Role of paxillin in metabolic oxidative stress-induced cytoskeletal reorganization: Involvement of SAPK signal transduction pathway and PTP-PEST geneexpression
Sh. Song et al., Role of paxillin in metabolic oxidative stress-induced cytoskeletal reorganization: Involvement of SAPK signal transduction pathway and PTP-PEST geneexpression, FREE RAD B, 29(1), 2000, pp. 61-70
Previous studies have shown that glucose deprivation-induced cell death is
associated with apoptosis, which is characterized by cellular membrane bleb
bing in multi-drug-resistant human breast carcinoma MCF-7/ADR cells. In thi
s study, we investigated the mechanism of glucose deprivation-induced cytos
keletal reorganization, which is known to be responsible fur the morphologi
cal alterations. An increase in the formation of focal adhesion and stress
fibers was observed during the early pried of glucose deprivation (1-2 h).
However, a disappearance of focal adhesion complexes and a loss of stress f
iber formation along with membrane blebbing were observed when glucose depr
ivation continued. These alterations were delayed in MCF-7/ADR cells transf
ected with bcl-2 and completely suppressed by treatment with an antioxidant
, N-acetyl-L-cysteine. These results indicated that glucose deprivation-ind
uced oxidative stress caused the cytoskeletal reorganization. The glucose d
eprivation-induced alteration of cytoskeletal organization was further inve
stigated by studying a modification of paxillin, one of the focal adhesion
proteins. Immunoblotting with anti-paxillin antibody showed that the paxill
in band shifted from 68 kDa to about 80 kDa during 1-4 h of glucose depriva
tion. The mobility shift indicated the modification of paxillin. This possi
bility was further studied by an immunoprecipitation assay with anti-paxill
in/anti-phosphotyrosine antibody and phosphoamino acid analysis (PAA). The
immunoprecipitation study revealed that the level of tyrosine phosphorylati
on of paxillin was maintained for 2 h and then markedly decreased without a
change in the total level of paxillin. The PAA study showed that paxillin
is dephosphorylated on tyrosine concurrent with phosphorylation on serine/t
hreonine. Expression of a dominant-negative mutant of c-Jun NH2-terminal ki
nase (JNK1) suppressed glucose deprivation-induced JNK1 activation, PTP-PES
T gene expression, and alteration of paxillin. Taken together, these result
s suggest that the alteration of the phosphorylation/dephosphorylation of p
axillin may he related to the cytoskeletal reorganization and these events
are mediated by glucose deprivation-induced oxidative stress and the stress
-activated protein kinase signal transduction pathway. (C) 2000 Elsevier Sc
ience Inc.