Amini, Rana et al. (2015) International Worm Meeting "Biogenesis in the C. elegans germline syncytium: from nucleation to maturation ."

Labbe, Jean-Claude, Amini, Rana, St-Pierre-See, Alexandre

[International Worm Meeting, 2015]

Cytokinesis consists in the physical separation of the two daughter cells, after mitosis. However, during development of certain tissues, mitotic division is followed by incomplete cytokinesis, giving rise to interconnected cells in a shared cytoplasm, or syncytium. Syncytial structures have been repeatedly described in female and male germlines in species ranging from insects to humans. Nevertheless, the mechanism of syncytium formation is poorly characterized. To understand this, we studied the syncytial C. elegans germline wherein germ cells (GCs) remain connected via stable intercellular bridges, also known as GC bridges. Using live-cell imaging, we found that ANI-1, ANI-2, NMY-2, UNC-59Septin, CYK-7 (a novel midbody component), ZEN-4Mklp1 are all stable components of GC bridges while RHO-1RhoA and ECT-2RhoGEF have a cortical localization throughout germline development from the L1 stage to adulthood. All GCs originate from the germline blastomere P4 that divides into the two primordial germ cells (PGCs) during embryogenesis. To understand the mechanism of syncytium formation, we monitored the dynamics of P4 cytokinesis. We found that P4, similar to its somatic neighbors, goes normally through the first phase of cytokinesis, also known as cytoplasmic isolation. Interestingly however, unlike somatic blastomeres, P4 fails to complete the second phase of cytokinesis wherein the midbody (MB) is released from the cell-cell boundary. While the MBs of somatic cells disappear soon after their cortical release, the MB connecting the two PGCs remains tightly associated to the cortex throughout embryogenesis. Our findings support a model in which incomplete cytokinesis of the P4 blastomere results in persistence of the MB between the two PGCs, thus forming a stable bridge that, in turn, promotes syncytium biogenesis.

Amini, Rana et al. (2013) International Worm Meeting "Anillin promotes syncytial organization and maintenance of the C. elegans germline."

Maddox, Amy S., Zetka, Monique, Labella, Sara, Chartier, Nicolas T., Amini, Rana, Labbe, Jean-Claude

[International Worm Meeting, 2013]

Cytokinesis, the last step of cell division, is required for the physical separation of the two daughter cells. During certain developmental stages however, incomplete cytokinesis gives rise to interconnected cells, forming a syncytium. The C. elegans adult germline is a syncytial organ enriched in ANI-2, a short homolog of the conserved scaffolding protein anillin, which controls cytokinesis. ANI-2 is present from the onset of germ cell specification and is enriched in intercellular syncytial bridges, suggesting that it might promote germline syncytium formation in C. elegans. To investigate this possibility, we developed a light microscopy-based assay to analyze the timing of syncytium formation during gonad development. We found that syncytial organization of the germline occurs progressively during larval development and completes shortly before animals enter the adult stage. Interestingly, while the germline of ani-2(-/-) mutants lacks defined syncytial openings, gonad development proceeds normally until animals reach the adult stage, when cytoplasmic partitions regress and germ cells progressively become polynucleated. This stage of gonad development is characterized by the initiation of cytoplasmic flows that promote oocyte growth, suggesting that ANI-2 is required to stabilize intercellular bridges when these flows initiate. In support of this, we did not observe severe multinucleation in male gonads or in tumorous germlines, which both lack cytoplasmic flows. Furthermore, time-lapse analysis of the proximal gonad at fertilization revealed that ANI-2 could confer elastic-like properties to the gonad, indicating that it can compensate for mechanical stress. We propose a model in which ANI-2 is required to promote syncytial organization of the germline and that its presence becomes essential to compensate for the mechanical stress resulting from cytoplasmic flows at the onset of oogenesis.

Herrmann, Audrey et al. (2019) International Worm Meeting "Regulation of Caenorhabditis elegans primordial germ cell abscission."

Amini, Rana, Labbe, Jean-Claude, Herrmann, Audrey, Goupil, Eugenie

[International Worm Meeting, 2019]

Cytokinesis occurs after mitosis, when the two daughter cells are physically separated. The last step of cytokinesis, termed abscission, involves numerous conserved regulators that coordinate all processes enabling resolution of the transient intercellular bridge, including midbody processing, cytoskeletal rearrangements membrane trafficking and scission. Certain cell types however undergo incomplete cytokinesis, generating cells that remain connected by a stable intercellular bridge, thus forming a syncytium. Stable intercellular bridges are a conserved feature during metazoan gametogenesis. In dividing mouse spermatocytes for instance, the protein TEX14 inhibits the recruitment of ESCRT-I regulators at the midbody ring, leading to a block of abscission and stabilization of the intercellular bridge. TEX14 is only found in vertebrates however, suggesting that this molecular mechanism of cytokinesis incompletion and syncytium formation is not conserved in most metazoans. The C. elegans syncytial gonad represents a relevant model to study how differential regulation of cytokinesis impacts syncytial organization. In C. elegans, all germ cells originate from the unique embryonic blastomere, P4, which divides incompletely to form two primordial germ cells that remain stably interconnected, Z2 and Z3. We hypothesize that an active mechanism prevents the completion of P4 cytokinesis during embryogenesis, and that this is required for proper syncytiogenesis of the larval and adult gonad. To define this mechanism and identify the step where abscission is blocked, we are using live-imaging approaches to monitor abscission regulators and compare their dynamics in P4 to those in neighboring somatic cells, which undergo complete cytokinesis. We found that the kinetics of Aurora B and midbody microtubules processing are identical in P4 and somatic cells. This indicates that the block of abscission in P4 is unlikely to be due to a defect in midbody maturation. We also found that the ESCRT-I regulator TSG-101 is loaded at the midbody ring with comparable timing in P4 and somatic cells, indicating that the mechanism of abscission block is different from that described in mouse spermatocytes. We are currently monitoring the dynamics of regulators that act at later steps in the process. Our work will help to define the mechanisms that enable differential cytokinetic regulation during gonad development.