Characterization of the Regulatory Elements of the Zebrafish Type II Keratin Gene, zfCKII
Date Issued
2004
Date
2004
Author(s)
Lu, Chun-Hung
DOI
zh-TW
Abstract
Type I (acidic) and type II (basic) keratins, two major classes of intermediate filament proteins, is encoded by at least 30 genes and are mainly expressed in epithelial cells in mammals. The regulation of keratin expression and transcription factors have been studied in mammals. However, the role of regulation is still unclear although there are many studies on the expression pattern of keratin in zebrafish (Danio rerio). In this report, we characterized an upstream 2274 bp (-2274/-1) of zebrafish type II cytokeratin (zfCKII) by transgenic analysis. Series of upstream deletions linked to the red fluorescent protein (RFP) reporter were constructed and microinjected into one-cell fertilized eggs. Transient transgenic analysis showed that the segments of -2274/-1, -1500/-1, -1000/-1, -600/-1, -500/-1, -400/-1 and -200/-1 enabled to direct RFP to be expressed in skin. But, the expression of RFP was completely abolished in the transgenic fish when the segment of -90/-1 and -100/-1 were transferred, indicating that -200/-1 is the minimal promoter to drive RFP expression in skin. Nevertheless, the segments of -1500/-1, -1000/-1, -600/-1, and -500/-1 were capable of directing RFP expression not only in skin but also in skeletal muscle. The expression rate in muscle ranged from 50 to 70%, compared to 0% of -2274/-1-injected embryos. Therefore, we propose that a muscle repressive element may exist in -2274/-1500, although the GFP expression rate in muscle of embryos injected with -2274/-1500 fused with -300/-1 of muscle regulatory factor myf-5 did not decrease significantly. Sequence analysis of zfCKII -200/-100 showed that there is a putative krüppel-like factor binding site located at -151/-121. When we mutated the segment -146/-143 from GGGG to AAAA, we found that this mutated segment almost lost its function to drive RFP expression in skin, suggesting that -146/-143 is an essential sequence for keratin expression in the skin of zebrafish.
Subjects
皮膚
調控
角質蛋白
Skin
Regulation
Keratin
Type
other
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