Molecular and in silico characterization of a promoter module and C/EBP element that mediate LPS‐induced RANTES/CCL5 expression in monocytic cells

• Published in: The FASEB journal 2001-03, Vol.15 (3), p.577-579
• Main Authors: Fessele, Sabine, Boehlk, Sabine, Mojaat, Anke, Miyamoto, Neil G., Werner, Thomas, Nelson, Edward L., Schloendorff, Detlef, Nelson, Peter J.
• Format: Article
• Language: English
• Subjects: bioinformatic; C/EBP; CCAAT-Enhancer-Binding Proteins - metabolism; Cell Line; Chemokine CCL5 - genetics; Chemokine CCL5 - metabolism; Computer Simulation; Dimerization; Genes, Reporter - genetics; Humans; Lipopolysaccharides - pharmacology; LPS; monocyte; Monocytes - drug effects; Monocytes - metabolism; NF-kappa B - genetics; NF-kappa B - metabolism; Promoter Regions, Genetic - genetics; RANTES/CCL5; Rel; Sp1; Sp1 Transcription Factor - genetics; Sp1 Transcription Factor - metabolism; transcriptional‐regulation; Transfection
• ISSN: 0892-6638; 1530-6860
• DOI: 10.1096/fj.00-0459fje

ABSTRACT The chemokine RANTES/CCL5 is a proinflammatory agent produced by a variety of tissues in response to specific stimuli. In human monocytes, RANTES/CCL5 transcription is up‐regulated rapidly and transiently in response to LPS. We describe here two regions that help control LPS‐driven transcription from the human RANTES/CCL5 promoter in monocytic cells. These sites were analyzed by using DNase I footprinting, transient transfection assays, site‐directed mutagenesis, and EMSA. RANTES site E (R(E), ‐125/‐99) constitutively binds C/EBP proteins in monocytic Mono Mac 6 cells. Mutation of region R(E) led to a significant (40%–50%) reduction in LPS‐induced promoter reporter activity. Region R(AB) is composed of tandem κB‐like elements R(A) and R(B) (‐73/‐34). These sites working in concert act as an LPS‐responsive promoter module. R(A) constitutively binds Sp1, and Rel p50/p65 following LPS stimulation. Either factor can mediate transcriptional effects at R(A). Induced Rel p50/p50 binding to site R(B) is required for LPS regulation of RANTES/CCL5 transcription. A series of computer models based on the RANTES/CCL5 promoter were generated to represent the organization of these functional elements. The models could identify LPS‐regulated promoters in human, other vertebrate, and viral sequences in various databases.