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Principles of Polarity – Integrating genetic, biophysical and computational approaches to understand cell and tissue polarity

Deliverables

Analysis of Axin1, Axin2, and APC roles in liver and intestinal cell division

Analysis of Axin1, Axin2, and APC roles in liver and intestinal cell division involves a) cell-biological analysis of Axin1, Axin2, and APC null phenotypes in liver and intestinal cell division, and b) Mechanistic analysis of links between altered G2/M mitotic programme and increased cell division following Axin loss.

Quantitative analyses of local polarity perturbations on polarized growth

Quantitative analyses of local polarity perturbations on polarized growth will involve a) generation of strains which can generate locally increased concentrations of PS, PIP2, activated Cdc42 and activated Rho1, b) Quantitative analyses of local perturbation of PS, PIP2, activated Cdc42 and activated Rho1 levels on external mediated polarized growth and force generation.

Analysis of cell contact dependent transmission of Wnt-signal in seam cell PCP

Analysis of cell contact dependent transmission of Wnt-signal in seam cell PCP involves a) C. elegans clonal analysis of lineage-specific knock out mutants, b) analysis of cell-cell contact dependent transmission of the Wnt-signal in seam cell PCP, and c) identification of components critical for mitotic spindle and cleavage plane positioning.

Biochemical characterization of the Scrib polarity complex

Biochemical characterization of the Scrib polarity complex will involve a) identification of new partners of the mammalian polarity protein Scrib, b) biochemical characterization of the Scrib polarity complex in migrating cells and polarized epithelial cells, and c) characterization of the polarity function of Scrib partners in migrating cells and polarized epithelial cells.

Semi-automated culturing of organoids in a bioreactor

Semi-automated culturing of organoids in a bioreactor involves a) determining the correlation between processing conditions and organoid morphology and function, and b) bespoke hollow fibre bioreactor design for processing of organoids.

Mechanistic role of microtubules in polarization of 3D-cultured epithelial cells

Elucidating the mechanistic role of microtubules in polarization of 3D-cultured epithelial cells involves a) quantitative description of the role of microtubule-attachment complexes in cortical polarization in 3D breast epithelia acini and their modification during EMT and invasive growth in 3D, b) characterization of the biochemical connection between microtubule attachment complexes and the Scrib complex, c) quantitative characterization of the role of microtubule dynamics in polarity maintenance in 3D-cultured epithelial cells and the effects of microtubule-targeting agents on this process.

Role of phosphoinositides in control of cell polarity during collective migration

Characterizing the role of phosphoinositides in control of cell polarity during collective migration will involve a) characterization of phosphoinositide distribution during collective migration, b) characterisation of the role of phosphoinositides and more precisely of the lipid phosphatase PTEN in the control of cell polarity during collective migration, c) identification of signalling cascades controlled downstream of phosphoinositide signalling, and d) elucidation of the consequences of PTEN loss of functions in tumor spreading.

Mechanochemical model of PAR polarity patterning

1. Development of a mechanochemical model of PAR polarity patterning will involve a) simultaneous and complete midplane concentration and flowfields of markers of polarity and the actomyosin cortex under wild type and RNAi conditions, b) Identication of which markers of polarity control what activities of the cellular cytoskeleton, c) Development of a quantitative and predictive description of PAR polarity as a process of mechanochemical patterning formation in a regulated thin film active fluid.

Comprehensive, quantitative mathematical description of PAR polarity

Developing a comprehensive, quantitative mathematical description of PAR polarity will involve a) Defining mobility parameters for PAR network components, including diffusion coefficients and membrane association/dissociation rates, b) Defining binding parameters governing pairwise interactions between PAR com-ponents. c) Construction and analysis of a model of PAR polarity based on parameters in (a) and (b) to identify core design principles of PAR network.

Computational model of epithelial regeneration after wounding

Developing a computational model of epithelial regeneration after wounding involves a) live imaging of Drosophila wing disc regeneration and polarity re-establishment, b) quantification of changes in tissue mechanics after the wound and during the regeneration process, and c) building of a computational model of epithelial regeneration.

GTPase-regulator circuits essential for yeast and C. elegans polarity

Elucidation of GTPase-regulator circuits essential for yeast and C. elegans polarity will involve a) identification of GTPase-regulator circuits essential for polarized growth in yeast and polarity establishment and maintenance in C. elegans early embryos, b) characterization of the role of Arf/Rab GTPases in epithelial polarity and tissue organization, and c) generation of a number of C. elegans strains expressing GFP or mCherry tagged Arf and Rab GTPases and their effectors, and generation of specific point mutations therein using the CRISPR-Cas9 system.

Characterization of PAR, Crumbs and Scribble in C. elegans larval epithelia

Characterization of PAR, Crumbs and Scribble in C. elegans larval epithelia will involve a) generation of genetic null alleles of members of the Par, Crumbs, and Scribble complexes for which such alleles do not exist yet, and b) analysis of inactivation of members of these complexes on polarity in the intestine and seam cells.

Review article by the ESRs incorporating data obtained by PolarNet

Review article by the ESRs incorporating data obtained by PolarNet

Analysis of the mechanisms that maintain tissue integrity in seam epithelium

Analysis of the mechanisms that maintain tissue integrity in seam epithelium involves a) generation of strains in which apical-basal polarity, cell adhesion, tissue polarity, and various tumour suppressors can be disrupted specifically in the seam epithelium, and b) analysis of genetic interactions between these regulators, with the goal of causing seam cells to leave the epithelium.

Draft Periodic Report

compulsory deliverable

Progress Report

compulsory deliverable

Mechanisms of mechanotransduction during division in epithelial tissue

Elucidating the mechanisms of mechanotransduction during division in epithelial tissue involves a) integrated understanding of cytokinesis in epithelial tissue, b) uncovering the mechanisms of mechanotrandsuction during division in epithelial tissue, and c) force measurements in whole animal.

Super-resolution localization of polarity regulators at the plasma membrane

Super-resolution localization of polarity regulators at the plasma membrane will involve a) immunization of llama's with MDCK cells to generate a MDCK-immune library, b) production and purification of the membrane anchoring component for selection of specific nanobodies, c) selection and characterization of nanobodies binding to anchoring proteins, d) imaging of membrane anchoring of the spindle in 3D by super-resolution microscopy of 2D MDCK cell cultures, and e) quantitative characterization of the organization of the spindle orientation by anchoring complexes and the effect of EGF thereon.

Video highlighting importance of polarity for website & youtube

Video highlighting importance of polarity for website & youtube

Summaries of ESRs results placed on website by the ESRs

Summaries of ESRs results placed on website by the ESRs

Bi-weekly ‘Polarizing Cells’ blog entry on website

Bi-weekly ‘Polarizing Cells’ blog entry on website

Web presence

Web presence established.

Symposium 3

Closing symposium

Midterm review Meeting

compulsory deliverable

Summer School

Network Summer School

Symposium 2

Network Symposium

Symposium 1

Network symposium

ESRs evaluated for Doctorate degree application

All ESR are evaluated by the Supervisory Board for eligibility to apply for a Doctorate degree.

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Publications

Talin-KANK1 interaction controls the recruitment of cortical microtubule stabilizing complexes to focal adhesions

Author(s): Benjamin P Bouchet, Rosemarie E Gough, York-Christoph Ammon, Dieudonnée van de Willige, Harm Post, Guillaume Jacquemet, AF Maarten Altelaar, Albert JR Heck, Benjamin T Goult, Anna Akhmanova
Published in: eLife, Issue 5, 2016, ISSN 2050-084X
DOI: 10.7554/eLife.18124

Mesenchymal Cell Invasion Requires Cooperative Regulation of Persistent Microtubule Growth by SLAIN2 and CLASP1

Author(s): Benjamin P. Bouchet, Ivar Noordstra, Miranda van Amersfoort, Eugene A. Katrukha, York-Christoph Ammon, Natalie D. ter Hoeve, Louis Hodgson, Marileen Dogterom, Patrick W.B. Derksen, Anna Akhmanova
Published in: Developmental Cell, Issue 39/6, 2016, Page(s) 708-723, ISSN 1534-5807
DOI: 10.1016/j.devcel.2016.11.009

Regulated Activation of the PAR Polarity Network Ensures a Timely and Specific Response to Spatial Cues

Author(s): Jacob D. Reich, Lars Hubatsch, Rukshala Illukkumbura, Florent Peglion, Tom Bland, Nisha Hirani, Nathan W. Goehring
Published in: Current Biology, Issue 29/12, 2019, Page(s) 1911-1923.e5, ISSN 0960-9822
DOI: 10.1016/j.cub.2019.04.058

Anterior-enriched filopodia create the appearance of asymmetric membrane microdomains in polarizing C. elegans zygotes

Author(s): Nisha Hirani, Rukshala Illukkumbura, Tom Bland, Grégoire Mathonnet, Delphine Suhner, Anne-Cecile Reymann, Nathan W. Goehring
Published in: Journal of Cell Science, Issue 132/14, 2019, Page(s) jcs230714, ISSN 0021-9533
DOI: 10.1242/jcs.230714

Patterning and polarization of cells by intracellular flows

Author(s): Rukshala Illukkumbura, Tom Bland, Nathan W. Goehring
Published in: Current Opinion in Cell Biology, Issue 62, 2020, Page(s) 123-134, ISSN 0955-0674
DOI: 10.1016/j.ceb.2019.10.005

Wnt ligand and receptor patterning in the liver

Author(s): Eider Valle-Encinas, Trevor C. Dale
Published in: Current Opinion in Cell Biology, Issue 62, 2020, Page(s) 17-25, ISSN 0955-0674
DOI: 10.1016/j.ceb.2019.07.014

Intermediate filaments control collective migration by restricting traction forces and sustaining cell–cell contacts

Author(s): Chiara De Pascalis, Carlos Pérez-González, Shailaja Seetharaman, Batiste Boëda, Benoit Vianay, Mithila Burute, Cécile Leduc, Nicolas Borghi, Xavier Trepat, Sandrine Etienne-Manneville
Published in: The Journal of Cell Biology, Issue 217/9, 2018, Page(s) 3031-3044, ISSN 0021-9525
DOI: 10.1083/jcb.201801162

A cell-size threshold limits cell polarity and asymmetric division potential

Author(s): Lars Hubatsch, Florent Peglion, Jacob D. Reich, Nelio T. L. Rodrigues, Nisha Hirani, Rukshala Illukkumbura, Nathan W. Goehring
Published in: Nature Physics, Issue 15/10, 2019, Page(s) 1078-1085, ISSN 1745-2473
DOI: 10.1038/s41567-019-0601-x

Microtubule acetylation but not detyrosination promotes focal adhesion dynamics and astrocyte migration

Author(s): Bertille Bance, Shailaja Seetharaman, Cécile Leduc, Batiste Boëda, Sandrine Etienne-Manneville
Published in: Journal of Cell Science, Issue 132/7, 2019, Page(s) jcs225805, ISSN 0021-9533
DOI: 10.1242/jcs.225805

Microtubules at focal adhesions – a double-edged sword

Author(s): Shailaja Seetharaman, Sandrine Etienne-Manneville
Published in: Journal of Cell Science, Issue 132/19, 2019, Page(s) jcs232843, ISSN 0021-9533
DOI: 10.1242/jcs.232843

Oriented cell divisions in epithelia: from force generation to force anisotropy by tension, shape and vertices

Author(s): Eric Victor van Leen, Florencia di Pietro, Yohanns Bellaïche
Published in: Current Opinion in Cell Biology, Issue 62, 2020, Page(s) 9-16, ISSN 0955-0674
DOI: 10.1016/j.ceb.2019.07.013

Secretory Vesicle Clustering in Fungal Filamentous Cells Does Not Require Directional Growth

Author(s): Patrícia M. Silva, Charles Puerner, Agnese Seminara, Martine Bassilana, Robert A. Arkowitz
Published in: Cell Reports, Issue 28/8, 2019, Page(s) 2231-2245.e5, ISSN 2211-1247
DOI: 10.1016/j.celrep.2019.07.062

Systematic identification of recognition motifs for the hub protein LC8

Author(s): Nathan Jespersen, Aidan Estelle, Nathan Waugh, Norman E Davey, Cecilia Blikstad, York-Christoph Ammon, Anna Akhmanova, Ylva Ivarsson, David A Hendrix, Elisar Barbar
Published in: Life Science Alliance, Issue 2/4, 2019, Page(s) e201900366, ISSN 2575-1077
DOI: 10.26508/lsa.201900366

Force-Dependent Regulation of Talin–KANK1 Complex at Focal Adhesions

Author(s): Miao Yu, Shimin Le, York-Christoph Ammon, Benjamin T. Goult, Anna Akhmanova, Jie Yan
Published in: Nano Letters, Issue 19/9, 2019, Page(s) 5982-5990, ISSN 1530-6984
DOI: 10.1021/acs.nanolett.9b01732

Dissection of GTPase-activating proteins reveals functional asymmetry in the COPI coat of budding yeast

Author(s): Eric C. Arakel, Martina Huranova, Alejandro F. Estrada, E-Ming Rau, Anne Spang, Blanche Schwappach
Published in: Journal of Cell Science, Issue 132/16, 2019, Page(s) jcs232124, ISSN 0021-9533
DOI: 10.1242/jcs.232124

New insights into apical-basal polarization in epithelia

Author(s): Amalia Riga, Victoria G Castiglioni, Mike Boxem
Published in: Current Opinion in Cell Biology, Issue 62, 2020, Page(s) 1-8, ISSN 0955-0674
DOI: 10.1016/j.ceb.2019.07.017

The Golgi apparatus and cell polarity: Roles of the cytoskeleton, the Golgi matrix, and Golgi membranes

Author(s): Yamini Ravichandran, Bruno Goud, Jean-Baptiste Manneville
Published in: Current Opinion in Cell Biology, Issue 62, 2020, Page(s) 104-113, ISSN 0955-0674
DOI: 10.1016/j.ceb.2019.10.003

Multicellular scale front-to-rear polarity in collective migration

Author(s): Lavinia Capuana, Astrid Boström, Sandrine Etienne-Manneville
Published in: Current Opinion in Cell Biology, Issue 62, 2020, Page(s) 114-122, ISSN 0955-0674
DOI: 10.1016/j.ceb.2019.10.001

C. elegans Runx/CBFβ suppresses POP-1 TCF to convert asymmetric to proliferative division of stem cell-like seam cells

Author(s): Suzanne E. M. van der Horst, Janine Cravo, Alison Woollard, Juliane Teapal, Sander van den Heuvel
Published in: Development, Issue 146/22, 2019, Page(s) dev180034, ISSN 0950-1991
DOI: 10.1242/dev.180034