Expression of heat shock-regulated human growth hormone genes containing or lacking introns by NIH-3T3 and Wish cell lines.
A plasmid containing the complete genomic DNA of the human growth hormone (ghGH) comprising four introns and driven by the human promoter of the human gene of the 70 kDa heat shock protein (hsp70) has been used to transfect mouse NIH-3T3 and human Wish cells. Selected cell lines were characterized for stable hGH secretion. Similarly in the same NIH-3T3 cells, the stable expression of the same plasmid construct, but containing the complementary DNA of the hGH gene (chGH), was compared in terms of the effect of introns on heterologous protein synthesis. Genomic hGH recombined cells synthetized, in a heat regulated fashion, matured hsp70/hGH hybrid mRNA able to drive the secretion of a 22 kDa polypeptide. Like the natural hGH, this polypeptide expressed the functional hormonal activity of prolactin on casein secretion by mammary cells. The time course of hGH secretion was prolonged in ghGH transcripts, while that of mRNA degradation appeared delayed, especially in Wish cells, as compared to chGH expression. In the human Wish cells the decay of endogenous hsp mRNA has been compared to that of recombinant hsp mRNA, demonstrating that this human hsp70/hGH hybrid mRNA was present in the cytoplasm during a longer period than the human endogenous hsp70 mRNA. In conclusion, similar levels of expression and resulting gene products were expressed from the chGH or the ghGH gene in an inducible manner.
Alouani
S
S
Department of Genetic Engineering, IntraCell S.A., Geneva-Carouge, Switzerland.
L'Hote
P
P
Marq
J B
JB
Houdebine
L M
LM
Montandon
F
F
Chessebeuf-Padieu
M
M
Dreano
M
M
D016475:3T3 Cells; D000818:Animals; D002364:Caseins / Q000378:metabolism; D002460:Cell Line; D004247:DNA; D005786:Gene Expression Regulation; D013006:Growth Hormone / Q000235:genetics* / Q000378:metabolism; D006360:Heat-Shock Proteins / Q000235:genetics* / Q000378:metabolism; D006358:Hot Temperature; D006801:Humans; D007438:Introns; D051379:Mice; D010957:Plasmids; D011233:Precipitin Tests; D011388:Prolactin / Q000378:metabolism; D011401:Promoter Regions, Genetic; D012333:RNA, Messenger / Q000235:genetics; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism
D002364:Caseins; D006360:Heat-Shock Proteins; D012333:RNA, Messenger; D011993:Recombinant Fusion Proteins; D011388:Prolactin; D013006:Growth Hormone; D004247:DNA