TGTCACA Motif Is a Novel cis-Regulatory Enhancer Element Involved in Fruit-specific Expression of the cucumisin Gene
| Autor: | Yasuo Aizono, Kiyoaki Yonesu, Ayako Hirata, Hiroshi Yamagata |
|---|---|
| Rok vydání: | 2002 |
| Předmět: |
DNA
Plant Transcription Genetic Molecular Sequence Data Negative regulatory element Chimeric gene Regulatory Sequences Nucleic Acid Biology Biochemistry Cucumis Sequence Homology Nucleic Acid Cloning Molecular Nuclear protein Promoter Regions Genetic Enhancer Molecular Biology Gene Serine protease Reporter gene Base Sequence Serine Endopeptidases Nuclear Proteins food and beverages Promoter Cell Biology Molecular biology Cell biology Enhancer Elements Genetic biology.protein |
| Zdroj: | Journal of Biological Chemistry. 277:11582-11590 |
| ISSN: | 0021-9258 |
| DOI: | 10.1074/jbc.m109946200 |
| Popis: | Cucumisin, a subtilisin-like serine protease, is expressed at high levels in the fruit of melon (Cucumis melo L.) and accumulates in the juice. We investigated roles of the promoter regions and DNA-protein interactions in fruit-specific expression of the cucumisin gene. In transient expression analysis, a chimeric gene construct containing a 1.2-kb cucumisin promoter fused to a beta-glucuronidase (GUS) reporter gene was expressed in fruit tissues at high levels, but the promoter activities in leaves and stems were very low. Deletion analysis indicated that a positive regulatory region is located between nucleotides -234 and -214 relative to the transcriptional initiation site. Gain-of-function experiments revealed that this 20-bp sequence conferred fruit specificity and contained a regulatory enhancer. Gel mobility shift experiments demonstrated the presence of fruit nuclear factors that interact with the cucumisin promoter. A typical G-box (GACACGTGTC) present in the 20-bp sequence did not bind fruit protein, but two possible cis-elements, an I-box-like sequence (AGATATGATAAAA) and an odd base palindromic TGTCACA motif, were identified in the promoter region between positions -254 and -215. The I-box-like sequence bound more tightly to fruit nuclear protein than the TGTCACA motif. The I-box-like sequence functions as a negative regulatory element, and the TGTCACA motif is a novel enhancer element necessary for fruit-specific expression of the cucumisin gene. Specific nucleotides responsible for the binding of fruit nuclear protein in these two elements were also determined. |
| Databáze: | OpenAIRE |
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