Non random activation of endogenous interleukin-2, (IL-2), IL-2 receptor alpha and IL-2 receptor beta genes after transfection of mouse fibroblasts with a cDNA for the alpha chain of the human IL-2 receptor.
Mouse fibroblasts do not ordinarily express components for the interleukin-2 receptor (IL-2R alpha, beta, and gamma). An analysis of these cells by reverse transcription followed by polymerase chain reaction, however, indicates the presence of transcripts specific for the IL-2R beta and gamma genes. Transfection of the cDNA for the alpha chain of the human IL-2R into LTK- mouse fibroblast cell line (L3 cells) leads, in long-term cultures, to the formation of transcripts of endogenous mouse IL-2, IL-2R alpha and beta genes, as detected by Northern blotting. Based upon the results of the binding of 125I-labeled IL-2 to the transfected cells, three IL-2-binding proteins of 55 kDa, 65 kDa and 75 kDa were expressed by the transfected cells. The 65-kDa and 75-kDa proteins bound IL-2 in the presence of monoclonal antibodies for the IL-2R alpha chain. These polypeptides assembled to form high-affinity IL-2R, as shown by Scatchard binding analyses. The receptors were functionally active, since the expression of H-2k major histocompatibility complex antigens on the surface membranes of L3 cells was enhanced by exposing the cells to IL-2. Activation of the IL-2 gene was also observed in long-term cultures of L alpha beta cells, another LTK- transfectant expressing the human IL-2R alpha chain. This type of gene activation was not observed in LTK- fibroblasts transfected with cDNA for human IL-2 or IL-2R beta genes. In L3 and L alpha beta cells, transcription of the endogenous IL-2 gene was suppressed by cyclosporin A and enhanced by cycloheximide. These data may have implications for gene therapy of cancer cells.
Han
D
D
INSERM U 268, Villejuif, France.
Plaisance
S
S
Rubinstein
E
E
Alileche
A
A
Pottin-Clemenceau
C
C
Kim
T S
TS
Cohen
E P
EP
Jasmin
C
C
Azzarone
B
B
D000818:Animals; D015152:Blotting, Northern; D002460:Cell Line; D003513:Cycloheximide / Q000494:pharmacology; D016572:Cyclosporine / Q000494:pharmacology; D017931:DNA Primers / Q000737:chemistry; D005347:Fibroblasts; D005786:Gene Expression Regulation / Q000187:drug effects; D005805:Genes, MHC Class I; D006183:H-2 Antigens / Q000235:genetics; D006801:Humans; D007376:Interleukin-2 / Q000378:metabolism; D051379:Mice; D016133:Polymerase Chain Reaction; D012333:RNA, Messenger / Q000235:genetics; D015375:Receptors, Interleukin-2 / Q000235:genetics* / Q000378:metabolism; D015533:Transcriptional Activation; D014162:Transfection
D017931:DNA Primers; D006183:H-2 Antigens; D007376:Interleukin-2; D012333:RNA, Messenger; D015375:Receptors, Interleukin-2; D016572:Cyclosporine; D003513:Cycloheximide