| Bombyx mori is an economically important organism and has been used extensively in sericulture industry for the commercial production of silk. The silk glands are functionally divided into three distinct compartments, the anterior (ASG), middle (MSG) and posterior (PSG) silk glands. Fibroins, the fibroin L and H chains and fibrohexamerin (formerly known as P25), are expressed exclusively in posterior silk gland (PSG), whereas the sericins are produced specifically in MSG. In this regard, the silk gland of Bombyx mori constitutes an attractive model for studying mechanism governing spatially and temporally programmed transcription.Sericin is a group of glue proteins which ensure the cohesion of the cocoon by aticking the silk threads together. Six kinds of sericin protein molecules are coded by two genes, ser1 and ser2, through alternative splicing. The Ser1 gene is the major constituents of sericins. Within the upstream sequence of the Ser1 gene, three specific protein binding sites are well known now. SA (-103 to -85), SB (-149 to -135) and SC (-204 to -183) have been characterized by in vitro transcription and DNase I foot printing. Silk gland factor I (SGF-1) that binds to the SA site exists specifically in the silk gland extract while SGF-3/POU-M1 was found to interact with the SC site. Current knowledge concerning the Ser1 promoter and factors associated with transcription regulation has been either partially or totally derived from in vitro assays. Recently reports have shown that there are some silkworm strains susceptible to Autographa californica multiple nuclear polyhedrosis virus (AcMNPV), which could serve as a novel in vivo gene delivery tool for the silkworm. Therefore developments in recombinant DNA technology and the advent of in vivo baculovirus expression system offer an approach to the study of the molecular mechanisms of transcriptional controls in silk gene in vivo.Owning to the poor understanding of mechanism responsible for thetissue-specific expression of the Ser\ gene, the present studies are carried to analyze the related region in the sericin-1 gene promoter. In the present study, we address to explore the possible cis-acting region(s) that takes part in determining the tissue-specific expression of the sericin-1 gene in vivo.To identify element involved in this transcription-dependent spatial restriction, EGFP reporter gene with SerJ promoter truncated in 5' terminal is introduced into the larvae with recombinant AcMNPV vector. A 209 bp DNA sequence upstream from the transcriptional start site (-586~-378) is found necessary to suppress the ectopic expresson. Analysis of this 209 bp region by overlapping deletion studies showed that a 25 bp region (-500~-476) suppresses the ectopic expression of the Serl promoter. Moreover, electrophoretic mobility shift assays (EMSA) were performed for a preliminarily analysis of the protein binding to this inhibitory region. Results showed that an unknown factor that binds to this 25 bp fragment is abundant in fat body. These results suggest that this 25 bp region and the unknown factor are necessary for determining the tissue-specificity of the sericin-1 promoter.Together with what have been reported, we supposed that there have been two kinds of mechanism involved in the control the expression of silk genes, they are activating in silk gland and repressing in other tissues. The specific expression of the sericin-1 gene could be the result of the co-ordination of multiple factors and cis-acting elements. This still needs further investigate.In general, the present studies may not only contribute to understanding the mechanisms of tissue-specific gene expression and benefit to improving the future production of silk-manufacture industry, but also do good to develop silk gland bioreactor in the future.
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