| Budget Amount *help |
¥3,800,000 (Direct Cost: ¥3,800,000)
Fiscal Year 2001: ¥1,700,000 (Direct Cost: ¥1,700,000)
Fiscal Year 2000: ¥2,100,000 (Direct Cost: ¥2,100,000)
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| Research Abstract |
Human endogenous retroviruses (HERV) sequences are dispersed throughout the human genome, which may be useful as a genetic marker for human genome research. In addition, it is possible that HERV may contribute to the pathogenesis of autoimmune diereses, either by activating the surrounding genes via its promoter activiites or generating virus proteins and thereby yielding immune complexes. Since demethylation of CpG islands is Associated with transcriptional activities, demethylated HERV may be transcriptionally active in vivo. In the present study, we investigated the methylation pattern of genes surrounding HERV, using suppression PCR method, for the purpose of obtaining the information as to the HERV associated with autoimmune diseases. In brief, genomic DNA was digested with methylation-sensitive restriction enzyme, HpaII, and was ligated to adapter primer. PCR was subsequently performed using adapter and HERV-specific primer. As control, methylation-independent enzyme, Mspl, was used. The fragment was ananalyzed by an automated sequencer. When we compare the results between DNA isolated from neutrophils and that of lymph nodes, the pattern of MspI fragment was the same, indicating that there exists no genetic difference, in terms of HpaII/MspI RFLP, between the samples. In contrast, HpaII fragment Pattern was different from each other, presumably reflecting the epigenetic difference (methylation) between them. In RA, HpaII fragment pattern of peripheral blood was different from that of synovila fluid cells. There was also a difference of HpaII pattern of neutrophils, before and after anti-rheumatic treatments. Suppression PCR may thus be a convenient way, to study the difference of methylation pattern around HERV in various autoimmune diseases,
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