Plasminogen activator inhibitor type-1 protein, mRNA and gene transcription are increased by phorbol esters in human rhabdomyosarcoma cells.
By use of an enzyme-linked immunosorbent assay, we have found that phorbol 12-myristate 13-acetate (PMA) causes an approximately 10-fold increase in the level of type-1 plasminogen activator inhibitor (PAI-1) accumulated in conditioned medium of the human rhabdomyosarcoma cell line. Half-maximal stimulation occurred at approximately equal to 15 nM PMA. The effect was only observed with phorbol esters that are tumor promoting. Maximal levels of secreted PAI-1 were observed 24 h after PMA addition. The increase in secreted PAI-1 was preceded by a transient approximately 10-fold increase in intracellular PAI-1 content, maximal at 8 h after PMA addition. There was a 20-fold increase in the cellular level of two 2.3- and 3.4-kilobase PAI-1 mRNAs and a more than 5-fold increase in the PAI-1 gene transcription rate. The protein synthesis inhibitor cycloheximide (10 micrograms/ml) also increased the level of PAI-1 mRNA, and when both cycloheximide and PMA were used, an additive effect was observed. Cycloheximide changed the ratio between the two PAI-1 mRNAs in favor of the 3.4-kilobase species. Overall, the data show that transcriptional activation of the PAI-1 gene forms part of the pleiotropic responses to tumor-promoting phorbol esters.
Mayer
M
M
Finsen Laboratory, Rigshospitalet, Copenhagen, Denmark.
Lund
L R
LR
Riccio
A
A
Skouv
J
J
Nielsen
L S
LS
Stacey
S N
SN
Danø
K
K
Andreasen
P A
PA
D015152:Blotting, Northern; D002460:Cell Line; D003513:Cycloheximide / Q000494:pharmacology; D004797:Enzyme-Linked Immunosorbent Assay; D006023:Glycoproteins / Q000235:genetics*; D006801:Humans; D010703:Phorbol Esters / Q000494:pharmacology*; D015849:Plasminogen Inactivators; D012333:RNA, Messenger / Q000378:metabolism*; D012208:Rhabdomyosarcoma / Q000378:metabolism*; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D014158:Transcription, Genetic / Q000187:drug effects*
D006023:Glycoproteins; D010703:Phorbol Esters; D015849:Plasminogen Inactivators; D012333:RNA, Messenger; C008876:phorbol-12,13-didecanoate; D003513:Cycloheximide; D013755:Tetradecanoylphorbol Acetate