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Journal of Virology, October 1998, p. 8181-8190, Vol. 72, No. 10
Department of Molecular Biology and
Biochemistry, University of California, Irvine, Irvine, California
92697-3900
Received 15 May 1998/Accepted 3 July 1998
Previous work on the strict late (
0022-538X/98/$04.00+0
Copyright © 1998, American Society for Microbiology. All rights reserved.
Purification and Characterization of a Cellular Protein That
Binds to the Downstream Activation Sequence of the Strict Late
UL38 Promoter of Herpes Simplex Virus Type 1
) UL38 promoter of
herpes simplex virus type 1 identified three cis-acting
elements required for wild-type levels of transcription: a TATA box at
31, a consensus mammalian initiator element at the transcription
start site, and a downstream activation sequence (DAS) at +20 to +33.
DAS is found in similar locations on several other late promoters,
suggesting an important regulatory role in late gene expression. In
this communication, we further characterize the interaction between DAS
and a cellular protein which is found in both uninfected and infected
nuclear extracts. This protein was purified from HeLa nuclear extracts
and identified as the DNA binding component (Ku heterodimer) of
DNA-dependent protein kinase (DNA-PK) by peptide mapping. Highly
purified DNA-PK was able to stimulate UL38 transcription in
vitro approximately 10-fold. DAS is similar in sequence to another
element, nuclear regulatory element 1 (NRE1) of the
glucocorticoid-responsive mouse mammary tumor virus long terminal
repeat. NRE1 is known to specifically bind Ku in the absence of DNA
ends. We demonstrated that NRE1 is able to substitute for DAS in the
UL38 promoter to activate transcription as measured by in
vitro transcription and in vivo during infection of tissue culture
cells with recombinant virus. Also, we found that the binding of DNA-PK
to DAS involves the bases demonstrated to be important in
UL38 transcription and that the 70-kDa subunit of Ku binds
to DAS.
*
Corresponding author. Mailing address: Department of
Molecular Biology and Biochemistry, University of California, Irvine, Irvine, CA 92697-3900. Phone: (949) 824-5370. Fax: (949)
824-8551. E-mail: ewagner{at}uci.edu.
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