Expression of human pancreatic polypeptide precursors from a dicistronic mRNA in mammalian cells.
The ability of eukaryotic ribosomes to reinitiate translation at downstream AUG codons on polycistronic mRNAs was used to select transfected CHO clones that secreted a precursor to the human pancreatic polypeptide (PP). In the in vitro constructed transcription unit, a viral promoter directed the synthesis of a dicistronic mRNA. The PP cDNA was placed in the 5'-part of this transcript, while a DHFR cDNA was placed 3' to the PP. This dicistronic expression unit was transfected into CHO cells, and methotrexate-resistant colonies were selected. RNA-blots verified that the PP precursor and DHFR were expressed from the same dicistronic mRNA. The CHO cells synthesized the hormone precursor and secreted it through the constitutive secretory pathway.
D000818:Animals; D001345:Autoradiography; D002460:Cell Line; D003001:Cloning, Molecular; D004247:DNA; D005796:Genes; D006801:Humans; D010191:Pancreatic Polypeptide; D010957:Plasmids; D014176:Protein Biosynthesis; D011498:Protein Precursors / Q000096:biosynthesis* / Q000235:genetics; D012333:RNA, Messenger / Q000378:metabolism*; D012270:Ribosomes / Q000502:physiology*; D013762:Tetrahydrofolate Dehydrogenase / Q000235:genetics; D014158:Transcription, Genetic; D014170:Transformation, Genetic
C043710:PPY protein, human; D011498:Protein Precursors; D012333:RNA, Messenger; D010191:Pancreatic Polypeptide; D004247:DNA; D013762:Tetrahydrofolate Dehydrogenase