Constitutive expression of a microinjected glucose-regulated protein (grp78) fusion gene during early Xenopus laevis development.
In this study we have found that a rat glucose-regulated protein (grp) 78 chloramphenicol acetyltransferase (CAT) fusion gene deleted to -456 bp at the 5' end and injected into fertilized Xenopus eggs was first expressed in a constitutive manner in late blastula stage embryos and displayed increased expression as the embryos developed to the gastrula and neurula stages. Using a series of internal deletion mutants and linker-scanner mutants of the rat grp78 promoter, we have found that a CCAAT box and CCAAT-like element within the region -129 to -90 were essential for constitutive expression of the chimeric genes in neurula stage embryos. These results suggest conservation of the regulatory sequences within the grp78 promoter between rat and Xenopus. Interestingly, deletion or alteration of sequences between -130 and -149 had a dramatic stimulatory effect on basal promoter activity. This effect, which was not observed previously in rat cells, may be the result of upstream elements that are transcriptionally active in Xenopus and that can compensate for the mutated or deleted sequences. It is also possible that these results indicate the presence of a negative regulatory element that is recognized by the Xenopus transcriptional apparatus.
Vezina
C
C
Department of Biology, University of Waterloo, Ontario, Canada.
Wooden
S K
SK
Lee
A S
AS
Heikkila
J J
JJ
D000818:Animals; D001483:Base Sequence; D015500:Chloramphenicol O-Acetyltransferase / Q000235:genetics; D003001:Cloning, Molecular; D005260:Female; D005809:Genes, Regulator; D018840:HSP70 Heat-Shock Proteins / Q000235:genetics*; D008565:Membrane Proteins / Q000235:genetics*; D008845:Microinjections; D008969:Molecular Sequence Data; D009154:Mutation; D011401:Promoter Regions, Genetic; D014982:Xenopus laevis / Q000235:genetics*
D018840:HSP70 Heat-Shock Proteins; D008565:Membrane Proteins; C018551:glucose-regulated proteins; D015500:Chloramphenicol O-Acetyltransferase