Anti-sense inhibition of tissue plasminogen activator production in differentiated F9 teratocarcinoma cells.
F9 teratocarcinoma cells secrete the serine protease, tissue plasminogen activator (t-PA), upon differentiation induced in vitro by retinoic acid (RA) or RA and dibutyryl cAMP (RA/dbcAMP). A recombinant plasmid capable of directing the production of t-PA anti-sense RNA was constructed and transfected into F9 stem cells in an attempt to create a hypomorphic phenotype for t-PA synthesis. Several colonies were isolated which contained anti-sense RNA and which showed greater than a 50% reduction in t-PA activity upon differentiation. One such colony, 3b4, exhibited a 75% reduction in t-PA activity and was analyzed further. Large quantities of t-PA anti-sense transcript were expressed in the stem cells which are characterized by the absence of t-PA gene expression. In the induced cells, which normally express t-PA, the amount of detectable anti-sense transcript was significantly decreased. The amount of t-PA mRNA in differentiated cells containing t-PA anti-sense RNA was comparable to that in differentiated control cells. Subcellular localization of the mRNA in induced 3b4 cells appeared to be the same as induced control cells. Expression of collagen type IV, another marker of differentiation, was also monitored and was unaffected by the presence of t-PA anti-sense RNA in RA/dbcAMP-treated cells. The inhibition of differentiation-specific gene expression by anti-sense RNA may be useful for further studies of developmentally regulated genes.
Pecorino
L T
LT
Program in Cellular and Developmental Biology, State University of New York, Stony Brook 11794-8651.
Rickles
R J
RJ
Strickland
S
S
D000818:Animals; D003994:Bucladesine / Q000494:pharmacology; D002454:Cell Differentiation / Q000187:drug effects; D002460:Cell Line; D003094:Collagen / Q000096:biosynthesis; D051379:Mice; D010641:Phenotype; D010957:Plasmids; D012313:RNA / Q000032:analysis; D018075:RNA, Complementary; D012333:RNA, Messenger / Q000032:analysis; D013724:Teratoma / Q000378:metabolism*; D010959:Tissue Plasminogen Activator / Q000096:biosynthesis* / Q000235:genetics
D018075:RNA, Complementary; D012333:RNA, Messenger; D012313:RNA; D003994:Bucladesine; D003094:Collagen; D010959:Tissue Plasminogen Activator