Regulation of cell motility by tyrosine phosphorylated villin.
Temporal and spatial regulation of the actin cytoskeleton is vital for cell migration. Here, we show that an epithelial cell actin-binding protein, villin, plays a crucial role in this process. Overexpression of villin in doxycyline-regulated HeLa cells enhanced cell migration. Villin-induced cell migration was modestly augmented by growth factors. In contrast, tyrosine phosphorylation of villin and villin-induced cell migration was significantly inhibited by the src kinase inhibitor 4-amino-5-(4-chlorophenyl)-7-(t-butyl)pyrazolo[3,4-d]pyrimidine (PP2) as well as by overexpression of a dominant negative mutant of c-src. These data suggest that phosphorylation of villin by c-src is involved in the actin cytoskeleton remodeling necessary for cell migration. We have previously shown that villin is tyrosine phosphorylated at four major sites. To further investigate the role of tyrosine phosphorylated villin in cell migration, we used phosphorylation site mutants (tyrosine to phenylalanine or tyrosine to glutamic acid) in HeLa cells. We determined that tyrosine phosphorylation at residues 60, 81, and 256 of human villin played an essential role in cell migration as well as in the reorganization of the actin cytoskeleton. Collectively, these studies define how biophysical events such as cell migration are actuated by biochemical signaling pathways involving tyrosine phosphorylation of actin binding proteins, in this case villin.
Tomar
Alok
A
Department of Physiology, University of Tennessee Health Science Center, Memphis, TN 38163, USA.
Wang
Yaohong
Y
Kumar
Narendra
N
George
Sudeep
S
Ceacareanu
Bogdan
B
Hassid
Aviv
A
Chapman
Kenneth E
KE
Aryal
Ashish M
AM
Waters
Christopher M
CM
Khurana
Seema
S
D000199:Actins / Q000737:chemistry / Q000378:metabolism; D000256:Adenoviridae / Q000235:genetics; D001665:Binding Sites; D000081247:CSK Tyrosine-Protein Kinase; D002465:Cell Movement; D049109:Cell Proliferation; D003599:Cytoskeleton / Q000378:metabolism; D018076:DNA, Complementary / Q000378:metabolism; D005799:Genes, Dominant; D006367:HeLa Cells; D006801:Humans; D008840:Microfilament Proteins / Q000737:chemistry* / Q000502:physiology*; D008856:Microscopy, Fluorescence; D009154:Mutation; D010766:Phosphorylation; D011505:Protein-Tyrosine Kinases / Q000235:genetics; D011743:Pyrimidines / Q000494:pharmacology; D015398:Signal Transduction; D013997:Time Factors; D014162:Transfection; D014443:Tyrosine / Q000737:chemistry*; D019061:src-Family Kinases
C412373:AG 1879; D000199:Actins; D018076:DNA, Complementary; D008840:Microfilament Proteins; D011743:Pyrimidines; C026615:villin; D014443:Tyrosine; D011505:Protein-Tyrosine Kinases; D000081247:CSK Tyrosine-Protein Kinase; D019061:src-Family Kinases; C000631346:CSK protein, human