Anti-apoptotic role of focal adhesion kinase (FAK): Induction of inhibitor-of-apoptosis proteins and apoptosis suppression by the overexpression of FAK in a human leukemic cell line, HL-60

Yoshiko Sonoda, Yaeko Matsumoto, Megumi Tago, Daisuke Yamamoto, Steven K. Hanks, Tadashi Kasahara

Research output: Contribution to journalArticle

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Abstract

Focal adhesion kinase (FAK) has an anti-apoptotic role in anchorage- dependent cells via an unknown mechanism. To elucidate the role of FAK in anti-apoptosis, we have established several FAK cDNA-transfected HL-60 cell lines and examined whether FAK-transfected cells have resistance to apoptotic stimuli. FAK-transfected HL-60 (HL-60/FAK) cells were highly resistant to apoptosis induced with hydrogen peroxide (1 mM) and etoposide (50 μg/ml) compared with the parental HL-60 cells or the vector-transfected cells, when determined using viability assay, DNA fragmentation, and flow cytometry analysis. Because no proteolytic cleavage of pro-caspase 3 to mature caspase 3 fragment was observed in HL-60/FAK cells, FAK was presumed to inhibit an upstream signal pathway leading to the activation of caspase 3. HL-60/FAK activated the phosphatidylinositide 3'-OH-kinase-Akt survival pathway and exhibited significant activation of NF-κB with marked induction of inhibitor-of-apoptosis proteins (IAPs: cIAP-1, cIAP-2, XIAP), regardless of the hydrogen peroxide-treated or untreated conditions, whereas no significant IAPs were detected in the parental or vector-transfected HL-60 cells. Apoptotic agents induced higher NF-(K)B activation in HL-60/FAK cells than in HL-60/Vect cells, and it appeared that sustained NF-κB activation is critical to the anti-apoptotic states in HL-60/FAK cells. Mutagenesis of FAK cDNA revealed that Y397 and Y925, which are involved in the tyrosine- phosphorylation sites, were prerequisite for the anti-apoptotic activity as well as induction of IAPs, and that K454, which is involved in the kinase activity, was also required for the full anti-apoptotic activity of FAK. Taken together, we have demonstrated definitively that FAK-transfected HL-60 cells, otherwise sensitive to apoptosis, become resistant to the apoptotic stimuli. We conclude that FAK activates the phosphatidylinositide 3'-OH- kinase-Akt survival pathway with the concomitant activation of NF-kB and induction of IAPs, which ultimately inhibit apoptosis by inhibiting caspase-3 cascade.

Original languageEnglish
Pages (from-to)16309-16315
Number of pages7
JournalJournal of Biological Chemistry
Volume275
Issue number21
DOIs
Publication statusPublished - 2000 May 26

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Inhibitor of Apoptosis Proteins
Focal Adhesion Protein-Tyrosine Kinases
Cells
Apoptosis
Cell Line
HL-60 Cells
Caspase 3
Chemical activation
Phosphotransferases
Hydrogen Peroxide
Complementary DNA
Mutagenesis
Phosphorylation
Flow cytometry
NF-kappa B
DNA Fragmentation
Etoposide

ASJC Scopus subject areas

  • Biochemistry

Cite this

Anti-apoptotic role of focal adhesion kinase (FAK) : Induction of inhibitor-of-apoptosis proteins and apoptosis suppression by the overexpression of FAK in a human leukemic cell line, HL-60. / Sonoda, Yoshiko; Matsumoto, Yaeko; Tago, Megumi; Yamamoto, Daisuke; Hanks, Steven K.; Kasahara, Tadashi.

In: Journal of Biological Chemistry, Vol. 275, No. 21, 26.05.2000, p. 16309-16315.

Research output: Contribution to journalArticle

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abstract = "Focal adhesion kinase (FAK) has an anti-apoptotic role in anchorage- dependent cells via an unknown mechanism. To elucidate the role of FAK in anti-apoptosis, we have established several FAK cDNA-transfected HL-60 cell lines and examined whether FAK-transfected cells have resistance to apoptotic stimuli. FAK-transfected HL-60 (HL-60/FAK) cells were highly resistant to apoptosis induced with hydrogen peroxide (1 mM) and etoposide (50 μg/ml) compared with the parental HL-60 cells or the vector-transfected cells, when determined using viability assay, DNA fragmentation, and flow cytometry analysis. Because no proteolytic cleavage of pro-caspase 3 to mature caspase 3 fragment was observed in HL-60/FAK cells, FAK was presumed to inhibit an upstream signal pathway leading to the activation of caspase 3. HL-60/FAK activated the phosphatidylinositide 3'-OH-kinase-Akt survival pathway and exhibited significant activation of NF-κB with marked induction of inhibitor-of-apoptosis proteins (IAPs: cIAP-1, cIAP-2, XIAP), regardless of the hydrogen peroxide-treated or untreated conditions, whereas no significant IAPs were detected in the parental or vector-transfected HL-60 cells. Apoptotic agents induced higher NF-(K)B activation in HL-60/FAK cells than in HL-60/Vect cells, and it appeared that sustained NF-κB activation is critical to the anti-apoptotic states in HL-60/FAK cells. Mutagenesis of FAK cDNA revealed that Y397 and Y925, which are involved in the tyrosine- phosphorylation sites, were prerequisite for the anti-apoptotic activity as well as induction of IAPs, and that K454, which is involved in the kinase activity, was also required for the full anti-apoptotic activity of FAK. Taken together, we have demonstrated definitively that FAK-transfected HL-60 cells, otherwise sensitive to apoptosis, become resistant to the apoptotic stimuli. We conclude that FAK activates the phosphatidylinositide 3'-OH- kinase-Akt survival pathway with the concomitant activation of NF-kB and induction of IAPs, which ultimately inhibit apoptosis by inhibiting caspase-3 cascade.",
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AU - Hanks, Steven K.

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