Hierarchical cleavage of focal adhesion kinase by caspases alters signal transduction during apoptosis of intestinal epithelial cells.
OBJECTIVE
Purified intestinal epithelial cells die of detachment-induced apoptosis due to loss of cell anchorage during isolation. Anchorage-dependent cells form focal adhesions, sites of enhanced cell-matrix attachment that confer survival signals. Focal adhesion kinase (FAK), a component of the focal adhesion signaling complex, transduces these antiapoptotic signals. In this report, the molecular events leading to cleavage of FAK by caspases during apoptosis and its functional implications are defined.


METHODS
Cytosolic extracts of human intestinal epithelial cells undergoing detachment-induced apoptosis were analyzed by Western blotting, immunoprecipitation, and kinase assay.


RESULTS
FAK is cleaved by the ordered proteolytic activity of 2 different members of the caspase-3 family. The first cleavage is mediated by caspase-3, generating a 94/92-kilodalton-terminal fragment, which is processed by caspase-6 to an 84-kilodalton fragment. After apoptosis is initiated, the level of FAK phosphorylation is rapidly decreased, and the phosphorylation pattern of FAK-associated proteins is dramatically modified, showing significant yet divergent changes in signal transduction.


CONCLUSIONS
Cleavage of FAK during apoptosis of normal human cells is an example of the sequential, highly regulated, and coordinate action of caspases that not only dismantle a cell by proteolysis, but also alter the cell's signaling machinery.
Grossmann
J
J
Department of Medicine I, University of Regensburg, Germany.
Artinger
M
M
Grasso
A W
AW
Kung
H J
HJ
Schölmerich
J
J
Fiocchi
C
C
Levine
A D
AD
D000590:Amino Acid Chloromethyl Ketones / Q000494:pharmacology; D017209:Apoptosis / Q000502:physiology*; D020170:Caspase 1 / Q000378:metabolism; D053148:Caspase 3; D061945:Caspase Inhibitors; D020169:Caspases / Q000378:metabolism*; D003106:Colon / Q000166:cytology / Q000201:enzymology; D015853:Cysteine Proteinase Inhibitors / Q000494:pharmacology; D051417:Focal Adhesion Kinase 1; D051416:Focal Adhesion Protein-Tyrosine Kinases; D006801:Humans; D007413:Intestinal Mucosa / Q000166:cytology* / Q000201:enzymology*; D009842:Oligopeptides / Q000494:pharmacology; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism; D010766:Phosphorylation; D011505:Protein-Tyrosine Kinases / Q000378:metabolism*; D015398:Signal Transduction / Q000502:physiology*; D014443:Tyrosine / Q000378:metabolism
D000590:Amino Acid Chloromethyl Ketones; D061945:Caspase Inhibitors; D015853:Cysteine Proteinase Inhibitors; D009842:Oligopeptides; C108637:aspartyl-glutamyl-valyl-aspartal; C096713:benzyloxycarbonylvalyl-alanyl-aspartyl fluoromethyl ketone; D014443:Tyrosine; D019869:Phosphatidylinositol 3-Kinases; D011505:Protein-Tyrosine Kinases; D051417:Focal Adhesion Kinase 1; D051416:Focal Adhesion Protein-Tyrosine Kinases; C496050:PTK2 protein, human; C505407:CASP3 protein, human; D053148:Caspase 3; D020169:Caspases; D020170:Caspase 1