victoria J
authors.initials: VJ
authors.lastname: Allan
mesh_terms: D015153:Blotting, Western; D002460:Cell Line; D045744:Cell Line, Tumor; D000072159:Dynactin Complex; D004398:Dyneins / Q000235:genetics / Q000378:metabolism*; D011992:Endosomes / Q000378:metabolism*; D006801:Humans; D008869:Microtubule-Associated Proteins / Q000235:genetics / Q000378:metabolism*; D016133:Polymerase Chain Reaction; D034622:RNA Interference; D020696:rab5 GTP-Binding Proteins / Q000235:genetics / Q000378:metabolism
chemical_list: C000606360:DCTN1 protein, human; D000072159:Dynactin Complex; D008869:Microtubule-Associated Proteins; D004398:Dyneins; D020696:rab5 GTP-Binding Proteins