Expression of truncated forms of the bovine growth hormone gene in cultured mouse cells.
A synthetic oligonucleotide, 5'-d(CTAGT-CTAGACTAG)-3' which encodes translational termination codons in three reading frames, was inserted into either exon IV (pbGH-4A) or V (pbGH-5A) of the bovine growth hormone gene. The resultant plasmids, under the transcriptional regulation of the mouse metallothionein 1 promoter, were introduced into cultured mouse L-cells or rat GH3 cells. Compared to wild type bGH RNA, bGH-specific RNA transiently expressed from pBGH-5A or pBGH-4A DNA in mouse L-cells was similar or slightly smaller in size, respectively. Unexpectedly, bGH-4A RNA lacked exon IV sequences. Immunofluorescence and immunoprecipitation analyses revealed that wild type bGH was localized to the Golgi apparatus, while truncated hormones were confined to the cytoplasmic compartment of transfected cells. In addition, truncated hormones were shown to be secretion-defective albeit the bGH signal peptide was efficiently and precisely processed. Thus, structural alterations in the bGH gene can dramatically affect bGH precursor mRNA processing and hormone localization within cultured mouse fibroblast or rat pituitary cells.
Department of Zoological and Biomedical Sciences, Ohio University, Athens 45701.
S J
SJ
McAndrew
N Y
NY
Chen
P
P
Wiehl
L
L
DiCaprio
J
J
Yun
T E
TE
Wagner
S
S
Okada
J J
JJ
Kopchick
D000818:Animals; D001483:Base Sequence; D015152:Blotting, Northern; D002417:Cattle; D003001:Cloning, Molecular; D004252:DNA Mutational Analysis; D005091:Exons; D005455:Fluorescent Antibody Technique; D015870:Gene Expression; D005796:Genes; D013006:Growth Hormone / Q000235:genetics* / Q000378:metabolism; D007739:L Cells; D051379:Mice; D008969:Molecular Sequence Data; D009690:Nucleic Acid Conformation; D009841:Oligonucleotides / Q000737:chemistry; D011499:Protein Processing, Post-Translational; D012326:RNA Splicing; D012333:RNA, Messenger / Q000235:genetics; D011994:Recombinant Proteins / Q000378:metabolism
D009841:Oligonucleotides; D012333:RNA, Messenger; D011994:Recombinant Proteins; D013006:Growth Hormone