Tyrosine phosphorylation of WIP releases bound WASP and impairs podosome assembly in macrophages

J Cell Sci. 2015 Jan 15;128(2):251-65. doi: 10.1242/jcs.154880. Epub 2014 Nov 20.

Abstract

Podosomes are integrin-containing adhesion structures commonly found in migrating leukocytes of the monocytic lineage. The actin cytoskeletal organisation of podosomes is based on a WASP- and Arp2/3-mediated mechanism. WASP also associates with a second protein, WIP (also known as WIPF1), and they co-localise in podosome cores. Here, we report for the first time that WIP can be phosphorylated on tyrosine residues and that tyrosine phosphorylation of WIP is a trigger for release of WASP from the WIP-WASP complex. Using a knockdown approach together with expression of WIP phosphomimics, we show that in the absence of WIP-WASP binding, cellular WASP is rapidly degraded, leading to disruption of podosomes and a failure of cells to degrade an underlying matrix. In the absence of tyrosine phosphorylation, the WIP-WASP complex remains intact and podosome lifetimes are extended. A screen of candidate kinases and inhibitor-based assays identified Bruton's tyrosine kinase (Btk) as a regulator of WIP tyrosine phosphorylation. We conclude that tyrosine phosphorylation of WIP is a crucial regulator of WASP stability and function as an actin-nucleation-promoting factor.

Keywords: Actin cytoskeleton; Macrophages; Matrix degradation; Podosome; Tyrosine phosphorylation; WIP; WIPF1.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Actin Cytoskeleton / genetics
  • Actin Cytoskeleton / metabolism
  • Agammaglobulinaemia Tyrosine Kinase
  • Animals
  • Cytoskeletal Proteins / genetics
  • Cytoskeletal Proteins / metabolism*
  • Extracellular Matrix / genetics
  • Extracellular Matrix / metabolism*
  • Humans
  • Intracellular Signaling Peptides and Proteins / genetics
  • Intracellular Signaling Peptides and Proteins / metabolism*
  • Macrophages / metabolism
  • Phosphorylation / genetics
  • Podosomes / metabolism
  • Protein Binding
  • Protein-Tyrosine Kinases / genetics
  • Protein-Tyrosine Kinases / metabolism*
  • Tyrosine / metabolism
  • Wiskott-Aldrich Syndrome Protein / genetics
  • Wiskott-Aldrich Syndrome Protein / metabolism*

Substances

  • Cytoskeletal Proteins
  • Intracellular Signaling Peptides and Proteins
  • WIPF1 protein, human
  • Wiskott-Aldrich Syndrome Protein
  • Tyrosine
  • Protein-Tyrosine Kinases
  • Agammaglobulinaemia Tyrosine Kinase
  • BTK protein, human