J Biol Chem, Vol. 275, Issue 9, 6600-6607, March 3, 2000
Characterization of a Repressor Element and a Juxtaposed
Tissue-restricted Activator Element Located on the Distal
neu Gene Promoter*
Ting-Chung
Suen
and
Paul E.
Goss§¶
From the Breast Cancer Prevention Program, The
Toronto Hospital, Oncology Research Laboratories and
§ Breast Group, Department of Medical Oncology, Princess
Margaret Hospital, Toronto, Ontario, M5G 2M9, Canada
The proto-oncogene neu (HER2 or c-erbB2) is
overexpressed with or without gene amplification in 20-30% of breast
cancers. In patients, neu amplification or overexpression
in breast and ovarian cancer correlates with poor prognosis and tumor
resistance to chemotherapy. neu-induced transformation can
be reversed by the suppression of neu gene transcription.
To further understand how neu gene transcription is
regulated and to identify a possible transcriptional repressor(s) of
neu, we identified a negative regulatory element known
previously to be located within a 1-kilobase (kb) DNA fragment of an
unknown sequence, upstream of the proximal neu gene
promoter. One of several DNA fragments subcloned from this region
suppressed transcriptional activity of the proximal neu
gene promoter. Sequencing of the 1-kb fragment confirmed the location
of the repressor element to be between an AluI and a RsaI sites, around 1.4 kb upstream to the translation start
site. Various deletions were introduced into the
AluI-RsaI fragment and subcloned into both the
native neu promoter and a heterologous thymidine kinase promoter.
Subsequent transfections and reporter gene assays in cell lines of
various tissues of origin confirmed and narrowed the repressor activity
to a 120-base pair NlaIV-MslI fragment located
between 
1385 and 1266. Importantly, specific protein binding
activity to this element could be detected with nuclear extracts
isolated from these cell lines. In contrast, a 28-base pair
MslI-RsaI fragment (1265 to 1238), located
immediately 3' of the putative repressor element, was found to form
protein-DNA complexes with only nuclear extracts isolated from a colon
carcinoma cell line. This specific protein binding activity correlated
with a previously unknown transcriptional stimulatory activity only in
this cell line.
*
This work was supported by the Breast Cancer Prevention
Program of The Toronto Hospital (to P. E. G.).The costs of publication of this
article were defrayed in part by the
payment of page charges. The article
must therefore be hereby marked
"advertisement" in accordance with 18 U.S.C. Section
1734 solely to indicate this fact.
The nucleotide sequence(s) reported in this paper has been submitted to the GenBankTM/EMBL Data Bank with accession number(s) AF208052.
CiteULike 
Complore 
Connotea 
Del.icio.us 
Digg 
Reddit 
Technorati What's this?
