Molecular pathology for down-regulation of erythroid-specific genes in prion diseases

朊病毒疾病中红细胞特异性基因下调的分子病理学

基本信息

批准号：
16208030
负责人：
INABA Mutsumi
金额：
$ 32.2万
依托单位：
Hokkaido University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (A)
财政年份：
2004
资助国家：
日本
起止时间：
2004 至 2006
项目状态：
已结题

项目摘要

AHSP is an erythroid-specific molecular chaperone that stabilizes newly synthesised a-globin. Previous studies demonstrated that mRNA levels of AHSP were specifically reduced in hematopoietic tissues of prion-infected animals. The purpose of the present study is to clarify the mechanism for down-regulation of AHSP transcription. A series of truncation and mutation analyses on 2.5-kb 5' upstream region of the AHSP gene in MELhide8 cells and electrophoretic mobility shift assay showed that the minimal 5'-promoter region located at-328-286 to the translation initiation site including a GATA binding motif. Either of two upstream GATA elements at-403 and-381 enhanced reporter gene transcription only in the presence of the minimal GATA element described above. These findings indicate that an erythroid-specific transcription factor GATA-1 is essential to AHSP gene expression and suggest that the down-regulation of AHSP involves changes in GATA-1 transcriptional activation. There was no significant change in gene expression of AHSP, a-globin, b-globin, GATA-1, EKLF, and NF-E2 in MELhide8 cells when the cells were incubated with brain homogenates from scrapie-infected mice for up to 120 hours. Moreover, MELhide8 cells exhibited no accumulation of PrPsc even after 16 passages. These data demonstrated that Prrc has no direct effect on AHSP gene expression in erythroid cells. Instead, IL-6 significantly and IL-1β weakly reduced the expression of AHSP mRNA levels and the AHSP promoter-reporter gene expression in MELhide8 cells in a dose-dependent manner. The reduction was recovered in the presence of the inhibitor of the STAT3 pathway, suggesting that the signal transduction of an inflammatory cytokine IL-6 through STAT3 pathway would modulate GATA-1/AHSP promoter interaction and subsequently causes down-regulation of the AHSP gene.

AHSP是一种红细胞特异性的分子链酮，可稳定新合成的A-珠蛋白。先前的研究表明，在原始动物的造血组织中，AHSP的mRNA水平明确降低。本研究的目的是阐明AHSP转录下调的机制。在MELHIDE8细胞中AHSP基因的2.5-kb 5'上游区域和电泳迁移率转移测定法上进行了一系列截短和突变分析，表明，位于翻译起始位点的最小5'Promoter区域AT-328-286 at-328-286，包括GATA结合基序。仅在上述最小的GATA元件的情况下，两个上游GATA元件中的任何一个AT-403和-381增强了基因转录。这些发现表明，红细胞特异性转录因子GATA-1对于AHSP基因表达至关重要，并表明AHSP的下调涉及GATA-1转录激活的变化。当Melhide8细胞中AHSP，A-Globin，B-珠蛋白，B-珠蛋白，GATA-1，EKLF和NF-E2的基因表达没有显着变化，当时将细胞与来自羊皮纸感染的小鼠的脑匀浆孵育长达120小时时。此外，即使经过16个传递，Melhide8细胞也没有表现出PRPSC的积累。这些数据表明，PRRC对红细胞细胞中AHSP基因的表达没有直接影响。取而代之的是，IL-6显着降低了AHSP mRNA水平的表达和以剂量依赖性方式在MELHIDE8细胞中AHSP启动子 - 重生基因表达。在STAT3途径的抑制剂存在下恢复了还原，这表明炎性细胞因子IL-6通过STAT3途径的信号转移会调节GATA-1/AHSP启动子的相互作用，并随后引起AHSP基因的下调。