title: In vitro systems for the study of microtubule-based cell polarity in fission yeast.
abstract: Establishment of cell polarity is essential for processes such as growth and division. In fission yeast, as well as other species, polarity factors travel at the ends of microtubules to cortical sites where they associate with the membrane and subsequently maintain a polarized activity pattern despite their ability to diffuse in the membrane. In this chapter we present methods to establish an in vitro system that captures the essential features of this process. This bottom-up approach allows us to identify the minimal molecular requirements for microtubule-based cell polarity. We employ microfabrication techniques combined with surface functionalization to create rigid chambers with affinity for proteins, as well as microfluidic techniques to create and shape emulsion droplets with functionalized lipid boundaries. Preliminary results are shown demonstrating that a properly organized microtubule cytoskeleton can be confined to these confined spaces, and proteins traveling at the ends of growing microtubules can be delivered to their boundaries.
authors.affiliation: Department of Bionanoscience, Kavli Institute of Nanoscience, Delft University of Technology, Delft, The Netherlands.
authors.forename: Núria
authors.initials: N
authors.lastname: Taberner
authors.affiliation: FOM Institute AMOLF, Amsterdam, The Netherlands.
authors.forename: Andries
authors.initials: A
authors.lastname: Lof
authors.affiliation: Department of Bionanoscience, Kavli Institute of Nanoscience, Delft University of Technology, Delft, The Netherlands.
authors.forename: Sophie
authors.initials: S
authors.lastname: Roth
authors.affiliation: FOM Institute AMOLF, Amsterdam, The Netherlands.
authors.forename: Dimitry
authors.initials: D
authors.lastname: Lamers
authors.affiliation: FOM Institute AMOLF, Amsterdam, The Netherlands.
authors.forename: Hans
authors.initials: H
authors.lastname: Zeijlemaker
authors.affiliation: Department of Bionanoscience, Kavli Institute of Nanoscience, Delft University of Technology, Delft, The Netherlands.
authors.forename: Marileen
authors.initials: M
authors.lastname: Dogterom
mesh_terms: D008841:Actin Cytoskeleton / Q000502:physiology; D002462:Cell Membrane / Q000378:metabolism; D016764:Cell Polarity / Q000502:physiology*; D002846:Chromatography, Affinity / Q000379:methods; D006639:Histidine / Q000737:chemistry; D044085:Microfluidics / Q000379:methods*; D008869:Microtubule-Associated Proteins / Q000378:metabolism; D008870:Microtubules / Q000502:physiology*; D009532:Nickel / Q000737:chemistry; D009571:Nitrilotriacetic Acid / Q000737:chemistry; D009842:Oligopeptides / Q000737:chemistry; D010749:Phosphoprotein Phosphatases / Q000378:metabolism; D010766:Phosphorylation; D011485:Protein Binding / Q000502:physiology; D011494:Protein Kinases / Q000378:metabolism; D012568:Schizosaccharomyces / Q000254:growth & development / Q000502:physiology*; D029702:Schizosaccharomyces pombe Proteins / Q000378:metabolism; D064048:Spindle Pole Bodies / Q000378:metabolism
chemical_list: C471213:His-His-His-His-His-His; D008869:Microtubule-Associated Proteins; D009842:Oligopeptides; D029702:Schizosaccharomyces pombe Proteins; C432671:Tea1 protein, S pombe; C467709:Tea3 protein, S pombe; C501888:Tea4 protein, S pombe; C474999:mod5 protein, S pombe; D006639:Histidine; D009532:Nickel; D011494:Protein Kinases; C112212:Pom1 protein, S pombe; C502092:Dis2 protein, S pombe; D010749:Phosphoprotein Phosphatases; D009571:Nitrilotriacetic Acid
keywords: Cell polarity; Cortical patterns; Droplet storage; Fission yeast; Microfabrication; Microfluidics; Microtubules; Ni-NTA; Protein delivery; Surface functionalization