Detection of c-sis proto-oncogene transcripts by direct enzyme-labeled cDNA probes and in situ hybridization.
Using in situ hybridization and platelet-derived growth factor (PDGF) cDNA probes labeled with horseradish peroxidase, PDGF-A and -B (c-cis proto-oncogene) mRNA transcripts were identified and localized in proliferating cultures. A human retinal pigment epithelial (RPE) cell line and a glial cell line were treated with either transforming growth factor beta-1 (TGFB1), phorbol-12-myristate-13-acetate (PMA), or thrombin from human plasma and compared for their ability to stimulate the production of PDGF-A and -B. Expression of both PDGF-A and -B transcripts were found to be localized predominantly in the cytoplasm of TGFB1-treated RPE cells, with a portion of these cells displaying a hybridization response in the nuclear region. When compared to PMA- and thrombin-treated cells, TGFB1 stimulated the RPE cell line to yield the greatest amount of detectable PDGF mRNA. In addition, the hybridization response observed in TGFB1-treated cells was shown to be RNA dependent.
McClintock
J T
JT
Digene Diagnostics, Inc., Silver Spring, Maryland 20904.
Chan
I J
IJ
Thaker
S R
SR
Katial
A
A
Taub
F E
FE
Aotaki-Keen
A E
AE
Hjelmeland
L M
LM
D002455:Cell Division; D002460:Cell Line; D015342:DNA Probes; D015870:Gene Expression / Q000187:drug effects; D006801:Humans; D066298:In Vitro Techniques; D009693:Nucleic Acid Hybridization; D010982:Platelet-Derived Growth Factor / Q000235:genetics*; D000090063:Proto-Oncogene Mas; D011519:Proto-Oncogenes; D012333:RNA, Messenger / Q000235:genetics; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D013917:Thrombin / Q000494:pharmacology; D016212:Transforming Growth Factor beta / Q000494:pharmacology
D015342:DNA Probes; C000715470:MAS1 protein, human; D010982:Platelet-Derived Growth Factor; D000090063:Proto-Oncogene Mas; D012333:RNA, Messenger; D016212:Transforming Growth Factor beta; D013917:Thrombin; D013755:Tetradecanoylphorbol Acetate