Transactivation of the grp78 promoter by malfolded proteins, glycosylation block, and calcium ionophore is mediated through a proximal region containing a CCAAT motif which interacts with CTF/NF-I.
The 78-kDa glucose-regulated protein (GRP78) is a major endoplasmic reticulum (ER) protein that can form stable associations with a variety of proteins retained in the ER because of underglycosylation or other conformational changes. In this study, we provide evidence at the transcriptional level that a conformationally abnormal protein, an altered herpes simplex virus type 1 envelope protein that is retained in the ER of a mammalian cell line, transactivates the grp78 promoter. In contrast, the normal viral envelope glycoprotein does not elevate grp78 promoter activity. Using a series of 5' deletions, linker-scanning, and internal deletion mutations spanning a 100-bp region from -179 to -80, we correlate the cis-acting regulatory elements mediating the activation of grp78 by malfolded proteins, glycosylation block, and the calcium ionophore A23187. We show that they all act through the same control elements, suggesting that they share a common signal. We report here that the highly conserved grp element, while important for basal level and induced grp78 expression, is functionally redundant. The single most important element, by linker-scanning analysis, is a 10-bp region that contains a CCAAT motif. It alone is not sufficient for promoter activity, but a 40-bp region (-129 to -90) that contains this motif is essential for mediating basal level and stress inducibility of the grp78 promoter. We show that the transcription factor CTF/NF-I is able to transactivate the grp78 promoter through interaction with this CCAAT motif.
Wooden
S K
SK
Department of Biochemistry, University of Southern California School of Medicine, Los Angeles 90033.
Li
L J
LJ
Navarro
D
D
Qadri
I
I
Pereira
L
L
Lee
A S
AS
D000818:Animals; D001483:Base Sequence; D022762:CCAAT-Enhancer-Binding Proteins; D000001:Calcimycin / Q000494:pharmacology*; D002352:Carrier Proteins / Q000235:genetics*; D002460:Cell Line; D002872:Chromosome Deletion; D004268:DNA-Binding Proteins / Q000378:metabolism*; D000091342:Endoplasmic Reticulum Chaperone BiP; D006031:Glycosylation; D006360:Heat-Shock Proteins / Q000235:genetics*; D018832:Molecular Chaperones; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D051839:NFI Transcription Factors; D009687:Nuclear Proteins / Q000378:metabolism*; D009841:Oligonucleotides; D010957:Plasmids; D016133:Polymerase Chain Reaction; D011401:Promoter Regions, Genetic; D011487:Protein Conformation; D018139:Simplexvirus / Q000235:genetics; D014157:Transcription Factors / Q000378:metabolism*; D015533:Transcriptional Activation / Q000187:drug effects; D014162:Transfection
D022762:CCAAT-Enhancer-Binding Proteins; D002352:Carrier Proteins; D004268:DNA-Binding Proteins; D000091342:Endoplasmic Reticulum Chaperone BiP; D006360:Heat-Shock Proteins; D018832:Molecular Chaperones; D051839:NFI Transcription Factors; D009687:Nuclear Proteins; D009841:Oligonucleotides; D014157:Transcription Factors; D000001:Calcimycin