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Endothelial cell retraction is induced by PAK2 monophosphorylation of myosin II

Q Zeng1, D Lagunoff, R Masaracchia

  • 1Department of Pathology, St Louis University School of Medicine St Louis, Missouri 63104-1028, USA.

Journal of Cell Science
|January 20, 2000
PubMed

Insights

p21-activated kinase 2 (PAK2) triggers endothelial cell retraction and actin cytoskeleton changes by directly activating myosin II. This involves monophosphorylation of the myosin II regulatory light chain, confirmed by inhibitor studies.

Area of Science:

  • Cell Biology
  • Molecular Biology
  • Biochemistry

Background:

  • The p21-activated kinase (PAK) family, including PAK1 and PAK2, are key regulators of the actin cytoskeleton, influenced by GTPases Rac1 and Cdc42.
  • While PAK1's role in cytoskeletal dynamics is established, PAK2's function in intact cells, particularly its interaction with myosin II, remains less understood.
  • Previous studies indicated PAK2's ability to monophosphorylate myosin II regulatory light chain and induce retraction in permeabilized endothelial cells.

Purpose of the Study:

  • To investigate the morphological and biochemical effects of constitutively active PAK2 on intact endothelial cells.
  • To elucidate the mechanism by which PAK2 induces cytoskeletal rearrangements and cell retraction.
  • To determine the specific role of myosin II activation in PAK2-mediated cellular changes.

Main Methods:

  • Microinjection of constitutively active PAK2 into intact endothelial cells.
  • Utilizing selective inhibitors: Staurosporine (PAK2 inhibitor), Butanedione monoxime (myosin ATPase inhibitor), and KT5926 (myosin light chain kinase inhibitor).
  • Analysis of actin cytoskeleton modifications, cell margin retraction, and myosin II regulatory light chain monophosphorylation.
  • Investigating the association between endogenous PAK2 and myosin II.

Main Results:

  • Microinjection of active PAK2 induced endothelial cell retraction and actin cytoskeleton reorganization.
  • A significant increase in myosin II regulatory light chain monophosphorylation was observed following PAK2 activation.
  • PAK2 inhibition by Staurosporine blocked PAK2-induced effects, while myosin ATPase inhibition by Butanedione monoxime also prevented these changes.
  • Myosin light chain kinase inhibition by KT5926 did not affect PAK2-induced cytoskeletal alterations.
  • Endogenous PAK2 was found to associate with myosin II.

Conclusions:

  • PAK2 directly activates myosin II through monophosphorylation of its regulatory light chain, leading to endothelial cell retraction.
  • The observed cytoskeletal rearrangements are dependent on myosin II activity, but not mediated by myosin light chain kinase.
  • These findings highlight a direct mechanism for PAK2 in regulating cell morphology and motility via myosin II activation.

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