QUICK SEARCH:   [advanced] Author: Keyword(s):
Home     Help     Feedback     Subscriptions     Archive     Search     Table of Contents

First published online August 2, 2005
doi: 10.1242/10.1242/dev.01956

This Article Summary Full Text Full Text (PDF) Supplementary Material Alert me when this article is cited Alert me if a correction is posted Services Email this article to a friend Related articles in Development Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Serbus, L. R. Articles by Saxton, W. M. Search for Related Content PubMed PubMed Citation Articles by Serbus, L. R. Articles by Saxton, W. M. Social Bookmarking                         What's this?

Dynein and the actin cytoskeleton control kinesin-driven cytoplasmic streaming in Drosophila oocytes

¹ Department of Biology, Indiana University, Bloomington, 1001 East 3rd Street, IN 47405, USA
² Program in Cell Dynamics, University of Massachusetts, 55 Lake Avenue, North Worcester, MA 01655, USA

View larger version (118K): [in a new window]   Fig. 1. Effects of Khc mutations on endosome motion in oocytes during stage 9. (A-D and Movies 1-3 in the supplementary material) Fluorescent yolk endosomes were imaged in egg chambers during slow streaming stages. Each panel is a projection of 10 images from one focal plane acquired at 15-second intervals. Moving endosomes appear as elongated streaks, while non-moving endosomes appear as circular spots. Genotypes are noted in each panel. Arrows in A and B indicate areas that had streaming movements (see Movie 1 in the supplementary material). (E) Overall mean and peak endosome velocities are shown for each genotype. Numbers in or above bars show sample sizes. Peak velocities were calculated as means of the fastest 10% of endosomes of each genotype. Scale bar: 25 µm.

View larger version (114K): [in a new window]   Fig. 2. Hypomorphic Khc alleles that block slow streaming did not block oskar mRNA localization. In situ hybridization with fluorescent oskar RNA probes in stage 10A-10B shows localization of oskar mRNA in oocytes with the indicated genotypes. (A) Wild type. (B,C) Some oskar accumulated and persisted at the centers of the hypomorphic mutant oocytes and concentrated strongly at the posterior (see also Fig. S2 in the supplementary material). (D) In Khc-null oocytes, oskar did not accumulate at either the center or posterior, but there was an elevated concentration around the cortex. Scale bar: 25 µm.

View larger version (99K): [in a new window]   Fig. 3. Effects of Khc mutations on endosome motion in oocytes during stages 10B-11. (A-D and Movies 4-6 in the supplementary material) Fluorescent yolk endosomes were imaged in isolated egg chambers during fast streaming stages. Each panel is a projection of 10 images from one focal plane acquired over 150 seconds. Fast-moving endosomes appear as strings of dots, while slower endosomes appear as blurred streaks. Non-moving endosomes appear as circular spots. Genotypes are indicated in each panel. Arrows in B,C indicate areas that had streaming movements. In C, the radial streaks at the upper right corner are from movement of debris outside the egg chamber. (E) Overall mean and peak endosome velocities are shown for each genotype. Numbers in or above bars show sample sizes for overall means. Peak velocities were calculated as means of the fastest 10% of endosomes for each genotype. Scale bar: 25 µm.

View larger version (120K): [in a new window]   Fig. 4. Effects of anti-dynein injections on endosome movement in oocytes during stages 9-10A. Each panel is a projection of eight images from one focal plane acquired at 20-second intervals. Moving endosomes appear as elongated streaks while non-moving endosomes appear as circular spots. (A-D) Wild-type oocytes are shown before injection (A,C) and after injection with antibodies specific for (B) dynein intermediate chain (anti-DIC) or (D) dynein heavy chain (anti-Dhc). (E) A wild-type oocyte after injection with nonspecific IgG. (F) A Khc-null oocyte after injection with anti-DIC. Better views of the effects of antibody injections are provided in Movies 7 and 8 in the supplementary material. Scale bar: 25 µm.

View larger version (122K): [in a new window]   Fig. 5. Anti-Khc injections blocked premature fast streaming in capu and spir mutants. Each panel is a projection of eight images from one focal plane acquired at 20-second intervals (see Movies 9-11 in the supplementary material). (A,B) Stage 11 wild-type oocytes injected with control antibody (A) or anti-Khc (B) show that antibody inhibition of kinesin-1 stopped normal fast streaming (see also Movies 9 and 10 in the supplementary material). (C,D) A capu1/capuHK oocyte before (C) and after (D) anti-Khc injection. (E,F) A spir1/spir1 oocyte before (E) and after (F) anti-Khc injection (Movie 11 in the supplementary material). Scale bars: in B, 50 µm for A,B; in D, 50 µm for C-F.

View larger version (96K): [in a new window]   Fig. 6. Oocyte microtubule behavior during slow and fast streaming stages. Spinning disk confocal images of (A) a fixed stage 9 oocyte (slow streaming stage) with fluorescence from -tubulin antibody staining and of (B) a live stage 9 oocyte with fluorescence from transgenic GFP::-tubulin. Time-lapse movies of GFP fluorescence show that randomly oriented microtubules underwent dynamic churning motions (which can only be seen in Movie 12 in the supplementary material). (C) A fixed stage 10B oocyte (fast streaming stage) with fluorescence from -tubulin antibody staining shows large microtubule bundles. (D) A live stage 10B fast streaming oocyte with fluorescence from GFP::-tubulin. Time-lapse movies of the GFP show that microtubules joined in loose, parallel arrays rather than tight bundles. The arrays were dynamic transient structures that aligned and bent with streaming currents (which can only be seen in Movie 13 in the supplementary material). (E) A fixed stage 10B Khc17 oocyte stained with -tubulin antibody that shows some aligned fine filamentous structures, but no large microtubule bundles. Scale bar: 25 µm.

View larger version (34K): [in a new window]   Fig. 7. A speculative model for slow and fast streaming in Drosophila oocytes. The lower panels represent expanded views of the regions enclosed by blue boxes in the upper panels. The underlying premise is that unidirectional transport of organelles along microtubules can impel surrounding cytoplasm to stream. (A) Before stage 10B, competition between dynein and kinesin-1 suppresses concerted plus-end organelle transport and prevents parallel ordering of microtubules, allowing only slow short-range currents. (B) During stage 10B, a dynein inhibitory signal is released from the actin-rich cortex. This allows concerted kinesin-driven organelle transport to impel strong plus-end-directed currents and sweep microtubules into parallel, but flexible, dynamic arrays.

CiteULike

Complore

Connotea

Del.icio.us

Digg

Reddit

Technorati

Twitter