Precise levels of the Drosophila adaptor protein Dreadlocks maintain the size and stability of germline ring canals.
Ontology highlight
ABSTRACT:
SUBMITTER: Stark K
PROVIDER: S-EPMC8106954 | biostudies-literature |
REPOSITORIES: biostudies-literature
Similar Datasets
Project description:Intercellular bridges are conserved structures that allow neighboring cells to exchange cytoplasmic material; defects in intercellular bridges can lead to infertility in many organisms. Here, we use the Drosophila egg chamber to study the mechanisms that regulate intercellular bridges. Within the developing egg chamber, the germ cells (15 nurse cells and 1 oocyte) are connected to each other through intercellular bridges called ring canals, which expand over the course of oogenesis to support the transfer of materials from the nurse cells to the oocyte. The ring canals are enriched in actin and actin binding proteins, and many proteins have been identified that localize to the germline ring canals and control their expansion and stability. Here, we demonstrate a novel role for the Ste20 family kinase, Misshapen (Msn), in regulation of the size of the germline ring canals. Msn localizes to ring canals throughout most of oogenesis, and depletion of Msn led to the formation of larger ring canals. Over-expression of Msn decreased ring canal diameter, and expression of a membrane tethered form of Msn caused ring canal detachment and nurse cell fusion. Altering the levels or localization of Msn also led to changes in the actin cytoskeleton and altered the localization of E-cadherin, which suggests that Msn could be indirectly limiting ring canal size by altering the structure or dynamics of the actin cytoskeleton and/or adherens junctions.
| S-EPMC6082181 | biostudies-literature
Project description:The Drosophila Kelch protein is required to organize the ovarian ring canal cytoskeleton. Kelch binds and cross-links F-actin in vitro, and it also functions with Cullin 3 (Cul3) as a component of a ubiquitin E3 ligase. How these two activities contribute to cytoskeletal remodeling in vivo is not known. We used targeted mutagenesis to investigate the mechanism of Kelch function. We tested a model in which Cul3-dependent degradation of Kelch is required for its function, but we found no evidence to support this hypothesis. However, we found that mutant Kelch deficient in its ability to interact with Cul3 failed to rescue the kelch cytoskeletal defects, suggesting that ubiquitin ligase activity is the principal activity required in vivo. We also determined that the proteasome is required with Kelch to promote the ordered growth of the ring canal cytoskeleton. These results indicate that Kelch organizes the cytoskeleton in vivo by targeting a protein substrate for degradation by the proteasome.
| S-EPMC4649639 | biostudies-literature
Project description:Although intercellular bridges resulting from incomplete cytokinesis were discovered in somatic Drosophila tissues decades ago, the impact of these structures on intercellular communication and tissue biology is largely unknown. In this work, we demonstrate that the ~250-nanometer-diameter somatic ring canals permit diffusion of cytoplasmic contents between connected cells and across mitotic clone boundaries and enable the equilibration of protein between transcriptionally mosaic follicle cells in the Drosophila ovary. We obtained similar, although more restricted, results in the larval imaginal discs. Our work illustrates the lack of cytoplasmic autonomy in these tissues and suggests a role for somatic ring canals in promoting homogeneous protein expression within the tissue.
| S-EPMC3819220 | biostudies-literature
Project description:Orbit, a Drosophila ortholog of microtubule plus-end enriched protein CLASP, plays an important role in many developmental processes involved in microtubule dynamics. Previous studies have shown that Orbit is required for asymmetric stem cell division and cystocyte divisions in germline cysts and for the development of microtubule networks that interconnect oocyte and nurse cells during oogenesis. Here, we examined the cellular localization of Orbit and its role in cyst formation during spermatogenesis. In male germline stem cells, distinct localization of Orbit was first observed on the spectrosome, which is a spherical precursor of the germline-specific cytoskeleton known as the fusome. In dividing stem cells and spermatogonia, Orbit was localized around centrosomes and on kinetochores and spindle microtubules. After cytokinesis, Orbit remained localized on ring canals, which are cytoplasmic bridges between the cells. Thereafter, it was found along fusomes, extending through the ring canal toward all spermatogonia in a cyst. Fusome localization of Orbit was not affected by microtubule depolymerization. Instead, our fluorescence resonance energy transfer experiments suggested that Orbit is closely associated with F-actin, which is abundantly found in fusomes. Surprisingly, F-actin depolymerization influenced neither fusome organization nor Orbit localization on the germline-specific cytoskeleton. We revealed that two conserved regions of Orbit are required for fusome localization. Using orbit hypomorphic mutants, we showed that the protein is required for ring canal formation and for fusome elongation mediated by the interaction of newly generated fusome plugs with the pre-existing fusome. The orbit mutation also disrupted ring canal clustering, which is essential for folding of the spermatogonia after cytokinesis. Orbit accumulates around centrosomes at the onset of spermatogonial mitosis and is required for the capture of one of the duplicated centrosomes onto the fusome. Moreover, Orbit is involved in the proper orientation of spindles towards fusomes during synchronous mitosis of spermatogonial cysts.
| S-EPMC3592921 | biostudies-literature
Project description:Intercellular bridges are an essential structural feature found in both germline and somatic cells throughout the animal kingdom. Because of their large size, the germline intercellular bridges, or ring canals, in the developing fruit fly egg chamber are an excellent model to study the formation, stabilization, and growth of these structures. Within the egg chamber, the germline ring canals connect 15 supporting nurse cells to the developing oocyte, facilitating the transfer of materials required for successful oogenesis. The ring canals are derived from a stalled actomyosin contractile ring; once formed, additional actin and actin-binding proteins are recruited to the ring to support the 20-fold growth that accompanies oogenesis. These behaviors provide a unique model system to study the actin regulators that control incomplete cytokinesis, intercellular bridge formation, and growth. By temporally controlling their expression in the germline, we have demonstrated that the Arp2/3 complex and the formin, Diaphanous (Dia), coordinately regulate ring canal size and growth throughout oogenesis. Dia is required for successful incomplete cytokinesis and the initial stabilization of the germline ring canals. Once ring canals have formed, the Arp2/3 complex and Dia cooperate to determine ring canal size and maintain stability. Our data suggest that nurse cells must maintain a precise balance between the activity of these two nucleators during oogenesis.
| S-EPMC7170765 | biostudies-literature
Project description:Intercellular bridges called "ring canals" (RCs) resulting from incomplete cytokinesis play an essential role in intercellular communication in somatic and germinal tissues. During Drosophila oogenesis, RCs connect the maturing oocyte to nurse cells supporting its growth. Despite numerous genetic screens aimed at identifying genes involved in RC biogenesis and maturation, how RCs anchor to the plasma membrane (PM) throughout development remains unexplained. In this study, we report that the clathrin adaptor protein 1 (AP-1) complex, although dispensable for the biogenesis of RCs, is required for the maintenance of the anchorage of RCs to the PM to withstand the increased membrane tension associated with the exponential tissue growth at the onset of vitellogenesis. Here we unravel the mechanisms by which AP-1 enables the maintenance of RCs' anchoring to the PM during size expansion. We show that AP-1 regulates the localization of the intercellular adhesion molecule E-cadherin and that loss of AP-1 causes the disappearance of the E-cadherin-containing adhesive clusters surrounding the RCs. E-cadherin itself is shown to be required for the maintenance of the RCs' anchorage, a function previously unrecognized because of functional compensation by N-cadherin. Scanning block-face EM combined with transmission EM analyses reveals the presence of interdigitated, actin- and Moesin-positive, microvilli-like structures wrapping the RCs. Thus, by modulating E-cadherin trafficking, we show that the sustained E-cadherin-dependent adhesion organizes the microvilli meshwork and ensures the proper attachment of RCs to the PM, thereby counteracting the increasing membrane tension induced by exponential tissue growth.
| S-EPMC4611665 | biostudies-literature
Project description:Animal germ cells communicate directly with each other during gametogenesis through intercellular bridges, often called ring canals (RCs), that form as a consequence of incomplete cytokinesis during cell division. Developing germ cells in Drosophila have an additional specialized organelle connecting the cells called the fusome. Ring canals and the fusome are required for fertility in Drosophila females, but little is known about their roles during spermatogenesis. With live imaging, we directly observe the intercellular movement of GFP and a subset of endogenous proteins through RCs during spermatogenesis, from two-cell diploid spermatogonia to clusters of 64 post-meiotic haploid spermatids, demonstrating that RCs are stable and open to intercellular traffic throughout spermatogenesis. Disruption of the fusome, a large cytoplasmic structure that extends through RCs and is important during oogenesis, had no effect on spermatogenesis or male fertility under normal conditions. Our results reveal that male germline RCs allow the sharing of cytoplasmic information that might play a role in quality control surveillance during sperm development.
| S-EPMC7687857 | biostudies-literature
Project description:Mitochondrial reactive oxygen species (ROS) regulate a variety of biological processes by networking with signal transduction pathways to maintain homeostasis and support adaptation to stress. In this capacity, ROS have been shown to promote the differentiation of progenitor cells, including mammalian embryonic and hematopoietic stem cells and Drosophila hematopoietic progenitors (prohemocytes). However, many questions remain about how ROS alter the regulatory machinery to promote progenitor differentiation. Here, we provide evidence for the hypothesis that ROS reduce E-cadherin levels to promote Drosophila prohemocyte differentiation. Specifically, we show that knockdown of the antioxidants, Superoxide dismutatase 2 and Catalase reduce E-cadherin protein levels prior to the loss of Odd-skipped-expressing prohemocytes. Additionally, over-expression of E-cadherin limits prohemocyte differentiation resulting from paraquat-induced oxidative stress. Furthermore, two established targets of ROS, Enhancer of Polycomb and FOS, control the level of E-cadherin protein expression. Finally, we show that knockdown of either Superoxide dismutatase 2 or Catalase leads to an increase in the E-cadherin repressor, Serpent. As a result, antioxidants and targets of ROS can control E-cadherin protein levels, and over-expression of E-cadherin can ameliorate the prohemocyte response to oxidative stress. Collectively, these data strongly suggest that ROS promote differentiation by reducing E-cadherin levels. In mammalian systems, ROS promote embryonic stem cell differentiation, whereas E-cadherin blocks differentiation. However, it is not known if elevated ROS reduce E-cadherin to promote embryonic stem cell differentiation. Thus, our findings may have identified an important mechanism by which ROS promote stem/progenitor cell differentiation.
| S-EPMC4167200 | biostudies-literature
Project description:Germline cyst formation is essential for the propagation of many organisms including humans and flies. The cytoplasm of germline cyst cells communicate with each other directly via large intercellular bridges called ring canals. Ring canals are often derived from arrested contractile rings during incomplete cytokinesis. However how ring canal formation, maintenance and growth are regulated remains unclear. To better understand this process, we carried out an unbiased genetic screen in Drosophila melanogaster germ cells and identified multiple alleles of flapwing (flw), a conserved serine/threonine-specific protein phosphatase. Flw had previously been reported to be unnecessary for early D. melanogaster oogenesis using a hypomorphic allele. We found that loss of Flw leads to over-constricted nascent ring canals and subsequently tiny mature ring canals, through which cytoplasmic transfer from nurse cells to the oocyte is impaired, resulting in small, non-functional eggs. Flw is expressed in germ cells undergoing incomplete cytokinesis, completely colocalized with the Drosophila myosin binding subunit of myosin phosphatase (DMYPT). This colocalization, together with genetic interaction studies, suggests that Flw functions together with DMYPT to negatively regulate myosin activity during ring canal formation. The identification of two subunits of the tripartite myosin phosphatase as the first two main players required for ring canal constriction indicates that tight regulation of myosin activity is essential for germline cyst formation and reproduction in D. melanogaster and probably other species as well.
| S-EPMC3723691 | biostudies-literature
Project description:In this computational study, we report on the stability of cyclic phosphinyl radicals with an aim for a systematical assessment of stabilization effects. The radical stabilization energies (RSEs) were calculated using isodesmic reactions for a large number of carbocyclic radicals possessing different ring sizes and grades of unsaturation. In general, the RSE values range from -1.2 to -14.0 kcal·mol-1, and they show practically no correlation with the spin populations at the P-centers. The RSE values correlate with the reaction Gibbs free energies calculated for the dimerization of the studied simple radicals. Therefore, the more easily accessible RSE values offer a cost-effective estimation of global stability in a straightforward manner. To explore the effect of unsaturation on the RSE values, delocalization energies were determined using appropriate isodesmic reactions. Introducing unsaturations beside the P-center into the backbone of the rings leads to an additive increase in the magnitude of the delocalization energy (∼10, 20, and 30 kcal·mol-1, respectively, for radicals with one, two, and three C═C bonds in the conjugation). Parallelly, the spin populations at the P-centers also dwindle gradually by ∼0.1 e in the same order, indicating that the lone electron delocalizes over the π-system. Radicals containing exocyclic C═C π-bonds were also investigated, and all of these radicals have rather similar stabilities independently of the ring size, outlining the primary importance of the two exocyclic π-bonds in the conjugation. Among the radicals involved in our study, those with the best electronic stabilization are the unsaturated three-, five-, six-, and seven-membered rings containing the maximum number of conjugated vinyl fragments. The largest delocalization energy of 31.5 kcal·mol-1 and the lowest obtained spin population of 0.665 e were found for the fully unsaturated seven-membered radical (phosphepin derivative). Importantly, the electronic stabilization effects alone are insufficient for stabilizing the radicals in monomeric forms epitomized by the exothermic dimerization energies (-40 to -58 kcal·mol-1). Therefore, it is essential to apply sterically demanding bulky substituents on the α-C-atoms. Tweaking the steric congestion enabled us to propose radicals that are expected to be stable against dimerization and, consequently, may be realistic target species for synthetic investigations. The effects contributing to the stability of radicals having sterically encumbered substituents have also been explored.
| S-EPMC9583709 | biostudies-literature