title: LeftyA decreases Actin Polymerization and Stiffness in Human Endometrial Cancer Cells.
abstract: LeftyA, a cytokine regulating stemness and embryonic differentiation, down-regulates cell proliferation and migration. Cell proliferation and motility require actin reorganization, which is under control of ras-related C3 botulinum toxin substrate 1 (Rac1) and p21 protein-activated kinase 1 (PAK1). The present study explored whether LeftyA modifies actin cytoskeleton, shape and stiffness of Ishikawa cells, a well differentiated endometrial carcinoma cell line. The effect of LeftyA on globular over filamentous actin ratio was determined utilizing Western blotting and flow cytometry. Rac1 and PAK1 transcript levels were measured by qRT-PCR as well as active Rac1 and PAK1 by immunoblotting. Cell stiffness (quantified by the elastic modulus), cell surface area and cell volume were studied by atomic force microscopy (AFM). As a result, 2 hours treatment with LeftyA (25 ng/ml) significantly decreased Rac1 and PAK1 transcript levels and activity, depolymerized actin, and decreased cell stiffness, surface area and volume. The effect of LeftyA on actin polymerization was mimicked by pharmacological inhibition of Rac1 and PAK1. In the presence of the Rac1 or PAK1 inhibitor LeftyA did not lead to significant further actin depolymerization. In conclusion, LeftyA leads to disruption of Rac1 and Pak1 activity with subsequent actin depolymerization, cell softening and cell shrinkage.
authors.affiliation: Department of Cardiology, Vascular Medicine and Physiology, Eberhard Karls University of Tübingen, Germany.
authors.forename: Madhuri S
authors.initials: MS
authors.lastname: Salker
authors.affiliation: Institute of Applied Physics, Eberhard Karls University of Tübingen, Germany.
authors.forename: Nicolas
authors.initials: N
authors.lastname: Schierbaum
authors.affiliation: Department of Cardiology, Vascular Medicine and Physiology, Eberhard Karls University of Tübingen, Germany.
authors.forename: Nour
authors.initials: N
authors.lastname: Alowayed
authors.affiliation: Department of Cardiology, Vascular Medicine and Physiology, Eberhard Karls University of Tübingen, Germany.
authors.forename: Yogesh
authors.initials: Y
authors.lastname: Singh
authors.affiliation: Institute of Anatomy Eberhard Karls University of Tübingen, Germany.
authors.forename: Andreas F
authors.initials: AF
authors.lastname: Mack
authors.affiliation: Department of Biochemistry, University of Crete Medical School, Heraklion, Greece.
authors.forename: Christos
authors.initials: C
authors.lastname: Stournaras
authors.affiliation: Institute of Applied Physics, Eberhard Karls University of Tübingen, Germany.
authors.forename: Tilman E
authors.initials: TE
authors.lastname: Schäffer
authors.affiliation: Department of Cardiology, Vascular Medicine and Physiology, Eberhard Karls University of Tübingen, Germany.
authors.forename: Florian
authors.initials: F
authors.lastname: Lang
mesh_terms: D000199:Actins / Q000378:metabolism*; D048430:Cell Shape; D055119:Elastic Modulus; D016889:Endometrial Neoplasms / Q000378:metabolism / Q000473:pathology*; D005260:Female; D006801:Humans; D055458:Left-Right Determination Factors / Q000378:metabolism*; D058105:Polymerization; D012333:RNA, Messenger / Q000235:genetics; D014407:Tumor Cells, Cultured; D054462:p21-Activated Kinases / Q000235:genetics; D020830:rac1 GTP-Binding Protein / Q000235:genetics
chemical_list: D000199:Actins; C411603:LEFTY2 protein, human; D055458:Left-Right Determination Factors; C107953:RAC1 protein, human; D012333:RNA, Messenger; C515903:PAK1 protein, human; D054462:p21-Activated Kinases; D020830:rac1 GTP-Binding Protein