Effect of phorbol esters on activity of Ha-ras gene promoter in HeLa cells.
OBJECTIVE
To study the role of phorbol esters (PMA) in the activity of promoter of Ha-ras in HeLa cells.


METHODS
After treatment with PMA, the growth rate of HeLa cells was measured by MTT assay and the expression of Ha-ras gene was detected by reversed transcriptase polymerase chain reaction. The regulation fragment of Ha-ras was constructed into the plasmid enhanced yellow fluorescent protein (EYFP) which did not contain promoter element. The recombinant plasmid pRasEYFP was transient transfected to HeLa cells. Using the EYFP gene as reporter, the activity of promoter of Ha-ras after treatment of PMA was assayed.


RESULTS
The growth rate of PMA-treated HeLa cells was markedly reduced compared with untreated cells. The expression of Ha-ras gene was obviously decreased and the activity of promoter of Ha-ras was decreased by 34.0 %and 26.7 % respectively in HeLa cells after treated with PMA (100 microg/L) for 48 h and 72 h. In the meanwhile the activity of protein kinase C (PKC) was decreased by 76.3 % and 73.2 % compared with the control cells, respectively.


CONCLUSIONS
PMA play an important role in regulation of the activity of promoter of Ha-ras in HeLa cells. The molecular mechanism may be through the PKC pathway.
Zhang
Wei
W
The Key Laboratory of Cell Proliferation and Regulation Biology of Ministry of Education, College of Life Science, Beijing Normal University, Beijing 100875, China.
Wang
Yong-Jun
YJ
Liu
Hui-Tu
HT
Gao
Ping
P
D001426:Bacterial Proteins / Q000494:pharmacology; D002273:Carcinogens / Q000494:pharmacology*; D011905:Genes, ras / Q000187:drug effects / Q000235:genetics*; D006367:HeLa Cells; D006801:Humans; D008164:Luminescent Proteins / Q000494:pharmacology; D010703:Phorbol Esters / Q000494:pharmacology*; D010957:Plasmids / Q000235:genetics; D011401:Promoter Regions, Genetic / Q000187:drug effects*; D011493:Protein Kinase C / Q000378:metabolism; D011994:Recombinant Proteins / Q000235:genetics; D015398:Signal Transduction; D014162:Transfection; D018631:ras Proteins / Q000096:biosynthesis / Q000235:genetics
D001426:Bacterial Proteins; D002273:Carcinogens; D008164:Luminescent Proteins; D010703:Phorbol Esters; D011994:Recombinant Proteins; C054562:yellow fluorescent protein, Bacteria; D011493:Protein Kinase C; D018631:ras Proteins