Characterization of a polypurine/polypyrimidine sequence upstream of the mouse metallothionein-I gene

Nicole A. Becker, L. James Maher

Research output: Contribution to journalArticle

6 Scopus citations

Abstract

A 128 base pair long homopurine/homopyrimidine (R/Y) element is located ~1.2 kb upstream of the transcription start point of the mouse metallothionein-I (MT-I) gene. We present a detailed in vitro structural characterization of the MT-I R/Y sequence as determined by enzymatic and chemical probes. An ~190 bp fragment containing the MT-I R/Y sequence was subcloned into a recombinant vector. Low resolution analysis with S1 nuclease indicates that DNA in this region was unpaired in supercoiled plasmids treated at low pH. High resolution mapping with chemical probes selective for non-B DNA structures provides evidence that the MT-I R/Y sequence adopts one or more H-DNA structures. We also investigated this sequence to determine if it can influence transcriptional regulation. Promoter/reporter constructs were prepared in which the MT-I R/Y sequence was positioned in either orientation upstream of either the MT-I or HSV-TK promoters. Promoter/reporter activities were evaluated by transient transfection assays using mouse NIH3T3 cells. The MT-I R/Y sequence displayed no detectable activity as a cis-acting transcriptional regulatory element. These results demonstrate that although the MT-I R/Y sequence is able to adopt a non-B DNA structure under certain in vitro conditions, there is no evidence that this sequence plays a significant role in transcriptional regulation.

Original languageEnglish (US)
Pages (from-to)1951-1958
Number of pages8
JournalNucleic acids research
Volume26
Issue number8
DOIs
StatePublished - Apr 15 1998

ASJC Scopus subject areas

  • Genetics

Fingerprint Dive into the research topics of 'Characterization of a polypurine/polypyrimidine sequence upstream of the mouse metallothionein-I gene'. Together they form a unique fingerprint.

  • Cite this