Polyamine depletion inhibits etoposide-induced NF-kappaB activation in transformed mouse fibroblasts.
In a previous research, we have shown that adequate levels of polyamines are required in transformed mouse fibroblasts for the correlated activations of MAPK subtypes (ERK and JNK) and caspases induced by etoposide and leading to apoptosis. We report now that the treatment of fibroblasts with etoposide also elicited a progressive and sustained increase of NF-kappaB activation. The DNA binding activity of p65 NF-kappaB subunit was increased up to approximately 4-fold and was accompanied by enhancement of p65 phosphorylation. A two days pre-treatment of fibroblasts with alpha-difluoromethylornithine (DFMO), which caused polyamine depletion, provoked a slight activating effect when given alone, but markedly inhibited the etoposide-induced increases in p65 DNA binding and phosphorylation. The NF-kappaB inhibiting effect of DFMO was prevented by the addition of exogenous putrescine, which restored the intracellular content of polyamines. Selective inhibitors of the etoposide-stimulated MAPK subtypes also reduced NF-kappaB activation. Moreover, pharmacological NF-kappaB inhibition reduced the increase in caspase activity and cell death elicited by etoposide, suggesting that NF-kappaB is involved in signaling to apoptosis. The results of the present study, together with our previous findings, suggest that polyamines play a permissive role in the pathways triggered by etoposide and leading to cell death of fibroblasts, by supporting the activation of MAPKs, NF-kappaB and caspases.
Tantini
B
B
Department of Biochemistry G. Moruzzi, University of Bologna, Bologna, Italy. benedetta.tantini@unibo.it
Pignatti
C
C
Fattori
M
M
Fiumana
E
E
Facchini
A
A
Stefanelli
C
C
Caldarera
C M
CM
Pegg
A E
AE
Flamigni
F
F
D000818:Animals; D000972:Antineoplastic Agents, Phytogenic / Q000494:pharmacology*; D017209:Apoptosis; D015153:Blotting, Western; D020169:Caspases / Q000378:metabolism; D002461:Cell Line, Transformed; D002467:Cell Nucleus / Q000378:metabolism; D003374:Coumarins / Q000737:chemistry; D004247:DNA / Q000378:metabolism; D000518:Eflornithine / Q000737:chemistry; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005047:Etoposide / Q000737:chemistry / Q000494:pharmacology*; D005347:Fibroblasts / Q000378:metabolism*; D005419:Flavonoids / Q000494:pharmacology; D020287:In Situ Nick-End Labeling; D020935:MAP Kinase Signaling System; D051379:Mice; D016328:NF-kappa B / Q000378:metabolism*; D010766:Phosphorylation; D011073:Polyamines / Q000737:chemistry*; D011485:Protein Binding; D013997:Time Factors
D000972:Antineoplastic Agents, Phytogenic; D003374:Coumarins; D004791:Enzyme Inhibitors; D005419:Flavonoids; D016328:NF-kappa B; D011073:Polyamines; D005047:Etoposide; D004247:DNA; C030123:coumarin; D020169:Caspases; C093973:2-(2-amino-3-methoxyphenyl)-4H-1-benzopyran-4-one; D000518:Eflornithine