No description available.

Anillin is a highly conserved multi-domain cytoskeletal protein that provides a spatial and temporal scaffold for contractile ring proteins to ensure successful cytokinesis. We have looked at the temporal order of anillin and septin recruitment to the cleavage furrow using time-lapse microscopy and found that anillin localizes to the furrow in early anaphase while septins appear there later in an anillin-dependent manner. We also characterized the effect of anillin depletion in different cell lines and observed that septins and myosin delocalize in the absence of anillin in Tet-ON HeLa, AD293 and ARPE-19 cells but not in wild type HeLa cells. Asymmetric furrow formation was also investigated using the epithelial cell model: MDCK cells. Depletion of anillin and SEPT9 in MDCK cells was achieved using lentivirus shRNA constructs and this revealed that anillin or SEPT9 depletion did not affect asymmetric cytokinesis, although localization of SEPT 9 was affected by anillin depletion.

Anillin is a highly conserved multi-domain cytoskeletal protein that provides a spatial and temporal scaffold for contractile ring proteins to ensure successful cytokinesis. We have looked at the temporal order of anillin and septin recruitment to the cleavage furrow using time-lapse microscopy and found that anillin localizes to the furrow in early anaphase while septins appear there later in an anillin-dependent manner. We also characterized the effect of anillin depletion in different cell lines and observed that septins and myosin delocalize in the absence of anillin in Tet-ON HeLa, AD293 and ARPE-19 cells but not in wild type HeLa cells. Asymmetric furrow formation was also investigated using the epithelial cell model: MDCK cells. Depletion of anillin and SEPT9 in MDCK cells was achieved using lentivirus shRNA constructs and this revealed that anillin or SEPT9 depletion did not affect asymmetric cytokinesis, although localization of SEPT 9 was affected by anillin depletion.

Anillin is a highly conserved multi-domain cytoskeletal protein that provides a spatial and temporal scaffold for contractile ring proteins to ensure successful cytokinesis. We have looked at the temporal order of anillin and septin recruitment to the cleavage furrow using time-lapse microscopy and found that anillin localizes to the furrow in early anaphase while septins appear there later in an anillin-dependent manner. We also characterized the effect of anillin depletion in different cell lines and observed that septins and myosin delocalize in the absence of anillin in Tet-ON HeLa, AD293 and ARPE-19 cells but not in wild type HeLa cells. Asymmetric furrow formation was also investigated using the epithelial cell model: MDCK cells. Depletion of anillin and SEPT9 in MDCK cells was achieved using lentivirus shRNA constructs and this revealed that anillin or SEPT9 depletion did not affect asymmetric cytokinesis, although localization of SEPT 9 was affected by anillin depletion.

Anillin is a highly conserved multi-domain cytoskeletal protein that provides a spatial and temporal scaffold for contractile ring proteins to ensure successful cytokinesis. We have looked at the temporal order of anillin and septin recruitment to the cleavage furrow using time-lapse microscopy and found that anillin localizes to the furrow in early anaphase while septins appear there later in an anillin-dependent manner. We also characterized the effect of anillin depletion in different cell lines and observed that septins and myosin delocalize in the absence of anillin in Tet-ON HeLa, AD293 and ARPE-19 cells but not in wild type HeLa cells. Asymmetric furrow formation was also investigated using the epithelial cell model: MDCK cells. Depletion of anillin and SEPT9 in MDCK cells was achieved using lentivirus shRNA constructs and this revealed that anillin or SEPT9 depletion did not affect asymmetric cytokinesis, although localization of SEPT 9 was affected by anillin depletion.

Adherens junctions (AJs) and tight junctions (TJs) are characteristic features of differentiated epithelial cells and are critical for regulation of epithelial barriers and cell polarity. Integrity and remodeling of epithelial junctions depend on their interactions with underlying actomyosin cytoskeleton. Anillin is a multifunctional scaffold able to interact with different cytoskeletal proteins including F-actin and Myosin II. This project aimed to investigate roles of anillin in regulating epithelial AJs and TJs. Using A549 human lung epithelial and DU145 human prostate epithelial cells, we demonstrated the anillin depletion-induced loss of AJs and TJs. This was accompanied by disorganization of perijunctional actomyosin belt and disruption of the adducin-based membrane skeleton that links actin filaments to the plasma membrane and epithelial junctions. Depletion of anillin decreased protein levels of γ-adducin and downregulation of γ-adducin mimicked effects anillin knockdown on AJ and TJ integrity. These findings suggest a novel role for anillin in the assembly of epithelial junctions.

No description available.

No description available.

During embryonal development, axons of newly differentiated neurons need to properly interconnect and create a functional neuronal network. To achieve this, the cell requires a growth cone. The growth cone is a highly dynamic structure at the end of growing axons that serves both as the navigator and the propeller. Crosstalk between actin and microtubules is vital for proper axonal pathfinding. But the exact mechanism of this cooperation remains unknown. This diploma thesis investigates the possible role of a candidate scaffolding protein called anillin in this process. Anillin has been studied in two human cell lines. SH-SY5Y neuroblastoma cell line was used for overexpression and siRNA knock-down experiments. Anillin overexpression led to perturbed neurite morphology and growth cone dynamics in SH-SY5Y cells, whereas cells with lower anillin expression had fewer neurites. Next, neurons differentiated from human iPSC (induced pluripotent stem cells) expressing endogenous fluorescently tagged anillin were studied. Local dynamic high concentration spots of anillin have been observed at the base of cell protrusions of differentiating neurons. These anillin flares appeared during cell migration, early neurite initiation, and in newly created growth cones. These results suggest that anillin plays a...

La cytokinèse est un processus minutieusement régulé par une structure corticale appelée l'anneau contractile d'actomyosine, sous le contrôle de la petite GTPase RhoA. La protéine d'échafaudage Anilline est un effecteur de RhoA et organise les différents éléments de l'anneau permettant sa fermeture. Via son N-terminus, l'Anilline interagit avec la Citron kinase Sticky, la myosine II et l'actine-F ce qui permet la stabilisation de l'anneau contractile et sa maturation en anneau du corps central ou «midbody». Via son domaine C-terminal, l'Anilline interagit avec la RhoA-GTP et les septines pour ancrer l'anneau à la membrane. De précédentes études du laboratoire ont montré que Sticky et les septines ont des actions opposées sur l'Anilline. Nous avons donc défini plusieurs ensembles d'interactions entre l'Anilline et les cytosquelettes d'actomyosine et de septines, qui possèdent des fonctions différentes voire opposées dans la cytokinèse: ce sont des réseaux cytosquelettiques. L'Anilline est ainsi impliquée dans la coordination de ces réseaux opposés et RhoA-dépendants. Nous devons encore déterminer si ces interactions de l'Anilline en N-ter et C-ter peuvent se produire en même temps ou si elles sont mutuellement exclusives pour coordonner les différents éléments cytosquelettiques pendant la fermeture de l'anneau contractile. Les cellules S2 de drosophile ont été utilisées pour déterminer comment l'Anilline coordonnait les réseaux cytosqulettiques de l'anneau contractile. Deux modèles ont été proposés : l'un où une seule molécule d'Anilline se lie simultanément aux réseaux en N-ter et C-ter; l'autre modèle suggère qu'une ou plusieurs populations d'Anilline coordonnent les réseaux de façon mutuellement exclusive. Pour distinguer entre ces deux modèles, des allèles de séparation de fonction de l'Anilline ont été testés : l'AnillinΔ1-5 qui n'interagit plus avec Sticky, ainsi que l'AnillinRBD* qui n'interagit plus avec Rho1-GTP et qui ne recrute plus les septines à l'anneau contractile. Des expériences de sauvetage suite à la déplétion de l'Anilline endogène ont été réalisées et les tentatives de division ont été captées par microscopie en temps réel. L'expression de chaque mutant individuellement menait à une fermeture de l’anneau décalée, ralentie et souvent incomplète. En revanche, la co-expression de l'AnillinΔ1-5 et AnillinRBD* en trans dans les mêmes cellules a restauré la cinétique normale de la fermeture de l'anneau. Ce résultat supporte le modèle des populations multiples d'Anilline. Cette étude avance significativement nos connaissances de l'organisation de l'anneau contractile qui gère la division de toutes cellules animales. / Cytokinesis is a process thoroughly regulated by a cortical structure called the actomyosine contractile ring, under the control of the RhoA GTPase. The scaffolding protein Anillin is a RhoA effector organizing the several elements of the ring, thus permitting its closure. The AnillinN-terminus interacts with the Citron kinase Sticky, Myosin II and F-actin to stabilize the contractile ring and drive its maturation to the midbody ring. The AnillinC-terminus interacts with the RhoA GTPase and the septins to anchor the ring to the membrane. Previous works revealed that Sticky retains Anillin while the septins shed Anillin from the ring. Therefore, Anillin is involved in opposed RhoA-dependent cytoskeletal sub-networks to generate or reduce the tension at the membrane, and their balance is necessary to improve the ring closure. This study aims to decipher the coordination between these opposed sub-networks. We proposed two models : either sub-networks on AnillinN-ter and AnillinC-ter are simultaneously organized by the same molecule of Anillin, or several pools of Anillin coordinate separately the opposed subnetworks. We generated and expressed several inducible Anillin mutants in drosophila S2 cells : AnillinΔ1-5 prevents the interaction with Sticky; AnillinRBD* does not interact with RhoA and perturbs the Anillo-septin assembly. The expression of each mutant individually delayed, slowed down and failed the ring closure. However, co-expression of single mutants in trans rescued the ring closure. Moreover, Sticky over-expression improved AnillinRBD* recruitment in the ring. These results support the multiple pools of Anillin model. This study improves our knowledge on the contractile ring organization, necessary to succeed cytokinesis in animal cells.