scholarly journals Canoe binds RanGTP to promote PinsTPR/Mud-mediated spindle orientation

2011 ◽  
Vol 195 (3) ◽  
pp. 369-376 ◽  
Author(s):  
Brett Wee ◽  
Christopher A. Johnston ◽  
Kenneth E. Prehoda ◽  
Chris Q. Doe
Keyword(s):  
Asymmetric Cell Division ◽  
Scaffolding Protein ◽  
Spindle Orientation ◽  
Cell Cortex ◽  
Tetratricopeptide Repeat ◽  
Epithelial Tissue ◽  
Mitotic Apparatus ◽  
Tissue Integrity ◽  
Interaction Partners ◽  
Tpr Domain

Regulated spindle orientation maintains epithelial tissue integrity and stem cell asymmetric cell division. In Drosophila melanogaster neural stem cells (neuroblasts), the scaffolding protein Canoe (Afadin/Af-6 in mammals) regulates spindle orientation, but its protein interaction partners and mechanism of action are unknown. In this paper, we use our recently developed induced cell polarity system to dissect the molecular mechanism of Canoe-mediated spindle orientation. We show that a previously uncharacterized portion of Canoe directly binds the Partner of Inscuteable (Pins) tetratricopeptide repeat (TPR) domain. The Canoe–PinsTPR interaction recruits Canoe to the cell cortex and is required for activation of the PinsTPR-Mud (nuclear mitotic apparatus in mammals) spindle orientation pathway. We show that the Canoe Ras-association (RA) domains directly bind RanGTP and that both the CanoeRA domains and RanGTP are required to recruit Mud to the cortex and activate the Pins/Mud/dynein spindle orientation pathway.

Asymmetric cell division in fucoid algae: a role for cortical adhesions in alignment of the mitotic apparatus

2001 ◽  
Vol 114 (23) ◽  
pp. 4319-4328
Author(s):  
Sherryl R. Bisgrove ◽  
Darryl L. Kropf
Keyword(s):  
Cell Division ◽  
Asymmetric Cell Division ◽  
Cell Cortex ◽  
Mitotic Apparatus ◽  
Pharmacological Agents ◽  
Division Plane ◽  
Adhesion Sites ◽  
Spindle Poles ◽  
Pelvetia Compressa ◽  
Two Phases

The first cell division in zygotes of the fucoid brown alga Pelvetia compressa is asymmetric and we are interested in the mechanism controlling the alignment of this division. Since the division plane bisects the mitotic apparatus, we investigated the timing and mechanism of spindle alignments. Centrosomes, which give rise to spindle poles, aligned with the growth axis in two phases – a premetaphase rotation of the nucleus and centrosomes followed by a postmetaphase alignment that coincided with the separation of the mitotic spindle poles during anaphase and telophase. The roles of the cytoskeleton and cell cortex in the two phases of alignment were analyzed by treatment with pharmacological agents. Treatments that disrupted cytoskeleton or perturbed cortical adhesions inhibited pre-metaphase alignment and we propose that this rotational alignment is effected by microtubules anchored at cortical adhesion sites. Postmetaphase alignment was not affected by any of the treatments tested, and may be dependent on asymmetric cell morphology.

scholarly journals The Phosphatase PTPL1 Is Required for PTEN-Mediated Regulation of Apical Membrane Size

2018 ◽  
Vol 38 (12) ◽  
pp. e00102-18 ◽  
Author(s):  
Lucas J. M. Bruurs ◽  
Mirjam C. van der Net ◽  
Susan Zwakenberg ◽  
Axel K. M. Rosendahl Huber ◽  
Anneke Post ◽  
...  
Keyword(s):  
Brush Border ◽  
Gene Disruption ◽  
Apical Membrane ◽  
Cell Polarization ◽  
Cell Junctions ◽  
Cell Cortex ◽  
Epithelial Tissue ◽  
Intestinal Epithelial ◽  
Tissue Integrity ◽  
Short Palindromic Repeat

ABSTRACT PTEN is a tumor suppressor that is frequently lost in epithelial malignancies. A part of the tumor-suppressive properties of PTEN is attributed to its function in cell polarization and consequently its role in maintaining epithelial tissue integrity. However, surprisingly little is known about the function and regulation of PTEN during epithelial cell polarization. We used clustered regularly interspaced short palindromic repeat (CRISPR)/Cas9-mediated gene disruption to delete PTEN in intestinal epithelial Ls174T:W4 cells, which upon differentiation form a microvillus-covered apical membrane (brush border) on a part of the cell cortex, independent of cell-cell junctions. We show that loss of PTEN results in the formation of a larger brush border that, in a fraction of the cells, even spans the entire plasma membrane, revealing that PTEN functions in the regulation of apical membrane size. Depletion of the phosphatase PTPL1 resulted in a similar defect. PTPL1 interacts with PTEN, and this interaction is necessary for apical membrane enrichment of PTEN. Importantly, phosphatase activity of PTPL1 is not required, indicating that PTPL1 functions as an anchor protein in this process. Our work thus demonstrates a novel function for PTEN during cell polarization in controlling apical membrane size and identifies PTPL1 as a critical apical membrane anchor for PTEN in this process.

scholarly journals NuMA localization, stability, and function in spindle orientation involve 4.1 and Cdk1 interactions

2013 ◽  
Vol 24 (23) ◽  
pp. 3651-3662 ◽  
Author(s):  
Lindsey Seldin ◽  
Nicholas D. Poulson ◽  
Henry P. Foote ◽  
Terry Lechler
Keyword(s):  
Protein Complex ◽  
Fluorescence Recovery After Photobleaching ◽  
Force Generation ◽  
Spindle Orientation ◽  
Cell Cortex ◽  
Mitotic Apparatus ◽  
Productive Force ◽  
Asymmetric Divisions ◽  
And Function ◽  
Multilayered Epithelium

The epidermis is a multilayered epithelium that requires asymmetric divisions for stratification. A conserved cortical protein complex, including LGN, nuclear mitotic apparatus (NuMA), and dynein/dynactin, plays a key role in establishing proper spindle orientation during asymmetric divisions. The requirements for the cortical recruitment of these proteins, however, remain unclear. In this work, we show that NuMA is required to recruit dynactin to the cell cortex of keratinocytes. NuMA's cortical recruitment requires LGN; however, LGN interactions are not sufficient for this localization. Using fluorescence recovery after photobleaching, we find that the 4.1-binding domain of NuMA is important for stabilizing its interaction with the cell cortex. This is functionally important, as loss of 4.1/NuMA interaction results in spindle orientation defects, using two distinct assays. Furthermore, we observe an increase in cortical NuMA localization as cells enter anaphase. Inhibition of Cdk1 or mutation of a single residue in NuMA mimics this effect. NuMA's anaphase localization is independent of LGN and 4.1 interactions, revealing two distinct mechanisms responsible for NuMA cortical recruitment at different stages of mitosis. This work highlights the complexity of NuMA localization and reveals the importance of NuMA cortical stability for productive force generation during spindle orientation.

scholarly journals Cytoplasmic MTOCs control spindle orientation for asymmetric cell division in plants

2017 ◽  
Vol 114 (42) ◽  
pp. E8847-E8854 ◽  
Author(s):  
Ken Kosetsu ◽  
Takashi Murata ◽  
Moé Yamada ◽  
Momoko Nishina ◽  
Joanna Boruc ◽  
...  
Keyword(s):  
Cell Division ◽  
Asymmetric Cell Division ◽  
Physcomitrella Patens ◽  
De Novo ◽  
Critical Role ◽  
Fine Tuning ◽  
Spindle Orientation ◽  
Mitotic Apparatus ◽  
Division Plane ◽  
Division Axis

Proper orientation of the cell division axis is critical for asymmetric cell divisions that underpin cell differentiation. In animals, centrosomes are the dominant microtubule organizing centers (MTOC) and play a pivotal role in axis determination by orienting the mitotic spindle. In land plants that lack centrosomes, a critical role of a microtubular ring structure, the preprophase band (PPB), has been observed in this process; the PPB is required for orienting (before prophase) and guiding (in telophase) the mitotic apparatus. However, plants must possess additional mechanisms to control the division axis, as certain cell types or mutants do not form PPBs. Here, using live imaging of the gametophore of the moss Physcomitrella patens, we identified acentrosomal MTOCs, which we termed “gametosomes,” appearing de novo and transiently in the prophase cytoplasm independent of PPB formation. We show that gametosomes are dispensable for spindle formation but required for metaphase spindle orientation. In some cells, gametosomes appeared reminiscent of the bipolar MT “polar cap” structure that forms transiently around the prophase nucleus in angiosperms. Specific disruption of the polar caps in tobacco cells misoriented the metaphase spindles and frequently altered the final division plane, indicating that they are functionally analogous to the gametosomes. These results suggest a broad use of transient MTOC structures as the spindle orientation machinery in plants, compensating for the evolutionary loss of centrosomes, to secure the initial orientation of the spindle in a spatial window that allows subsequent fine-tuning of the division plane axis by the guidance machinery.

scholarly journals SLK-dependent activation of ERMs controls LGN–NuMA localization and spindle orientation

2014 ◽  
Vol 205 (6) ◽  
pp. 791-799 ◽  
Author(s):  
Mickael Machicoane ◽  
Cristina A. de Frutos ◽  
Jenny Fink ◽  
Murielle Rocancourt ◽  
Yannis Lombardi ◽  
...  
Keyword(s):  
Mitotic Spindle ◽  
Mammalian Cells ◽  
Molecular Level ◽  
Spindle Orientation ◽  
Cell Cortex ◽  
Mitotic Apparatus ◽  
Mitotic Entry ◽  
Spindle Microtubules ◽  
Force Generator

Mitotic spindle orientation relies on a complex dialog between the spindle microtubules and the cell cortex, in which F-actin has been recently implicated. Here, we report that the membrane–actin linkers ezrin/radixin/moesin (ERMs) are strongly and directly activated by the Ste20-like kinase at mitotic entry in mammalian cells. Using microfabricated adhesive substrates to control the axis of cell division, we found that the activation of ERMs plays a key role in guiding the orientation of the mitotic spindle. Accordingly, impairing ERM activation in apical progenitors of the mouse embryonic neocortex severely disturbed spindle orientation in vivo. At the molecular level, ERM activation promotes the polarized association at the mitotic cortex of leucine-glycine-asparagine repeat protein (LGN) and nuclear mitotic apparatus (NuMA) protein, two essential factors for spindle orientation. We propose that activated ERMs, together with Gαi, are critical for the correct localization of LGN–NuMA force generator complexes and hence for proper spindle orientation.

scholarly journals Specific polar subpopulations of astral microtubules control spindle orientation and symmetric neural stem cell division

eLife ◽  
2014 ◽  
Vol 3 ◽  
Author(s):  
Felipe Mora-Bermúdez ◽  
Fumio Matsuzaki ◽  
Wieland B Huttner
Keyword(s):  
Cell Division ◽  
Basal Cell ◽  
Mitotic Spindle ◽  
Asymmetric Cell Division ◽  
Spindle Cell ◽  
Spindle Orientation ◽  
Cell Cortex ◽  
Dividing Cells ◽  
Stem Cell Division

Mitotic spindle orientation is crucial for symmetric vs asymmetric cell division and depends on astral microtubules. Here, we show that distinct subpopulations of astral microtubules exist, which have differential functions in regulating spindle orientation and division symmetry. Specifically, in polarized stem cells of developing mouse neocortex, astral microtubules reaching the apical and basal cell cortex, but not those reaching the central cell cortex, are more abundant in symmetrically than asymmetrically dividing cells and reduce spindle orientation variability. This promotes symmetric divisions by maintaining an apico-basal cleavage plane. The greater abundance of apical/basal astrals depends on a higher concentration, at the basal cell cortex, of LGN, a known spindle-cell cortex linker. Furthermore, newly developed specific microtubule perturbations that selectively decrease apical/basal astrals recapitulate the symmetric-to-asymmetric division switch and suffice to increase neurogenesis in vivo. Thus, our study identifies a novel link between cell polarity, astral microtubules, and spindle orientation in morphogenesis.

scholarly journals KIF18B is a cell-type specific regulator of spindle orientation in the epidermis

2021 ◽  
pp. mbc.E21-06-0291
Author(s):  
Rebecca S. Moreci ◽  
Terry Lechler
Keyword(s):  
Cell Division ◽  
Hair Follicle ◽  
Cell Fate ◽  
Asymmetric Cell Division ◽  
Microtubule Dynamics ◽  
Spindle Orientation ◽  
Cell Cortex ◽  
Cell Fate Specification ◽  
Cell Fate Decisions ◽  
Fate Specification

Proper spindle orientation is required for asymmetric cell division and the establishment of complex tissue architecture. In the developing epidermis, spindle orientation requires a conserved cortical protein complex of LGN/NuMA/dynein-dynactin. However, how microtubule dynamics are regulated to interact with this machinery and properly position the mitotic spindle is not fully understood. Furthermore, our understanding of the processes that link spindle orientation during asymmetric cell division to cell fate specification in distinct tissue contexts remains incomplete. We report a role for the microtubule catastrophe factor KIF18B in regulating microtubule dynamics to promote spindle orientation in keratinocytes. During mitosis, KIF18B accumulates at the cell cortex, colocalizing with the conserved spindle orientation machinery. In vivo we find that KIF18B is required for oriented cell divisions within the hair placode, the first stage of hair follicle morphogenesis, but is not essential in the interfollicular epidermis. Disrupting spindle orientation in the placode, using mutations in either KIF18B or NuMA, results in aberrant cell fate marker expression of hair follicle progenitor cells. These data functionally link spindle orientation to cell fate decisions during hair follicle morphogenesis. Taken together, our data demonstrate a role for regulated microtubule dynamics in spindle orientation in epidermal cells. This work also highlights the importance of spindle orientation during asymmetric cell division to dictate cell fate specification. [Media: see text] [Media: see text]

scholarly journals KIF18B is a cell-type specific regulator of spindle orientation in the epidermis

2021 ◽  
Author(s):  
Rebecca S. Moreci ◽  
Terry Lechler
Keyword(s):  
Cell Division ◽  
Hair Follicle ◽  
Cell Fate ◽  
Asymmetric Cell Division ◽  
Microtubule Dynamics ◽  
Spindle Orientation ◽  
Cell Cortex ◽  
Cell Fate Specification ◽  
Cell Fate Decisions ◽  
Fate Specification

Proper spindle orientation is required for asymmetric cell division and the establishment of complex tissue architecture. In the developing epidermis, spindle orientation requires a conserved cortical protein complex of LGN/NuMA/dynein-dynactin. However, how microtubule dynamics are regulated to interact with this machinery and properly position the mitotic spindle is not fully understood. Furthermore, our understanding of the processes that link spindle orientation during asymmetric cell division to cell fate specification in distinct tissue contexts remains incomplete. We report a role for the microtubule catastrophe factor KIF18B in regulating microtubule dynamics to promote spindle orientation in keratinocytes. During mitosis, KIF18B accumulates at the cell cortex, colocalizing with the conserved spindle orientation machinery. In vivo we find that KIF18B is required for oriented cell divisions within the hair placode, the first stage of hair follicle morphogenesis, but is not essential in the interfollicular epidermis. Disrupting spindle orientation in the placode, using mutations in either KIF18B or NuMA, results in aberrant cell fate marker expression of hair follicle progenitor cells. These data functionally link spindle orientation to cell fate decisions during hair follicle morphogenesis. Taken together, our data demonstrate a role for regulated microtubule dynamics in spindle orientation in epidermal cells. This work also highlights the importance of spindle orientation during asymmetric cell division to dictate cell fate specification.

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