scholarly journals A novel mechanism of bulk cytoplasmic transport by cortical dynein in Drosophila ovary

2021 ◽  
Author(s):  
Wen Lu ◽  
Margot Lakonishok ◽  
Anna S. Serpinskaya ◽  
Vladimir I Gelfand
Keyword(s):  
Cytoplasmic Dynein ◽  
Cell Cortex ◽  
Nurse Cells ◽  
Oocyte Growth ◽  
Actin Cortex ◽  
Cytoplasmic Transport ◽  
Ring Canals ◽  
Microtubule Motor ◽  
Cytoplasmic Bridges ◽  
Drosophila Ovary

Cytoplasmic dynein, a major minus-end directed microtubule motor, plays essential roles in eukaryotic cells. Drosophila oocyte growth is mainly dependent on the contribution of cytoplasmic contents from the interconnected sister cells, nurse cells. We have previously shown that cytoplasmic dynein is required for Drosophila oocyte growth, and assumed that it transports cargoes along microtubule tracks from nurse cells to the oocyte. Here we report that instead transporting cargoes along microtubules into the oocyte, cortical dynein actively moves microtubules in nurse cells and from nurse cells to the oocyte via the cytoplasmic bridges, the ring canals. We demonstrate this microtubule movement is sufficient to drag even inert cytoplasmic particles through the ring canals to the oocyte. Furthermore, replacing dynein with a minus-end directed plant kinesin linked to the actin cortex is sufficient for transporting organelles and cytoplasm to the oocyte and driving its growth. These experiments show that cortical dynein can perform bulk cytoplasmic transport by gliding microtubules along the cell cortex and through the ring canals to the oocyte. We propose that the dynein-driven microtubule flow could serve as a novel mode of cargo transport for fast cytoplasmic transfer to support rapid oocyte growth.  

scholarly journals Short stop is a gatekeeper at the ring canals of Drosophila ovary

2020 ◽  
Author(s):  
Wen Lu ◽  
Margot Lakonishok ◽  
Vladimir I. Gelfand
Keyword(s):  
Actin Filaments ◽  
Cytoskeletal Proteins ◽  
Nurse Cells ◽  
Cellular Functions ◽  
Oocyte Growth ◽  
Ring Canals ◽  
Cytoplasmic Bridges ◽  
Drosophila Ovary ◽  
Open Nature ◽  
Sister Cells

SUMMARYMicrotubules and actin filaments are two major cytoskeletal components essential for a variety of cellular functions. Spectraplakins are a family of large cytoskeletal proteins cross-linking microtubules and actin filaments among other components. In this study, we aim to understand how Short stop (Shot), the single Drosophila spectraplakin, coordinates microtubules and actin filaments for oocyte growth. The oocyte growth completely relies on the acquisition of cytoplasmic materials from the interconnected sister cells (nurse cells), through ring canals, cytoplasmic bridges that remained open after incomplete germ cell division. Given the open nature of the ring canals, it is unclear how the direction of transport through the ring canal is controlled. Here we show that Shot controls the directionality of flow of material from the nurse cells towards the oocyte. Knockdown of shot changes the direction of transport of many types of cargo through the ring canals from unidirectional (toward the oocyte) to bidirectional, resulting in small oocytes that fail to grow over time. In agreement with this flow-directing function of Shot, we find that it is localized at the asymmetric actin fibers adjacent to the ring canals at the nurse cell side, and controls the uniform polarity of microtubules located in the ring canals connecting the nurse cells and the oocyte. Together, we propose that Shot functions as a gatekeeper directing the material flow from the nurse cells to the oocyte, via organization of microtubule tracks.

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

2016 ◽  
Vol 113 (34) ◽  
pp. E4995-E5004 ◽  
Author(s):  
Wen Lu ◽  
Michael Winding ◽  
Margot Lakonishok ◽  
Jill Wildonger ◽  
Vladimir I. Gelfand
Keyword(s):  
Cytoplasmic Streaming ◽  
Cell Types ◽  
Nurse Cells ◽  
Wild Type ◽  
Actin Cortex ◽  
Microtubule Motor ◽  
Multiple Cell ◽  
Cytoplasmic Microtubules ◽  
Two Populations

Cytoplasmic streaming in Drosophila oocytes is a microtubule-based 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.

scholarly journals A novel pattern of germ cell divisions in the production of hymenopteran insect eggs

2020 ◽  
Vol 16 (5) ◽  
pp. 20200137
Author(s):  
Katherine J. Eastin ◽  
Austin P. Huang ◽  
Patrick M. Ferree
Keyword(s):  
Germ Cell ◽  
Germ Cells ◽  
Radial Direction ◽  
Fruit Fly ◽  
Cell Number ◽  
Nurse Cells ◽  
Insect Eggs ◽  
Ring Canals ◽  
Cytoplasmic Bridges ◽  
Higher Eukaryotes

Egg development is a defining process of reproduction in higher eukaryotes. In the fruit fly, Drosophila melanogaster , this process begins with four mitotic divisions starting from a single germ cell, producing a cyst of 16 cystocytes; one of these cells will become the oocyte and the others supporting nurse cells. These mitotic divisions are exceptional because cytokinesis is incomplete, resulting in the formation of cytoplasmic bridges known as ring canals that interconnect the cystocytes. This organization allows all cystocytes to divide synchronously during each mitotic round, resulting in a final, power-of-2 number of germ cells. Given that numerous insects obey this power-of-2 rule, we investigated if strict cell doubling is a universal, underlying cause. Using confocal microscopy, we found striking departures from this paradigm in three different power-of-2 insects belonging to the Apocrita suborder (ants, bees and wasps). In these insects, the earliest-formed cystocytes cease to divide during the latter mitotic cycles while their descendants undergo further division, thereby producing a ‘radial’ direction of division activity. Such cystocyte division patterns that depart from strict cell doubling may be ‘fine-tuned’ in order to maintain a final, power-of-2 germ cell number.

scholarly journals A Dual Role for Actin and Microtubule Cytoskeleton in the Transport of Golgi Units from the Nurse Cells to the Oocyte Across Ring Canals

2009 ◽  
Vol 20 (1) ◽  
pp. 556-568 ◽  
Author(s):  
Emmanuelle Nicolas ◽  
Nicolas Chenouard ◽  
Jean-Christophe Olivo-Marin ◽  
Antoine Guichet
Keyword(s):  
Embryonic Development ◽  
Transport Process ◽  
Myosin Ii ◽  
Nurse Cells ◽  
Transport Mechanisms ◽  
Step Process ◽  
Actin Networks ◽  
Ring Canals ◽  
Cytoplasmic Bridges ◽  
Active Transport Process

Axis specification during Drosophila embryonic development requires transfer of maternal components during oogenesis from nurse cells (NCs) into the oocyte through cytoplasmic bridges. We found that the asymmetrical distribution of Golgi, between nurse cells and the oocyte, is sustained by an active transport process. We have characterized actin basket structures that asymmetrically cap the NC side of Ring canals (RCs) connecting the oocyte. Our results suggest that these actin baskets structurally support transport mechanisms of RC transit. In addition, our tracking analysis indicates that Golgi are actively transported to the oocyte rather than diffusing. We observed that RC transit is microtubule-based and mediated at least by dynein. Finally, we show that actin networks may be involved in RC crossing through a myosin II step process, as well as in dispatching Golgi units inside the oocyte subcompartments.

scholarly journals A Diaphanous and Enabled dependent asymmetric actin cable array repositions nuclei during Drosophila oogenesis

2020 ◽  
Author(s):  
Gregory Logan ◽  
Brooke M. McCartney
Keyword(s):  
Growth Rate ◽  
Nurse Cell ◽  
Cell Cortex ◽  
Nurse Cells ◽  
Nuclear Movement ◽  
Actin Cable ◽  
Rate Asymmetry ◽  
Ring Canals ◽  
Differential Localization ◽  
Actin Cables

AbstractCells reposition their nuclei for a diversity of specialized functions through a wide variety of cytoskeletal mechanisms. To complete oogenesis, Drosophila nurse cells employ novel actin cable arrays to reposition their nuclei. During oogenesis, 15 nurse cells connected by ring canals contract to “dump” their cytoplasmic contents into the oocyte. Just prior to dumping, actin cables initiate from the nurse cell cortex and elongate toward their nuclei, pushing them away from the ring canals to prevent obstruction. How the actin cable arrays generate directional nuclear movement is not known. We found regional differences in the actin cable growth rate that are dependent on the differential localization of the actin assembly factors Enabled (Ena) and Diaphanous (Dia). Mislocalization of Ena resulted in actin cable arrays with a uniform growth rate. In the absence of growth rate asymmetry, nuclear relocation was significantly altered and cytoplasmic dumping was incomplete. This novel mechanism for nuclear repositioning relies on the regulated cortical localization of Dia and Ena producing asymmetric actin cable arrays that push the nuclei away from the ring canals, enabling successful oogenesis.Summary statementThis work demonstrates that an asymmetric actin cable array regulated by the differential localization of Diaphanous and Enabled is necessary to reposition nurse cell nuclei and complete oogenesis in Drosophila.

scholarly journals Dynactin is required for bidirectional organelle transport

2003 ◽  
Vol 160 (3) ◽  
pp. 297-301 ◽  
Author(s):  
Sean W. Deacon ◽  
Anna S. Serpinskaya ◽  
Patricia S. Vaughan ◽  
Monica Lopez Fanarraga ◽  
Isabelle Vernos ◽  
...  
Keyword(s):  
Cell Types ◽  
Cytoplasmic Dynein ◽  
Opposite Polarity ◽  
Model System ◽  
Organelle Transport ◽  
Transport Activity ◽  
Microtubule Motors ◽  
Microtubule Motor ◽  
Dynactin Complex ◽  
Binding Subunit

Kinesin II is a heterotrimeric plus end–directed microtubule motor responsible for the anterograde movement of organelles in various cell types. Despite substantial literature concerning the types of organelles that kinesin II transports, the question of how this motor associates with cargo organelles remains unanswered. To address this question, we have used Xenopus laevis melanophores as a model system. Through analysis of kinesin II–mediated melanosome motility, we have determined that the dynactin complex, known as an anchor for cytoplasmic dynein, also links kinesin II to organelles. Biochemical data demonstrates that the putative cargo-binding subunit of Xenopus kinesin II, Xenopus kinesin II–associated protein (XKAP), binds directly to the p150Glued subunit of dynactin. This interaction occurs through aa 530–793 of XKAP and aa 600–811 of p150Glued. These results reveal that dynactin is required for transport activity of microtubule motors of opposite polarity, cytoplasmic dynein and kinesin II, and may provide a new mechanism to coordinate their activities.

scholarly journals Microtubule-based Endoplasmic Reticulum Motility in Xenopus laevis: Activation of Membrane-associated Kinesin during Development

1999 ◽  
Vol 10 (6) ◽  
pp. 1909-1922 ◽  
Author(s):  
Jon D. Lane ◽  
Victoria J. Allan
Keyword(s):  
Endoplasmic Reticulum ◽  
Cell Types ◽  
Cytoplasmic Dynein ◽  
Culture Cell ◽  
Tissue Culture Cell ◽  
Differential Interference Contrast Microscopy ◽  
Animal Cells ◽  
Microtubule Motor ◽  
Directed Motility ◽  
Conventional Kinesin

The endoplasmic reticulum (ER) in animal cells uses microtubule motor proteins to adopt and maintain its extended, reticular organization. Although the orientation of microtubules in many somatic cell types predicts that the ER should move toward microtubule plus ends, motor-dependent ER motility reconstituted in extracts ofXenopus laevis eggs is exclusively a minus end-directed, cytoplasmic dynein-driven process. We have used Xenopusegg, embryo, and somatic Xenopus tissue culture cell (XTC) extracts to study ER motility during embryonic development inXenopus by video-enhanced differential interference contrast microscopy. Our results demonstrate that cytoplasmic dynein is the sole motor for microtubule-based ER motility throughout the early stages of development (up to at least the fifth embryonic interphase). When egg-derived ER membranes were incubated in somatic XTC cytosol, however, ER tubules moved in both directions along microtubules. Data from directionality assays suggest that plus end-directed ER tubule extensions contribute ∼19% of the total microtubule-based ER motility under these conditions. In XTC extracts, the rate of ER tubule extensions toward microtubule plus ends is lower (∼0.4 μm/s) than minus end-directed motility (∼1.3 μm/s), and plus end-directed motility is eliminated by a function-blocking anti-conventional kinesin heavy chain antibody (SUK4). In addition, we provide evidence that the initiation of plus end-directed ER motility in somatic cytosol is likely to occur via activation of membrane-associated kinesin.

Real time imaging reveals a peroxisomal reticulum in living cells

2000 ◽  
Vol 113 (20) ◽  
pp. 3663-3671 ◽  
Author(s):  
M. Schrader ◽  
S.J. King ◽  
T.A. Stroh ◽  
T.A. Schroer
Keyword(s):  
Real Time ◽  
Cytoplasmic Dynein ◽  
Living Cells ◽  
Real Time Imaging ◽  
Peroxisomal Targeting ◽  
Microtubule Motor ◽  
Targeting Signal ◽  
Peroxisomal Targeting Signal

We have directly imaged the dynamic behavior of a variety of morphologically different peroxisomal structures in HepG2 and COS-7 cells transfected with a construct encoding GFP bearing the C-terminal peroxisomal targeting signal 1. Real time imaging revealed that moving peroxisomes interacted with each other and were engaged in transient contacts, and at higher magnification, tubular peroxisomes appeared to form a peroxisomal reticulum. Local remodeling of these structures could be observed involving the formation and detachment of tubular processes that interconnected adjacent organelles. Inhibition of cytoplasmic dynein based motility by overexpression of the dynactin subunit, dynamitin (p50), inhibited the movement of peroxisomes in vivo and interfered with the reestablishment of a uniform distribution of peroxisomes after recovery from nocodazole treatment. Isolated peroxisomes moved in vitro along microtubules in the presence of a microtubule motor fraction. Our data reveal that peroxisomal behavior in vivo is significantly more dynamic and interactive than previously thought and suggest a role for the dynein/dynactin motor in peroxisome motility.

The Drosophila fusome, a germline-specific organelle, contains membrane skeletal proteins and functions in cyst formation

Development ◽  
1994 ◽  
Vol 120 (4) ◽  
pp. 947-956 ◽  
Author(s):  
H. Lin ◽  
L. Yue ◽  
A.C. Spradling
Keyword(s):  
Cyst Formation ◽  
Nurse Cells ◽  
Germline Stem Cells ◽  
Cytoplasmic Structure ◽  
Ring Canals ◽  
Alpha Spectrin ◽  
Polarized Transport ◽  
Form Ring ◽  
Molecular Components ◽  
Number Of Cells

Oogenesis in Drosophila takes place within germline cysts that support polarized transport through ring canals interconnecting their 15 nurse cells and single oocyte. Developing cystocytes are spanned by a large cytoplasmic structure known as the fusome that has been postulated to help form ring canals and determine the pattern of nurse cell-oocyte interconnections. We identified the adducin-like hts product and alpha-spectrin as molecular components of fusomes, discovered a related structure in germline stem cells and documented regular associations between fusomes and cystocyte centrosomes. hts mutations completely eliminated fusomes, causing abnormal cysts containing a reduced number of cells to form. Our results imply that Drosophila fusomes are required for ovarian cyst formation and suggest that membrane skeletal proteins regulate cystocyte divisions.

The microtubule motor cytoplasmic dynein is required for spindle orientation during germline cell divisions and oocyte differentiation in Drosophila

Development ◽  
1997 ◽  
Vol 124 (12) ◽  
pp. 2409-2419 ◽  
Author(s):  
M. McGrail ◽  
T.S. Hays
Keyword(s):  
Asymmetric Cell Division ◽  
Fruit Fly ◽  
Cytoplasmic Dynein ◽  
Spindle Orientation ◽  
Chain Gene ◽  
Animal Development ◽  
Dynein Motor ◽  
Microtubule Motor ◽  
Two Stages ◽  
Oocyte Differentiation

During animal development cellular differentiation is often preceded by an asymmetric cell division whose polarity is determined by the orientation of the mitotic spindle. In the fruit fly, Drosophila melanogaster, the oocyte differentiates in a 16-cell syncytium that arises from a cystoblast which undergoes 4 synchronous divisions with incomplete cytokinesis. During these divisions, spindle orientation is highly ordered and is thought to impart a polarity to the cyst that is necessary for the subsequent differentiation of the oocyte. Using mutations in the Drosophila cytoplasmic dynein heavy chain gene, Dhc64C, we show that cytoplasmic dynein is required at two stages of oogenesis. Early in oogenesis, dynein mutations disrupt spindle orientation in dividing cysts and block oocyte determination. The localization of dynein in mitotic cysts suggests spindle orientation is mediated by the microtubule motor cytoplasmic dynein. Later in oogenesis, dynein function is necessary for proper differentiation, but does not appear to participate in morphogen localization within the oocyte. These results provide evidence for a novel developmental role for the cytoplasmic dynein motor in cellular determination and differentiation.

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