Molecular mechanisms of nitric oxide-dependent inhibition of TPA-induced matrix metalloproteinase-9 (MMP-9) in MCF-7 cells.
Matrix metalloproteinase-9 (MMP-9) is implicated in the invasion and metastasis of breast cancer cells. We investigated the modulatory effects of nitric oxide (NO) on the 12-O-tetradecanoylphorbol 13-acetate (TPA)-induced MMP-9 expression in MCF-7 cells. Different chemical NO donors inhibited the extracellular content of TPA-induced MMP-9 protein and MMP-9 activity as assessed by gelatin-zymography and ELISA, respectively. Concomitant with the reduction in the extracellular MMP-9 content NO strongly decreased the steady-state levels of MMP-9 mRNA which in turn leads to a lower recruitment of MMP-9 transcripts to polysomes and to a diminished MMP-9 translation. Reporter gene assays revealed that the inhibition in MMP-9 expression by NO is mainly attributed to a 0.67 kb fragment of the 5'-promoter region of the MMP-9 gene but independent of the 3'untranslated region thus indicating that MMP-9 suppression by NO mainly results from transcriptional events. Electrophoretic mobility shift assays (EMSA), showed that NO specifically interferes with the TPA-induced DNA binding affinity of c-Jun and c-Fos without affecting the TPA-induced increase in the levels of the transcription factors. Using pharmacological inhibitors and small interfering (si)RNA we found that PKCdelta is indispensably involved in the TPA-triggered MMP-9 expression. Concomitantly, the TPA-evoked increase in total PKC activity was strongly attenuated in the lysates from NO-treated MCF-7 cells, thus suggesting that NO attenuates TPA-triggered MMP-9 mainly through a direct inhibition of PKCdelta. Modulation of MMP-9 by NO highlights the complex roles of NO in the regulation of MMP-9 in breast cancer cells.
Jespersen
Christine
C
Pharmazentrum Frankfurt/ZAFES, Klinikum der Johann Wolfgang Goethe-Universität, Frankfurt am Main, Germany.
Doller
Anke
A
Akool
El-Sayed
el-S
Bachmann
Malte
M
Müller
Roswitha
R
Gutwein
Paul
P
Mühl
Heiko
H
Pfeilschifter
Josef
J
Eberhardt
Wolfgang
W
D020413:3' Untranslated Regions; D045744:Cell Line, Tumor / Q000187:drug effects / Q000502:physiology; D006152:Cyclic GMP / Q000378:metabolism; D004790:Enzyme Induction / Q000187:drug effects*; D015971:Gene Expression Regulation, Enzymologic / Q000187:drug effects*; D017930:Genes, Reporter; D006801:Humans; D020780:Matrix Metalloproteinase 9 / Q000235:genetics / Q000378:metabolism*; D009569:Nitric Oxide / Q000378:metabolism*; D020030:Nitric Oxide Donors / Q000378:metabolism; D051745:Protein Kinase C-delta / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D034741:RNA, Small Interfering / Q000235:genetics / Q000378:metabolism; D015398:Signal Transduction / Q000502:physiology; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology*; D014157:Transcription Factors / Q000378:metabolism
D020413:3' Untranslated Regions; D020030:Nitric Oxide Donors; D034741:RNA, Small Interfering; D014157:Transcription Factors; D009569:Nitric Oxide; D051745:Protein Kinase C-delta; D020780:Matrix Metalloproteinase 9; D006152:Cyclic GMP; D013755:Tetradecanoylphorbol Acetate