title: Effects of Plectin Depletion on Keratin Network Dynamics and Organization.
abstract: The keratin intermediate filament cytoskeleton protects epithelial cells against various types of stress and is involved in fundamental cellular processes such as signaling, differentiation and organelle trafficking. These functions rely on the cell type-specific arrangement and plasticity of the keratin system. It has been suggested that these properties are regulated by a complex cycle of assembly and disassembly. The exact mechanisms responsible for the underlying molecular processes, however, have not been clarified. Accumulating evidence implicates the cytolinker plectin in various aspects of the keratin cycle, i.e., by acting as a stabilizing anchor at hemidesmosomal adhesion sites and the nucleus, by affecting keratin bundling and branching and by linkage of keratins to actin filament and microtubule dynamics. In the present study we tested these hypotheses. To this end, plectin was downregulated by shRNA in vulvar carcinoma-derived A431 cells. As expected, integrin β4- and BPAG-1-positive hemidesmosomal structures were strongly reduced and cytosolic actin stress fibers were increased. In addition, integrins α3 and β1 were reduced. The experiments furthermore showed that loss of plectin led to a reduction in keratin filament branch length but did not alter overall mechanical properties as assessed by indentation analyses using atomic force microscopy and by displacement analyses of cytoplasmic superparamagnetic beads using magnetic tweezers. An increase in keratin movement was observed in plectin-depleted cells as was the case in control cells lacking hemidesmosome-like structures. Yet, keratin turnover was not significantly affected. We conclude that plectin alone is not needed for keratin assembly and disassembly and that other mechanisms exist to guarantee proper keratin cycling under steady state conditions in cultured single cells.
authors.affiliation: Institute of Molecular and Cellular Anatomy, RWTH Aachen University, Aachen, Germany.
authors.forename: Marcin
authors.initials: M
authors.lastname: Moch
authors.affiliation: Institute of Molecular and Cellular Anatomy, RWTH Aachen University, Aachen, Germany.
authors.forename: Reinhard
authors.initials: R
authors.lastname: Windoffer
authors.affiliation: Institute of Molecular and Cellular Anatomy, RWTH Aachen University, Aachen, Germany.
authors.forename: Nicole
authors.initials: N
authors.lastname: Schwarz
authors.affiliation: Institute of Molecular and Cellular Anatomy, RWTH Aachen University, Aachen, Germany.
authors.forename: Raphaela
authors.initials: R
authors.lastname: Pohl
authors.affiliation: Institute of Materials in Electrical Engineering 1, RWTH Aachen University, Aachen, Germany.
authors.forename: Andreas
authors.initials: A
authors.lastname: Omenzetter
authors.affiliation: Institute of Materials in Electrical Engineering 1, RWTH Aachen University, Aachen, Germany.
authors.forename: Uwe
authors.initials: U
authors.lastname: Schnakenberg
authors.affiliation: Institute of Imaging & Computer Vision, RWTH Aachen University, Aachen, Germany.
authors.forename: Fabian
authors.initials: F
authors.lastname: Herb
authors.affiliation: Institute of Imaging & Computer Vision, RWTH Aachen University, Aachen, Germany.
authors.forename: Kraisorn
authors.initials: K
authors.lastname: Chaisaowong
authors.affiliation: Institute of Imaging & Computer Vision, RWTH Aachen University, Aachen, Germany.
authors.forename: Dorit
authors.initials: D
authors.lastname: Merhof
authors.affiliation: Institute of Complex Systems, Forschungszentrum Jülich, Jülich, Germany.
authors.forename: Lena
authors.initials: L
authors.lastname: Ramms
authors.affiliation: Institute of Complex Systems, Forschungszentrum Jülich, Jülich, Germany.
authors.forename: Gloria
authors.initials: G
authors.lastname: Fabris
authors.affiliation: Institute of Complex Systems, Forschungszentrum Jülich, Jülich, Germany.
authors.forename: Bernd
authors.initials: B
authors.lastname: Hoffmann
authors.affiliation: Institute of Complex Systems, Forschungszentrum Jülich, Jülich, Germany.
authors.forename: Rudolf
authors.initials: R
authors.lastname: Merkel
authors.affiliation: Institute of Molecular and Cellular Anatomy, RWTH Aachen University, Aachen, Germany.
authors.forename: Rudolf E
authors.initials: RE
authors.lastname: Leube
mesh_terms: D008841:Actin Cytoskeleton / Q000378:metabolism; D000199:Actins / Q000378:metabolism; D002352:Carrier Proteins / Q000378:metabolism; D045744:Cell Line, Tumor; D002465:Cell Movement / Q000502:physiology; D003598:Cytoskeletal Proteins / Q000378:metabolism; D003599:Cytoskeleton / Q000378:metabolism; D000071821:Dystonin; D004847:Epithelial Cells / Q000378:metabolism; D022002:Hemidesmosomes / Q000378:metabolism; D006801:Humans; D039663:Integrin beta4 / Q000378:metabolism; D007381:Intermediate Filament Proteins / Q000378:metabolism; D007382:Intermediate Filaments / Q000378:metabolism; D015603:Keratinocytes / Q000378:metabolism; D007633:Keratins / Q000378:metabolism*; D009419:Nerve Tissue Proteins / Q000378:metabolism; D051190:Plectin / Q000378:metabolism*; D011485:Protein Binding / Q000502:physiology
chemical_list: D000199:Actins; D002352:Carrier Proteins; D003598:Cytoskeletal Proteins; C486268:DST protein, human; D000071821:Dystonin; D039663:Integrin beta4; D007381:Intermediate Filament Proteins; D009419:Nerve Tissue Proteins; D051190:Plectin; D007633:Keratins