1: J Neurovirol. 1997 Jun;3(3):212-24. 

Repression of the HSV-1 latency-associated transcript (LAT) promoter by the
early growth response (EGR) proteins: involvement of a binding site immediately
downstream of the TATA box.

Tatarowicz WA, Martin CE, Pekosz AS, Madden SL, Rauscher FJ 3rd, Chiang SY,
Beerman TA, Fraser NW.

Wistar Institute, Philadelphia, Pennsylvania 19104, USA.

During herpes simplex virus (HSV) latency, in neurons of the nervous system, a
single family of viral transcripts (the Latency-Associated Transcripts or LATs)
are synthesized. Within the LAT promoter region, we have identified a consensus
sequence for the EGR proteins in an unusual position immediately downstream of
the TATA box. The early growth response (EGR) proteins are rapidly induced in
cells by stimuli which also induce HSV to reactivate from latency. In order to
determine if EGR proteins play any role in control of LAT transcription, we have
analyzed the interactions between EGR proteins and the LAT promoter. Gel
retardation and DNase I protection assays demonstrated that EGR1 zinc finger
protein bound specifically to the LAT promoter region EGR consensus sequence. To
determine if EGR proteins could modulate transcription through the LAT promoter,
cotransfection assays were performed using chloramphenicol acetyltransferase
(CAT) reporter constructs driven by either the wild-type LAT promoter or a LAT
promoter with a mutated EGR binding site. Contransfection of the wild-type LAT
promoter construct with EGR expression plasmids resulted in inhibition of the
basal level of CAT activity with EGR-2 but not EGR-1 or 3. However, normal
levels of CAT activity were observed in cotransfections using the mutant LAT
promoter CAT construct suggesting that repression was mediated by the binding of
EGR-2 proteins to the LAT promoter. Furthermore, data from combination binding
assays using EGR1 and TATA binding protein (TBP) in vitro support the hypothesis
that binding of EGR proteins to the LAT promoter prevents binding of TBP and
thus suppresses transcription. These results may provide a link between stress
responses in neurons of the CNS which activate the EGR family of proteins and
HSV reactivation from latency due to the same stress response.

PMID: 9200069 [PubMed - indexed for MEDLINE]


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2: J Biol Chem. 1996 Sep 27;271(39):23999-4004. 

Effect of DNA-binding drugs on early growth response factor-1 and TATA
box-binding protein complex formation with the herpes simplex virus latency
promoter.

Chiang SY, Welch JJ, Rauscher FJ 3rd, Beerman TA.

Experimental Therapeutics Department, Roswell Park Cancer Institute, Buffalo,
New York 14263, USA.

Adjacent binding sites for early growth response factor-1 (EGR1) and TATA
box-binding protein (TBP) were identified on the herpes simplex virus latency
promoter in previous work. The binding of EGR1 to the GC-rich region prevented
TBP binding to the AT-rich region. With the simultaneous addition of both EGR1
and TBP, the intercalator nogalamycin prevented EGR1 complex formation,
resulting in a dose-dependent increase of the TBP.DNA complex. The minor groove
binder chromomycin A3 inhibited EGR1 complex formation but resulted in a smaller
increase of the TBP complex. In contrast, an alkylating intercalator hedamycin
strongly inhibited binding of both proteins. The ability of these GC-binding
drugs to prevent EGR1.DNA complex formation was in the following order:
hedamycin > nogalamycin > chromomycin A3, and the specificity was nogalamycin >
chromomycin A3 > hedamycin. With transcription factor IIA (TFIIA) in the assay,
TBP was able to bind the promoter whereas formation of the EGR1.DNA complex was
reduced. An AT minor groove-binding drug, distamycin A, disrupted the
TBP.TFIIA.DNA complex and restored the EGR1.DNA complex. We conclude that the
binding motif and sequence preference of DNA-interactive drugs are manifested in
their ability to inhibit the transcription factor-DNA complexes.

PMID: 8798634 [PubMed - indexed for MEDLINE]


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