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Department of Molecular and Experimental Medicine, W. M. Keck Autoimmune Disease Center, The Scripps Research Institute, La Jolla, CA 92037
The complement regulatory protein decay-accelerating factor (DAF or CD55) protects host tissue from complement-mediated injury by inhibiting the classical and alternative complement pathways. Besides its role in complement regulation, DAF has also been shown to be a key player in T cell immunity. Modulation of DAF expression could therefore represent a critical regulatory mechanism in both innate and adaptive immune responses. To identify and characterize key transcriptional regulatory elements controlling mouse Daf1 expression, a 2.5-kb fragment corresponding to the 5' flanking region of the mouse Daf1 gene was cloned. Sequence analysis showed that the mouse Daf1 promoter lacks conventional TATA and CCAAT boxes and displays a high guanine and cytosine content. RACE was used to identify one major and two minor transcription start sites 47, 20, and 17 bp upstream of the translational codon. Positive and negative regulatory regions were identified by transiently transfecting sequential 5'deletion constructs of the 5'flanking region into NIH/3T3, M12.4, and RAW264.7 cells. Mutational analyses of the promoter region combined with Sp1-specific ELISA showed that the transcription factor Sp1 is required for basal transcription and LPS-induced expression of the Daf1 gene. These findings provide new information on the regulation of the mouse Daf1 promoter and will facilitate further studies on the expression of Daf1 during immune responses.
The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked advertisement in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
1 This work was supported by National Institutes of Health Grants ES07511, ES09802, and ES08080, and The Stein Endowment Fund. D.M.C. is an Arthritis National Research Foundation Grant Recipient.
2 Address correspondence and reprint requests to Dr. K. Michael Pollard, Department of Molecular and Experimental Medicine, MEM131, The Scripps Research Institute, 10550 North Torrey Pines Road, La Jolla, CA 92037. E-mail address: mpollard{at}scripps.edu
3 Abbreviations used in this paper: DAF, decay-accelerating factor; RT, room temperature; GC, guanine and cytosine; TF, transcription factor.
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