72-bp element contains a critical control region for SV40 late expression in Xenopus laevis oocytes

Abstract

The SV40 late promoter is transcribed at least 10-fold more efficiently than the SV40 early promoter when SV40 DNA is injected into the germinal vesicle of Xenopus laevis oocytes. Late expression in the oocyte is independent of T antigen and does not require DNA replication. To identify DNA sequences required for SV40 late gene expression, 12 mutants spanning nucleotide position (np) 5187 to np 304 were injected into the germinal vesicles of X. laevis oocytes, and RNA was extracted 18 to 24 hr later. S1 nuclease analysis of the 5′ ends of SV40 late mRNA revealed that mutations in the origin of replication had no quantitative or qualitative effect on the 5′ late start sites. Mutants which deleted the 21-bp repeats did not reduce or alter use of the major RNA initiation site (np 295), but did reduce use of a minor initiation site within the 72-bp repeats. In contrast, deletion of or certain point mutatins in the 72-bp repeat decreased initiation from the major late start site. An 85-bp insertion containing a complete set of the 21-bp repeats positioned to the late side of the enhancer elements also decreased initiation from the major late start site. Thus, an element in the 72-bp repeat appears to be the major promoter element for late SV40 transcription in the oocyte.