Gas6-Axl pathway: the role of redox-dependent association of Axl with nonmuscle myosin IIB.
In vascular smooth muscle cells, Axl is a key receptor tyrosine kinase, because it is upregulated in injury, increases migration and neointima formation, and is activated by reactive oxygen species. Reaction of glutathione with cysteine residues (termed "glutathiolation") is an important posttranslational redox modification that may alter protein activity and protein-protein interactions. To investigate the mechanisms by which reactive oxygen species increase Axl-dependent vascular smooth muscle cell function we assayed for glutathiolated proteins that associated with Axl in a redox-dependent manner. We identified glutathiolated nonmuscle myosin heavy chain (MHC)-IIB as a novel Axl interacting protein. This interaction was specific in that other myosins did not interact with Axl. The endogenous ligand for Axl, Gas6, increased production of reactive oxygen species in vascular smooth muscle cells and also increased the association of Axl with MHC-IIB. Antioxidants ebselen and N-acetylcysteine decreased the association of Axl with MHC-IIB in response to both Gas6 and reactive oxygen species. Blocking the Axl-MHC-IIB interaction with the specific myosin II inhibitor blebbistatin decreased phosphorylation of Axl and activation of extracellular signal-regulated kinase 1/2 and Akt. Association of MHC-IIB with Axl was increased in balloon-injured rat carotid vessels. Finally, expression of MHC-IIB was upregulated in the neointima of the carotid artery after balloon injury similar to upregulation of Axl protein expression, as shown in our previous studies. These results demonstrate a novel interaction between Axl and MHC-IIB in response to reactive oxygen species. This interaction provides a direct link between Axl and molecular motors crucial for directed cell migration, which may mediate increased migration in vascular dysfunction.
Cavet
Megan E
ME
Aab Cardiovascular Research Institute, University of Rochester, Box 679, Rochester, NY 14642, USA.
Smolock
Elaine M
EM
Menon
Prashanthi
P
Konishi
Atsushi
A
Korshunov
Vyacheslav A
VA
Berk
Bradford C
BC
D000818:Animals; D015153:Blotting, Western; D002340:Carotid Artery Diseases / Q000378:metabolism* / Q000473:pathology; D002478:Cells, Cultured; D004195:Disease Models, Animal; D007150:Immunohistochemistry; D047468:Immunoprecipitation; D036341:Intercellular Signaling Peptides and Proteins / Q000378:metabolism*; D008297:Male; D009131:Muscle, Smooth, Vascular / Q000378:metabolism* / Q000473:pathology; D024501:Nonmuscle Myosin Type IIB / Q000378:metabolism*; D010084:Oxidation-Reduction; D011518:Proto-Oncogene Proteins / Q000378:metabolism*; D051381:Rats; D017207:Rats, Sprague-Dawley; D017382:Reactive Oxygen Species / Q000378:metabolism; D020794:Receptor Protein-Tyrosine Kinases / Q000378:metabolism*; D000094963:Axl Receptor Tyrosine Kinase; D000099255:Growth Arrest-Specific Protein 6
D036341:Intercellular Signaling Peptides and Proteins; D024501:Nonmuscle Myosin Type IIB; D011518:Proto-Oncogene Proteins; D017382:Reactive Oxygen Species; D020794:Receptor Protein-Tyrosine Kinases; D000094963:Axl Receptor Tyrosine Kinase; D000099255:Growth Arrest-Specific Protein 6