The influence of genetic polymorphisms in the ccr5promoter on surface CCR5 expression

• Main Author: 呂淑娟
• Other Authors: 黎慶
• Format: Others
• Language: zh-TW
• Published: 2000
• Online Access: http://ndltd.ncl.edu.tw/handle/80595050373524687341

碩士 === 國立成功大學 === 微生物暨免疫學研究所 === 88 === The ccr5 gene encodes a β-chemokine receptor that also serves as the major coreceptor for HIV R5 strains. Common allelic variants in this gene influence the rates of AIDS progression. The nucleotide polymorphism at the position 59029 (G to A) (ccr5-p59029 G/A) or 59653 (C to T) (ccr5-p59653 C/T) in the ccr5 promoter was confirmed associated with faster or slower HIV disease progression, respectively, and that has been well documented. Recently, the ccr5 promoter has been found to contain 10 additional nucleotide changes that can be categorized into 10 haplotypes (P1-P10), and the P1 haplotype is correlated to accelerated HIV disease progression. Since the heterozygous 32-bp-deletion in the ccr5 coding region in the Caucasians was related to the reduction of the surface CCR5 expression as well as to slower disease progression, we suggest that P1 promoter may expresses more surface CCR5 expression resulting in the increasing rate of HIV-1 spreading in vivo, as compared with other haplotypes. In order to confirm this hypothesis, we defined the ccr5 promoter haplotypes in 92 healthy, unrelated Chinese in Taiwan with automated nucleotide sequence analysis. The level of the surface CCR5 expression on PBMC from 61 individuals in this studied cohort was determined by flow cytometry. The result showed that only three haplotypes, P1, P2, and P4 were identified in this group, and the frequencies of P1/P1, P1/P2, P1/P4 and P4/P4 equaled to 20.65%, 1.08%, 41.30% and 36.96%, respectively. We further correlated the genotype with the levels of CCR5 expression on cell surface. There were no significant differences between the promoter genotypes (P1/P1, P1/P4 and P4/P4) and CCR5 expression levels had been detected. Interestingly, from the sequenced data, the P1 haplotype was found to be in linkage-disequilibrium with ccr5-p59029 A. Since p59029 A drives 2-fold greater CAT activity than p59029 G in vitro, we speculated that P1 has a stronger promoter activity. We further analyzed the correlation of the three ccr5-p59653 genotypes (C/C, C/T and T/T) with the level CCR5 expression on PBMC derived from P1 homozygote. From this FACScan data, we found a significantly lower percentage of PBMC and CD4+ cells expressing CCR5 which had two 59653 T alleles (p＜0.05). According to the results of gel retardation, the different binding pattern factors were detected due to the polymorphism of p59029 (G/A) or p59653 (C/T). Furthermore, we found that p59653 T may inhibit one or more cellular factor(s) from binding to p59029 A site and that may be resulted in reducing surface CCR5 expression. Finally, we used microarray hybridization to study the gene expression during monocyte (U937 cell) differentiation. In conclusion, a higher frequency of P1/P1 genotype was detected in Taiwanese population. The nucleotide polymorphism of ccr5-p59653 seemed influence the activity of ccr5 promoter.