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. 1989 Oct;86(20):7890–7894. doi: 10.1073/pnas.86.20.7890

Site-specific mutations alter in vitro factor binding and change promoter expression pattern in transgenic plants.

E Lam 1, P N Benfey 1, P M Gilmartin 1, R X Fang 1, N H Chua 1
PMCID: PMC298177  PMID: 2813365

Abstract

The 35S promoter of cauliflower mosaic virus (CaMV) is able to confer high-level gene expression in most organs of transgenic plants. A cellular factor from pea and tobacco leaf tissue, which recognizes nucleotides in a tandemly repeated TGACG motif at the -75 region of this promoter, has been detected by DNase I footprinting and gel retardation assays. This factor is named activation sequence factor 1 (ASF-1). A cellular factor binding to the two TGACG motifs can also be detected in tobacco root extracts. Mutations at these motifs inhibit binding of ASF-1 to the 35S promoter in vitro. When examined in transgenic tobacco, these mutations cause a 50% drop in leaf expression of the 35S promoter. In addition, these same mutations attenuate stem and root expression of the 35S promoter about 5- to 10-fold when compared to the level of expression in leaf. In contrast, mutations at two adjacent CCAAT-box-like sequences have no dramatic effect on promoter activity in vivo. A 21-base-pair element containing the two TGACG motifs is sufficient for binding of ASF-1 in vitro when inserted in a green-tissue-specific promoter. In vivo, the insertion of an ASF-1 binding site caused high levels of expression in root. Thus, a single factor binding site that is defined by site-specific mutations is shown to be sufficient to alter the expression pattern of promoters in vivo.

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Selected References

These references are in PubMed. This may not be the complete list of references from this article.

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