Synthetic promoter elements obtained by nucleotide sequence variation and selection for activity

Gerald M. Edelman, Robyn Meech, Geoffrey C. Owens, Frederick S. Jones

Research output: Contribution to journalArticle

54 Citations (Scopus)

Abstract

Eukaryotic transcriptional regulation in different cells involves large numbers and arrangements of cis and trans elements. To survey the number of cis regulatory elements that are active in different contexts, we have devised a high-throughput selection procedure permitting synthesis of active cis motifs that enhance the activity of a minimal promoter. This synthetic promoter construction method (SPCM) was used to identify >100 DNA sequences that showed increased promoter activity in the neuroblastoma cell line Neuro2A. After determining DNA sequences of selected synthetic promoters, database searches for known elements revealed a predominance of eight motifs: AP2, CEBP, GRE, Ebox, ETS, CREB, AP1, and SP1/MAZ. The most active of the selected synthetic promoters contain composites of a number of these motifs. Assays of DNA binding and promoter activity of three exemplary motifs (ETS, CREB, and SP1/MAZ) were used to prove the effectiveness of SPCM in uncovering active sequences. Up to 10% of 133 selected active sequences had no match in currently available databases, raising the possibility that new motifs and transcriptional regulatory proteins to which they bind may be revealed by SPCM. The method may find uses in constructing databases of active cis motifs, in diagnostics, and in gene therapy.

Original languageEnglish
Pages (from-to)3038-3043
Number of pages6
JournalProceedings of the National Academy of Sciences of the United States of America
Volume97
Issue number7
DOIs
Publication statusPublished - 28 Mar 2000
Externally publishedYes

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