Microtubule–microtubule sliding by kinesin-1 is essential for normal cytoplasmic streaming in Drosophila oocytes

Abstract

Cytoplasmic streaming in Drosophila oocytes is a microtubulebased bulk cytoplasmic movement. Streaming efficiently circulates and localizes mRNAs and proteins deposited by the nurse cells across the oocyte. This movement is driven by kinesin-1, a major microtubule motor. Recently, we have shown that kinesin-1 heavy chain (KHC) can transport one microtubule on another microtubule, thus driving microtubule-microtubule sliding in multiple cell types. To study the role of microtubule sliding in oocyte cytoplasmic streaming, we used a Khc mutant that is deficient in microtubule sliding but able to transport a majority of cargoes. We demonstrated that streaming is reduced by genomic replacement of wild-type Khc with this sliding-deficient mutant. Streaming can be fully rescued by wild-type KHC and partially rescued by a chimeric motor that cannot move organelles but is active in microtubule sliding. Consistent with these data, we identified two populations of microtubules in fast-streaming oocytes: a network of stable microtubules anchored to the actin cortex and free cytoplasmic microtubules that moved in the ooplasm. We further demonstrated that the reduced streaming in sliding-deficient oocytes resulted in posterior determination defects. Together, we propose that kinesin-1 slides free cytoplasmic microtubules against cortically immobilized microtubules, generating forces that contribute to cytoplasmic streaming and are essential for the refinement of posterior determinants.kinesin-1 | microtubules | cytoplasmic streaming | Drosophila | axis determination

Moonlighting Motors: Kinesin, Dynein, and Cell Polarity

“…Inhibition of sliding by a kinesin mutation ( Khc mutA , see section “Kinesin-1-driven microtubule sliding in neurons and non-neuronal cells”) resulted in a more diffuse pattern of posterior determinants in the oocyte. We propose that the cytoplasmic streaming refines the posterior determination by bringing determinants that are attached to moving microtubules to the posterior pole for proper anchorage [22]. This is consistent with previous studies that suggest streaming transports polarity determinants in the oocytes [68–70].…”

“…Both the motor domain and the ATP-independent microtubule binding site at the C-terminus of the heavy chain are highly evolutionary conserved [5, 6, 22] (Figure 1). Therefore, we propose that microtubule sliding can serve as a universal and essential mechanism driving reorganization of the microtubule networks.…”

“…Our recent study of Drosophila ooplasmic streaming revisited this hitchhiking hypothesis. We showed that kinesin-driven microtubule sliding is required for cytoplasmic streaming in Drosophila oocytes [22]. Unlike sliding in neurons, microtubule sliding in oocytes occurs between the cortically-anchored and free microtubules.…”

Role of kinesin-1–based microtubule sliding in Drosophila nervous system development

“…However, because the rescue was partial, MT sliding is likely important in other tissues as well. In support of this idea, an accompanying study demonstrates the role of MT sliding in cytoplasmic streaming and polarity determination during Drosophila oogenesis (35). We also tested whether we could rescue lethality in Khc 27 flies, a kinesin-1-null allele (14).…”

Mechanisms of Kinesin-1 activation by Ensconsin/MAP7in vivo

“…Stage 10 oocytes were isolated with two 27-gauge syringe needles in Halocarbon oil (Sigma), mounted and observed by light microscopy using a DMRXA2 microscope (Leica). Velocity analyses of the particles were performed as described in two separate studies ( 20,21 ) except that three kymographs were obtained for each oocyte along active flows and 5 particles (15 per oocyte) were analyzed for each kymograph using the Multi Kymograph plugin for Fiji.…”