Integrin-dependent protein tyrosine phosphorylation is a key regulatory event in collagen-IV-mediated adhesion and proliferation of human lung tumor cell line, Calu-1.
BACKGROUND
The clinical phenomenon of lung cancer metastasis to specific target organs is believed to be a direct interaction between tumor cells and extracellular matrix components. During invasion, tumor cells attach to the basement membrane protein, collagen type IV, degrade it, migrate through blood vessels, and adhere to extracellular matrix proteins.


METHODS
Four nonsmall-cell lung cancer cells were tested for adhesion, proliferation, migration and morphologic alterations on collagen type IV matrix by immunoprecipitation, Western blotting, phase contrast and time lapse video microscopy.


RESULTS
Collagen type IV promoted Calu-1 cell adhesion (< 15 minutes) and motility (< 6 hours) without any significant effect on proliferation. beta(1)-integrin is essential for collagen type IV adhesion and 8 to 10 fold higher expression of beta1-integrin on the surface of Calu-1 cells was identified. A protein tyrosine phosphatase inhibitor, peroxyvanadate, caused 50% inhibition in the adhesion process within 5 minutes but exposure to 60 micromol/L genistein for 72 hours, a protein tyrosine kinase inhibitor, drastically inhibits Calu-1 cell proliferation (> 70%). We examined the influence of beta1-integrin, peroxyvanadate and genistein on the spreading morphogenesis of Calu-1 cells on Collagen type IV. Use of either 1 microg of anti beta1-integrin inhibitory antibody (P5D2), 100 micromol/L peroxyvanadate or 60 micromol/L genistein had dramatic influence on the spreading of Calu-1 cells. Finally, Focal adhesion kinase was identified as a phosphoprotein target.


CONCLUSIONS
We have characterized an in vitro model of matrix-specific binding of a lung cancer cell line, Calu-1 to Coll IV. Calu-1 cells use primarily a beta1-integrin mediated intracellular tyrosine phosphorylation phenomenon as the key regulatory mechanism for its binding advantage to Coll IV matrix.
Division of Thoracic Surgery, Brigham and Women's Hospital, Harvard Medical School, Boston, MA 02115, USA.
Nishit K
NK
Mukhopadhyay
David
D
Gilchrist
Gavin J
GJ
Gordon
Chang-Jie
CJ
Chen
Raphael
R
Bueno
Michael L
ML
Lu
Ravi
R
Salgia
David J
DJ
Sugarbaker
Michael T
MT
Jaklitsch
D002289:Carcinoma, Non-Small-Cell Lung / Q000473:pathology*; D002448:Cell Adhesion / Q000187:drug effects / Q000502:physiology; D002455:Cell Division / Q000187:drug effects / Q000502:physiology; D045744:Cell Line, Tumor / Q000166:cytology / Q000187:drug effects; D002465:Cell Movement / Q000187:drug effects / Q000502:physiology; D048429:Cell Size / Q000187:drug effects; D024141:Collagen Type IV / Q000378:metabolism*; D016895:Culture Media, Serum-Free; D051417:Focal Adhesion Kinase 1; D051416:Focal Adhesion Protein-Tyrosine Kinases; D019833:Genistein / Q000494:pharmacology; D006801:Humans; D019012:Integrin beta1 / Q000502:physiology*; D008175:Lung Neoplasms / Q000473:pathology*; D018715:Microscopy, Video; D009361:Neoplasm Invasiveness / Q000503:physiopathology*; D009363:Neoplasm Proteins / Q000502:physiology*; D010766:Phosphorylation / Q000187:drug effects; D011499:Protein Processing, Post-Translational / Q000187:drug effects; D011505:Protein-Tyrosine Kinases / Q000502:physiology; D011554:Pseudopodia / Q000187:drug effects; D014638:Vanadates / Q000494:pharmacology
D024141:Collagen Type IV; D016895:Culture Media, Serum-Free; D019012:Integrin beta1; D009363:Neoplasm Proteins; C500599:peroxyvanadate; D014638:Vanadates; D019833:Genistein; D011505:Protein-Tyrosine Kinases; D051417:Focal Adhesion Kinase 1; D051416:Focal Adhesion Protein-Tyrosine Kinases; C496050:PTK2 protein, human