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Myelin basic protein as a novel genetic risk factor in rheumatoid arthritis--a genome-wide study combined with immunological analyses.

Terao C, Ohmura K, Katayama M, Takahashi M, Kokubo M, Diop G, Toda Y, Yamamoto N, Human Disease Genomics Working GroupRheumatoid Arthritis (RA) Clinical and Genetic Study ConsortiumShinkura R, Shimizu M, Gut I, Heath S, Melchers I, Manabe T, Lathrop M, Mimori T, Yamada R, Matsuda F - PLoS ONE (2011)

Bottom Line: The transcription of MBP was significantly elevated with the risk allele compared to the alternative allele (p<0.001).We also established by immunohistochemistry that MBP was expressed in the synovial lining layer of RA patients, the main target of inflammation in the disease.This is the first report of a genetic study in RA implicating MBP as a potential autoantigen and its involvement in pathogenesis of the disease.

View Article: PubMed Central - PubMed

Affiliation: Center for Genomic Medicine, Graduate School of Medicine, Kyoto University, Kyoto, Japan.

ABSTRACT
Rheumatoid arthritis (RA) is a major cause of adult chronic inflammatory arthritis and a typical complex trait. Although several genetic determinants have been identified, they account for only a part of the genetic susceptibility. We conducted a genome-wide association study of RA in Japanese using 225,079 SNPs genotyped in 990 cases and 1,236 controls from two independent collections (658 cases and 934 controls in collection1; 332 cases and 302 controls in collection2), followed by replication studies in two additional collections (874 cases and 855 controls in collection3; 1,264 cases and 948 controls in collection4). SNPs showing p<0.005 in the first two collections and p<10(-4) by meta-analysis were further genotyped in the latter two collections. A novel risk variant, rs2000811, in intron2 of the myelin basic protein (MBP) at chromosome 18q23 showed strong association with RA (p = 2.7×10(-8), OR 1.23, 95% CI: 1.14-1.32). The transcription of MBP was significantly elevated with the risk allele compared to the alternative allele (p<0.001). We also established by immunohistochemistry that MBP was expressed in the synovial lining layer of RA patients, the main target of inflammation in the disease. Circulating autoantibody against MBP derived from human brain was quantified by ELISA between patients with RA, other connective tissue diseases and healthy controls. As a result, the titer of anti-MBP antibody was markedly higher in plasma of RA patients compared to healthy controls (p<0.001) and patients with other connective tissue disorders (p<0.001). ELISA experiment using citrullinated recombinant MBP revealed that a large fraction of anti-MBP antibody in RA patients recognized citrullinated MBP. This is the first report of a genetic study in RA implicating MBP as a potential autoantigen and its involvement in pathogenesis of the disease.

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Allelic difference in MBP transcription using allele specific quantitative RT-PCR.The amount of MBP primary transcripts transcribed from chromosomes carrying rs2000811 risk (T) and alternative (C) alleles was compared in each cell line, and the ratio (T/C) was plotted. Genomic DNA was used as a control for equimoler biallelic representation. Experiments were done twice independently.
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pone-0020457-g002: Allelic difference in MBP transcription using allele specific quantitative RT-PCR.The amount of MBP primary transcripts transcribed from chromosomes carrying rs2000811 risk (T) and alternative (C) alleles was compared in each cell line, and the ratio (T/C) was plotted. Genomic DNA was used as a control for equimoler biallelic representation. Experiments were done twice independently.

Mentions: Quantitative RT-PCR experiments showed only very low levels of MBP expression in RNA from Epstein-Barr virus (EBV)-transformed human B-lymphoblastoid cell lines, and we could detect no discernable correlation of MBP transcript levels with different risk genotypes (p = 0.36, Figure S4). By a similar reason, in-silico expression analysis using GEO database did not return clear association [28]. However, when we performed allele-specific quantitative RT-PCR [16] using genomic DNA and cDNA of these cell lines, we observed elevated allele-specific transcription associated with the risk allele (p<0.001, Figure 2, for detailed procedure, see Materials and Methods). This suggests that rs2000811, and/or other variants in linkage disequilibrium with this marker, impact the quantitative pattern of MBP transcription. However, bioinformatics analysis identified no known cis-acting elements covering rs2000811 that could be inferred to have functional effects (Method S2). In addition, the alignment of the 4-kb region ranging between 2-kb centromeric and 2-kb telomeric to rs2000811 revealed that this segment has very low interspecies conservation among placental mammals.


Myelin basic protein as a novel genetic risk factor in rheumatoid arthritis--a genome-wide study combined with immunological analyses.

Terao C, Ohmura K, Katayama M, Takahashi M, Kokubo M, Diop G, Toda Y, Yamamoto N, Human Disease Genomics Working GroupRheumatoid Arthritis (RA) Clinical and Genetic Study ConsortiumShinkura R, Shimizu M, Gut I, Heath S, Melchers I, Manabe T, Lathrop M, Mimori T, Yamada R, Matsuda F - PLoS ONE (2011)

Allelic difference in MBP transcription using allele specific quantitative RT-PCR.The amount of MBP primary transcripts transcribed from chromosomes carrying rs2000811 risk (T) and alternative (C) alleles was compared in each cell line, and the ratio (T/C) was plotted. Genomic DNA was used as a control for equimoler biallelic representation. Experiments were done twice independently.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3108877&req=5

pone-0020457-g002: Allelic difference in MBP transcription using allele specific quantitative RT-PCR.The amount of MBP primary transcripts transcribed from chromosomes carrying rs2000811 risk (T) and alternative (C) alleles was compared in each cell line, and the ratio (T/C) was plotted. Genomic DNA was used as a control for equimoler biallelic representation. Experiments were done twice independently.
Mentions: Quantitative RT-PCR experiments showed only very low levels of MBP expression in RNA from Epstein-Barr virus (EBV)-transformed human B-lymphoblastoid cell lines, and we could detect no discernable correlation of MBP transcript levels with different risk genotypes (p = 0.36, Figure S4). By a similar reason, in-silico expression analysis using GEO database did not return clear association [28]. However, when we performed allele-specific quantitative RT-PCR [16] using genomic DNA and cDNA of these cell lines, we observed elevated allele-specific transcription associated with the risk allele (p<0.001, Figure 2, for detailed procedure, see Materials and Methods). This suggests that rs2000811, and/or other variants in linkage disequilibrium with this marker, impact the quantitative pattern of MBP transcription. However, bioinformatics analysis identified no known cis-acting elements covering rs2000811 that could be inferred to have functional effects (Method S2). In addition, the alignment of the 4-kb region ranging between 2-kb centromeric and 2-kb telomeric to rs2000811 revealed that this segment has very low interspecies conservation among placental mammals.

Bottom Line: The transcription of MBP was significantly elevated with the risk allele compared to the alternative allele (p<0.001).We also established by immunohistochemistry that MBP was expressed in the synovial lining layer of RA patients, the main target of inflammation in the disease.This is the first report of a genetic study in RA implicating MBP as a potential autoantigen and its involvement in pathogenesis of the disease.

View Article: PubMed Central - PubMed

Affiliation: Center for Genomic Medicine, Graduate School of Medicine, Kyoto University, Kyoto, Japan.

ABSTRACT
Rheumatoid arthritis (RA) is a major cause of adult chronic inflammatory arthritis and a typical complex trait. Although several genetic determinants have been identified, they account for only a part of the genetic susceptibility. We conducted a genome-wide association study of RA in Japanese using 225,079 SNPs genotyped in 990 cases and 1,236 controls from two independent collections (658 cases and 934 controls in collection1; 332 cases and 302 controls in collection2), followed by replication studies in two additional collections (874 cases and 855 controls in collection3; 1,264 cases and 948 controls in collection4). SNPs showing p<0.005 in the first two collections and p<10(-4) by meta-analysis were further genotyped in the latter two collections. A novel risk variant, rs2000811, in intron2 of the myelin basic protein (MBP) at chromosome 18q23 showed strong association with RA (p = 2.7×10(-8), OR 1.23, 95% CI: 1.14-1.32). The transcription of MBP was significantly elevated with the risk allele compared to the alternative allele (p<0.001). We also established by immunohistochemistry that MBP was expressed in the synovial lining layer of RA patients, the main target of inflammation in the disease. Circulating autoantibody against MBP derived from human brain was quantified by ELISA between patients with RA, other connective tissue diseases and healthy controls. As a result, the titer of anti-MBP antibody was markedly higher in plasma of RA patients compared to healthy controls (p<0.001) and patients with other connective tissue disorders (p<0.001). ELISA experiment using citrullinated recombinant MBP revealed that a large fraction of anti-MBP antibody in RA patients recognized citrullinated MBP. This is the first report of a genetic study in RA implicating MBP as a potential autoantigen and its involvement in pathogenesis of the disease.

Show MeSH
Related in: MedlinePlus