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Cloning and tissue-specific expression of the gene for mouse C-reactive protein
1Department of Microbiology, Ohio State University, Columbus 43210-1292.
Abstract:
C-reactive protein is a serum acute-phase reactant that increases several thousand-fold in concentration during inflammation in most mammals. However, mouse C-reactive protein is considered to be a minor acute-phase reactant, since its blood level increases only from approx. 0.1 to 1-2 micrograms/ml. A mouse genomic clone of approximately 5 kb was obtained to determine the molecular basis for the regulation of the expression of mouse C-reactive protein. Several cis-acting elements in the 5' flanking region that potentially regulate transcription were identified: two glucocorticoid-responsive elements, two CCAAT-enhancer-binding protein C (C/EBP) consensus elements that are required for the interleukin-1 responsiveness of some acute-phase reactant genes, an interleukin-6-responsive element, two hepatocyte nuclear factor-1 (HNF-1) elements and a single heat-shock element. Transfection of the hepatoma cell line Hep 3B.2 with a pCAT expression vector containing the 5' flanking sequence from -1083 to -3 bp from the transcriptional start site, and truncations of this sequence, localized elements that control the tissue-specific expression of mouse C-reactive protein to the two HNF-1 elements and a C/EBP, interleukin-1-responsive element located between -220 and -153, and -90 and -50 bp from the transcriptional start site. A constitutive nuclear protein from mouse-liver hepatocytes specifically binds to the HNF-1 elements. These findings explain the tissue-specific expression of the gene, as well as its limited expression during the acute-phase response.
Insights
Mouse C-reactive protein (CRP) shows limited acute-phase response. Regulatory elements, including hepatocyte nuclear factor-1 (HNF-1) and C/EBP, control its tissue-specific and inflammation-induced expression.
Area of Science:
- Molecular Biology
- Immunology
- Genetics
Background:
- C-reactive protein (CRP) is a major acute-phase reactant in most mammals, significantly increasing during inflammation.
- Mouse CRP is a minor acute-phase reactant, with only a modest increase in blood levels during inflammation.
Purpose of the Study:
- To investigate the molecular mechanisms regulating mouse C-reactive protein (CRP) gene expression.
- To identify cis-acting elements responsible for tissue-specific and acute-phase responsiveness.
Main Methods:
- Isolation and characterization of a mouse CRP genomic clone.
- Reporter gene assays using Hep 3B.2 hepatoma cells transfected with 5' flanking sequences and truncations.
- Identification of cis-acting regulatory elements including glucocorticoid-responsive elements, C/EBP, interleukin-6-responsive element, HNF-1 elements, and heat-shock element.
- Electrophoretic mobility shift assays to study protein binding to HNF-1 elements.
Main Results:
- Several cis-acting elements in the 5' flanking region of the mouse CRP gene were identified.
- Tissue-specific expression was localized to two HNF-1 elements and a C/EBP element.
- A constitutive nuclear protein from mouse liver specifically binds to the HNF-1 elements.
Conclusions:
- The identified regulatory elements explain the tissue-specific expression of mouse CRP.
- These elements also account for the limited induction of mouse CRP during the acute-phase response.
- Understanding these regulatory elements provides insight into CRP's role in mammalian inflammation.