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Structure and function of 5′-flanking regions of Bombyx mori fibroin heavy chain gene: Identification of a novel transcription enhancing element with a homeodomain protein-binding motif
- Source :
-
Insect Biochemistry & Molecular Biology . Jul2007, Vol. 37 Issue 7, p713-725. 13p. - Publication Year :
- 2007
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Abstract
- Abstract: We studied the promoter activity of a 5′-flanking region from −5000 to +24 (−5000/+24) in Bombyx mori fibroin heavy chain gene (fibH), fibH(−5000/+24). A luciferase reporter vector carrying fibH(−5000/+24) was bombarded to isolated posterior silk glands (PSGs). The PSGs showed a high luciferase activity when transplanted to larvae, indicating its potent promoter activity. Deletion experiments showed the requirement of fibH(−5000/−3844) and fibH(−2211/−542) for the promoter activity. These two regions and fibH(−541/+24) that contained the basal promoter were tandem fused to yield fibH(−5000/−3844:−2211/−542:−541/+24), which was found to retain 88% of the activity of fibH(−5000/+24). Germline transgenic silkworms bearing fibH(−5000/−3844:−2211/−542:−541/+24) as a promoter and enhanced green fluorescent protein (EGFP) gene as a reporter efficiently secreted EGFP in cocoons. The promoter activity of fibH(−2211/−542) was further investigated, because this contained a DNase I-hypersensitive site. The transient expression assay demonstrated that the activity of fibH(−2211/−542) required fibH(−1659/−1590), which contained the homeodomain protein-binding motif. Mutation experiments suggested a critical role of the motif for the promoter activity. Electrophoretic mobility shift assay (EMSA) demonstrated that a nuclear protein of PSGs bound to the motif. We propose fibH(−1659/−1590) as a novel transcription enhancer that plays a key role for the expression by recruiting a homeodomain protein. [Copyright &y& Elsevier]
- Subjects :
- *DEVELOPMENTAL biology
*FLUORESCENT polymers
*SILK
*LARVAE
Subjects
Details
- Language :
- English
- ISSN :
- 09651748
- Volume :
- 37
- Issue :
- 7
- Database :
- Academic Search Index
- Journal :
- Insect Biochemistry & Molecular Biology
- Publication Type :
- Academic Journal
- Accession number :
- 25342326
- Full Text :
- https://doi.org/10.1016/j.ibmb.2007.03.016