Many mRNAs, proteins and organelles, whose synthesis occurs in discrete regions or compartments of cells, need to be dispersed throughout the cytoplasm for delocalized functions, while others need to be concentrated asymmetrically for localized functions. Cytoplasm is a gel-like fluid that allows small molecules to diffuse freely but restricts diffusion of large molecules and supramolecular complexes (Luby-Phelps, 2000
). Thus, for both dispersal and for asymmetric localization of large complexes, cells have evolved machinery that actively transports cytoplasmic components that do not diffuse well (Saxton, 2001
; Scholey et al., 2003
; Shimmen and Yokota, 2004
; Vale, 2003
Intracellular arrays of cytoskeletal filaments (F-actin or microtubules) are required for most forms of active transport (Vale, 2003
). The filaments, which have structural polarity, act as directional tracks for the transport of organelles or other cargoes by molecular motors; myosins that move on F-actin or kinesins and dyneins that move on microtubules. Many microtubule motors act as force-producing crosslinks with a mechanochemical, filament-binding ‘head’ at one end and a cargo-binding ‘tail’ at the other. Motors of the kinesin-1 subfamily (conventional kinesins) move cargoes towards microtubule plus-ends, which are usually distal to microtubule organizing centers (MTOCs), while cytoplasmic dynein moves cargoes toward minus-ends, which are usually at or near MTOCs. Thus, the positions of MTOCs and the paths of microtubules that project away from them dictate the directions and paths of microtubule-based cargo movements.
It is thought that kinesins and cytoplasmic dynein can attach to the same cargo, which raises questions about conflict between their opposing forces. Live imaging has shown that cargoes usually move alternately toward microtubule plus- and minus-ends, in a saltatory manner. Net transport is accomplished by a bias in favor of one direction (Mallik and Gross, 2004
). Studies of melanosomes in Xenopus
(Deacon et al., 2003
), and lipid droplets, peroxisomes and mRNA particles in Drosophila
(Gross et al., 2002b
; Kural et al., 2005
; Ling et al., 2004
) suggest that minus- and plus-end microtubule motors strictly alternate rather than competing with one another in a tug-of-war. This raises the question of whether or not coordinated alternation of opposing motors is a universal feature of microtubule-based transport processes.
oocytes provide a good system for investigating microtubule-dependent transport. Microtubule motors are important both for targeted localization of polarity determinant mRNAs, and for dispersal of components delivered to the oocyte anterior from adjoining nurse cells. During midoogenesis, bicoid
) and gurken
) mRNAs are localized in the oocyte to their respective anterior, posterior and dorsal positions in a kinesin-1- and dynein-dependent manner (Brendza et al., 2000a
; Brendza et al., 2002
; Duncan and Warrior, 2002
; Januschke et al., 2002
). Throughout that period of targeted localization, slow microtubule-dependent bulk streaming movements occur (Gutzeit, 1986b
; Theurkauf et al., 1992
). After polarity determinant localizations are well established, microtubule-based streaming becomes fast and well-ordered just before nurse cells non-selectively dump their contents into the anterior end of the oocyte (Gutzeit and Koppa, 1982
). Coincident with the start of fast streaming, oocyte microtubules align to form bundles that lie parallel to the cortex (Theurkauf et al., 1992
). Although kinesin-1, dynein and microtubules are important for these processes, how they contribute and how they relate to one another remains poorly understood.
To address issues about the mechanism of streaming and how it influences targeted localization and dispersal transport processes, we used time-lapse confocal microscopy to study the behavior of endosomes, determinant mRNAs and microtubules during slow and fast streaming. Tests of kinesin-1 and cytoplasmic dynein suggest a novel competitive relationship during slow streaming stages. Suppression of dynein activity allows a transition to robust, fast, plus-end movement by kinesin-1 that aligns microtubules into parallel arrays, which orders and amplifies plus-end cargo motion and thus fast streaming of surrounding cytoplasm. An allelic series of Khc mutations revealed that while posterior oskar mRNA localization did not require streaming, it did require some kinesin-1 activity, supporting the hypothesis that kinesin-1 can form physical links with oskar RNPs that contribute to posterior oskar localization by direct microtubule-based transport.