Single-nucleotide polymorphisms in the C-reactive protein (CRP) gene promoter that affect transcription factor binding, alter transcriptional activity, and associate with differences in baseline serum CRP level

A. J. Szalai, J. Wu, E. M. Lange, M. A. McCrory, C. D. Langefeld, A. Williams, S. O. Zakharkin, Varghese George, D. B. Allison, G. S. Cooper, F. Xie, Z. Fan, J. C. Edberg, R. P. Kimberly

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Abstract

To investigate whether functional polymorphisms exist in the C-reactive protein (CRP) gene, i.e., ones that contribute directly to differences in baseline CRP among individuals, we sequenced a 1,156-nucleotide-long stretch of the CRP gene promoter in 287 ostensibly healthy people. We identified two single-nucleotide polymorphisms (SNPs), a bi-allelic one at nucleotide -409 (G→A), and a tri-allelic one at -390 (C→T→A), both resident within the hexameric core of transcription factor binding E-box elements. Electrophoretic mobility shift assays confirmed that the SNP within the sequence -412CACGTG-407 (E-box 1) modulates transcription factor binding, and that the one within -394CACTTG-389 (E-box 2) supports transcription factor binding only when the -390 T allele is present. The commonest of four E-box 1/E-box 2 haplotypes (-409G/-390T) identified in the population supported highest promoter activity in luciferase reporter assays, and the rarest one (-409A/-390T) supported the least. Importantly, serum CRP in people with these haplotypes reproduced this rank order, i.e., people with the -409G/-390T haplotype had the highest baseline serum CRP (mean±SEM 10.9±2.25 μg/ml) and people with the -409A/-390T haplotype had the lowest (5.01±1.56 μg/ml). Furthermore, haplotype-associated differences in baseline CRP were not due to differences in age, sex, or race, and were still apparent in people with no history of smoking. At least two other SNPs in the CRP promoter lie within E-box elements (-198 C→T, E-box 4, and -861 T→C, E-box 3), indicating that not only is the quality of E-box sites in CRP a major determinant of baseline CRP level, but also that the number of E-boxes may be important. These data confirm that the CRP promoter does encode functional polymorphisms, which should be considered when baseline CRP is being used as an indicator of clinical outcome. Ultimately, development of genetic tests to screen for CRP expression variants could allow categorization of healthy people into groups at high versus low future risk of inflammatory disease.

Original languageEnglish (US)
Pages (from-to)440-447
Number of pages8
JournalJournal of Molecular Medicine
Volume83
Issue number6
DOIs
StatePublished - Jun 1 2005

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C-Reactive Protein
Single Nucleotide Polymorphism
Blood Proteins
Transcription Factors
Genes
Haplotypes
E-Box Elements
Nucleotides
Core Binding Factors
Electrophoretic Mobility Shift Assay
Luciferases
Smoking
Alleles

Keywords

  • Acute-phase reactants
  • Gene regulation
  • Inflammation

ASJC Scopus subject areas

  • Molecular Medicine
  • Drug Discovery
  • Genetics(clinical)

Cite this

Single-nucleotide polymorphisms in the C-reactive protein (CRP) gene promoter that affect transcription factor binding, alter transcriptional activity, and associate with differences in baseline serum CRP level. / Szalai, A. J.; Wu, J.; Lange, E. M.; McCrory, M. A.; Langefeld, C. D.; Williams, A.; Zakharkin, S. O.; George, Varghese; Allison, D. B.; Cooper, G. S.; Xie, F.; Fan, Z.; Edberg, J. C.; Kimberly, R. P.

In: Journal of Molecular Medicine, Vol. 83, No. 6, 01.06.2005, p. 440-447.

Research output: Contribution to journalArticle

Szalai, AJ, Wu, J, Lange, EM, McCrory, MA, Langefeld, CD, Williams, A, Zakharkin, SO, George, V, Allison, DB, Cooper, GS, Xie, F, Fan, Z, Edberg, JC & Kimberly, RP 2005, 'Single-nucleotide polymorphisms in the C-reactive protein (CRP) gene promoter that affect transcription factor binding, alter transcriptional activity, and associate with differences in baseline serum CRP level', Journal of Molecular Medicine, vol. 83, no. 6, pp. 440-447. https://doi.org/10.1007/s00109-005-0658-0
Szalai, A. J. ; Wu, J. ; Lange, E. M. ; McCrory, M. A. ; Langefeld, C. D. ; Williams, A. ; Zakharkin, S. O. ; George, Varghese ; Allison, D. B. ; Cooper, G. S. ; Xie, F. ; Fan, Z. ; Edberg, J. C. ; Kimberly, R. P. / Single-nucleotide polymorphisms in the C-reactive protein (CRP) gene promoter that affect transcription factor binding, alter transcriptional activity, and associate with differences in baseline serum CRP level. In: Journal of Molecular Medicine. 2005 ; Vol. 83, No. 6. pp. 440-447.
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abstract = "To investigate whether functional polymorphisms exist in the C-reactive protein (CRP) gene, i.e., ones that contribute directly to differences in baseline CRP among individuals, we sequenced a 1,156-nucleotide-long stretch of the CRP gene promoter in 287 ostensibly healthy people. We identified two single-nucleotide polymorphisms (SNPs), a bi-allelic one at nucleotide -409 (G→A), and a tri-allelic one at -390 (C→T→A), both resident within the hexameric core of transcription factor binding E-box elements. Electrophoretic mobility shift assays confirmed that the SNP within the sequence -412CACGTG-407 (E-box 1) modulates transcription factor binding, and that the one within -394CACTTG-389 (E-box 2) supports transcription factor binding only when the -390 T allele is present. The commonest of four E-box 1/E-box 2 haplotypes (-409G/-390T) identified in the population supported highest promoter activity in luciferase reporter assays, and the rarest one (-409A/-390T) supported the least. Importantly, serum CRP in people with these haplotypes reproduced this rank order, i.e., people with the -409G/-390T haplotype had the highest baseline serum CRP (mean±SEM 10.9±2.25 μg/ml) and people with the -409A/-390T haplotype had the lowest (5.01±1.56 μg/ml). Furthermore, haplotype-associated differences in baseline CRP were not due to differences in age, sex, or race, and were still apparent in people with no history of smoking. At least two other SNPs in the CRP promoter lie within E-box elements (-198 C→T, E-box 4, and -861 T→C, E-box 3), indicating that not only is the quality of E-box sites in CRP a major determinant of baseline CRP level, but also that the number of E-boxes may be important. These data confirm that the CRP promoter does encode functional polymorphisms, which should be considered when baseline CRP is being used as an indicator of clinical outcome. Ultimately, development of genetic tests to screen for CRP expression variants could allow categorization of healthy people into groups at high versus low future risk of inflammatory disease.",
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T1 - Single-nucleotide polymorphisms in the C-reactive protein (CRP) gene promoter that affect transcription factor binding, alter transcriptional activity, and associate with differences in baseline serum CRP level

AU - Szalai, A. J.

AU - Wu, J.

AU - Lange, E. M.

AU - McCrory, M. A.

AU - Langefeld, C. D.

AU - Williams, A.

AU - Zakharkin, S. O.

AU - George, Varghese

AU - Allison, D. B.

AU - Cooper, G. S.

AU - Xie, F.

AU - Fan, Z.

AU - Edberg, J. C.

AU - Kimberly, R. P.

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N2 - To investigate whether functional polymorphisms exist in the C-reactive protein (CRP) gene, i.e., ones that contribute directly to differences in baseline CRP among individuals, we sequenced a 1,156-nucleotide-long stretch of the CRP gene promoter in 287 ostensibly healthy people. We identified two single-nucleotide polymorphisms (SNPs), a bi-allelic one at nucleotide -409 (G→A), and a tri-allelic one at -390 (C→T→A), both resident within the hexameric core of transcription factor binding E-box elements. Electrophoretic mobility shift assays confirmed that the SNP within the sequence -412CACGTG-407 (E-box 1) modulates transcription factor binding, and that the one within -394CACTTG-389 (E-box 2) supports transcription factor binding only when the -390 T allele is present. The commonest of four E-box 1/E-box 2 haplotypes (-409G/-390T) identified in the population supported highest promoter activity in luciferase reporter assays, and the rarest one (-409A/-390T) supported the least. Importantly, serum CRP in people with these haplotypes reproduced this rank order, i.e., people with the -409G/-390T haplotype had the highest baseline serum CRP (mean±SEM 10.9±2.25 μg/ml) and people with the -409A/-390T haplotype had the lowest (5.01±1.56 μg/ml). Furthermore, haplotype-associated differences in baseline CRP were not due to differences in age, sex, or race, and were still apparent in people with no history of smoking. At least two other SNPs in the CRP promoter lie within E-box elements (-198 C→T, E-box 4, and -861 T→C, E-box 3), indicating that not only is the quality of E-box sites in CRP a major determinant of baseline CRP level, but also that the number of E-boxes may be important. These data confirm that the CRP promoter does encode functional polymorphisms, which should be considered when baseline CRP is being used as an indicator of clinical outcome. Ultimately, development of genetic tests to screen for CRP expression variants could allow categorization of healthy people into groups at high versus low future risk of inflammatory disease.

AB - To investigate whether functional polymorphisms exist in the C-reactive protein (CRP) gene, i.e., ones that contribute directly to differences in baseline CRP among individuals, we sequenced a 1,156-nucleotide-long stretch of the CRP gene promoter in 287 ostensibly healthy people. We identified two single-nucleotide polymorphisms (SNPs), a bi-allelic one at nucleotide -409 (G→A), and a tri-allelic one at -390 (C→T→A), both resident within the hexameric core of transcription factor binding E-box elements. Electrophoretic mobility shift assays confirmed that the SNP within the sequence -412CACGTG-407 (E-box 1) modulates transcription factor binding, and that the one within -394CACTTG-389 (E-box 2) supports transcription factor binding only when the -390 T allele is present. The commonest of four E-box 1/E-box 2 haplotypes (-409G/-390T) identified in the population supported highest promoter activity in luciferase reporter assays, and the rarest one (-409A/-390T) supported the least. Importantly, serum CRP in people with these haplotypes reproduced this rank order, i.e., people with the -409G/-390T haplotype had the highest baseline serum CRP (mean±SEM 10.9±2.25 μg/ml) and people with the -409A/-390T haplotype had the lowest (5.01±1.56 μg/ml). Furthermore, haplotype-associated differences in baseline CRP were not due to differences in age, sex, or race, and were still apparent in people with no history of smoking. At least two other SNPs in the CRP promoter lie within E-box elements (-198 C→T, E-box 4, and -861 T→C, E-box 3), indicating that not only is the quality of E-box sites in CRP a major determinant of baseline CRP level, but also that the number of E-boxes may be important. These data confirm that the CRP promoter does encode functional polymorphisms, which should be considered when baseline CRP is being used as an indicator of clinical outcome. Ultimately, development of genetic tests to screen for CRP expression variants could allow categorization of healthy people into groups at high versus low future risk of inflammatory disease.

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