title: Supervillin couples myosin-dependent contractility to podosomes and enables their turnover.
abstract: Podosomes are actin-rich adhesion and invasion structures. Especially in macrophages, podosomes exist in two subpopulations, large precursors at the cell periphery and smaller podosomes (successors) in the cell interior. To date, the mechanisms that differentially regulate these subpopulations are largely unknown. Here, we show that the membrane-associated protein supervillin localizes preferentially to successor podosomes and becomes enriched at precursors immediately before their dissolution. Consistently, podosome numbers are inversely correlated with supervillin protein levels. Using deletion constructs, we find that the myosin II regulatory N-terminus of supervillin [SV(1-174)] is crucial for these effects. Phosphorylated myosin light chain (pMLC) localizes at supervillin-positive podosomes, and time-lapse analyses show that enrichment of GFP-supervillin at podosomes coincides with their coupling to contractile myosin-IIA-positive cables. We also show that supervillin binds only to activated myosin IIA, and a dysregulated N-terminal construct [SV(1-830)] enhances pMLC levels at podosomes. Thus, preferential recruitment of supervillin to podosome subpopulations might both require and induce actomyosin contractility. Using siRNA and pharmacological inhibition, we demonstrate that supervillin and myosin IIA cooperate to regulate podosome lifetime, podosomal matrix degradation and cell polarization. In sum, we show here that podosome subpopulations differ in their molecular composition and identify supervillin, in cooperation with myosin IIA, as a crucial factor in the regulation of podosome turnover and function.
authors.affiliation: Institut für Medizinische Mikrobiologie, Virologie und Hygiene, Universitätsklinikum Eppendorf, Hamburg, Germany.
authors.forename: Ridhirama
authors.initials: R
authors.lastname: Bhuwania
authors.forename: Susanne
authors.initials: S
authors.lastname: Cornfine
authors.forename: Zhiyou
authors.initials: Z
authors.lastname: Fang
authors.forename: Marcus
authors.initials: M
authors.lastname: Krüger
authors.forename: Elizabeth J
authors.initials: EJ
authors.lastname: Luna
authors.forename: Stefan
authors.initials: S
authors.lastname: Linder
mesh_terms: D000199:Actins / Q000378:metabolism; D000205:Actomyosin / Q000378:metabolism; D016764:Cell Polarity / Q000502:physiology; D022081:Cell Surface Extensions / Q000502:physiology / Q000648:ultrastructure*; D006801:Humans; D008565:Membrane Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D008840:Microfilament Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D009000:Monocytes / Q000166:cytology* / Q000502:physiology / Q000648:ultrastructure; D018994:Myosin Light Chains / Q000235:genetics / Q000378:metabolism; D024507:Nonmuscle Myosin Type IIA / Q000235:genetics / Q000378:metabolism*; D010766:Phosphorylation; D010957:Plasmids; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D034741:RNA, Small Interfering; D059008:Time-Lapse Imaging; D014162:Transfection
chemical_list: D000199:Actins; D008565:Membrane Proteins; D008840:Microfilament Proteins; D018994:Myosin Light Chains; D034741:RNA, Small Interfering; C109413:SVIL protein, human; D000205:Actomyosin; D024507:Nonmuscle Myosin Type IIA