摘要
碱性神经酰胺酶2(alkaline ceramidase 2,ACER2)是一个参与脂质类信号分子神经酰胺代谢的酶分子,在细胞增殖、衰老和凋亡等过程中起重要作用。为了进一步研究ACER2基因的转录调控机制,该研究克隆鉴定了ACER2基因的启动子。首先,应用5'RACE(rapid amplification of cDNA ends,cDNA末端快速扩增)技术鉴定了ACER2基因的转录起始位点。然后,通过PCR定向克隆和定点突变策略,构建了三个长度不同的覆盖ACER2基因5'端侧翼区起始密码子ATG上游约1.3Kb区域的一系列ACER2基因启动子荧光素酶报告基因重组体。启动子活性分析表明ACER2基因启动子定位于转录起始位点附近约670 bp的区域内。转录因子结合位点分析结果表明,ZCER2基因启动子含有Sp1、GATA-1和AP-1等潜在的转录因子结合位点,提示Sp1和GATA-1等转录因子可能参与ACER2基因的转录调控。
Alkaline ceramidase 2, ACER2, is a ceramide-hydrolyzing enzyme which plays an important role in cell proliferation, senescence, and apoptosis etc. To further investigate its transcriptional regulatory mechanism, ACER2 gene promoter has been cloned and identified in the present study. At first, the transcriptional start sites for ACER2 gene have been identified by 5＇ RACE （rapid amplification of cDNA ends） and bioinformatic analysis. Fur- thermore, three overlapping genomic fragments from the 5＇-flanking region ofACER2 gene have been coloned into pGL3-basic vector to construct ACER2 promoter reporters. Luciferase reporter assay indicated that ACER2 pro-moter region is mainly located in a 670 bp region nearby the transcriptional start site. Transcriptional factor binding analysis indicated that, ACER2 gene promoter contains several putative binding sites for transcriptional factors such as Spl and GATA-1. These results suggested that transcriptional factors such as Spl and GATA-1 might be invovled in the transcriptional regualtion of ACER2 gene.
出处
《中国细胞生物学学报》
CAS
CSCD
2011年第6期674-680,共7页
Chinese Journal of Cell Biology
基金
国家自然科学基金(No.30801356
No.81001097)
重庆市科委自然科学基金(No.2010BB5357)
重庆市教委科学技术研究(No.KJ090305)资助项目~~
