WEB2基因在酿酒酵母S期检查点通路上调控RNR3基因表达  

RNR3 Gene Expression is Regulated by WEB2 Gene in S Checkpoint Pathway of S.cerevisiae

作　　者：李新鸣[1] 孙黎光[2] 刘萍[2] 付玥[2] 白抚生 宣忠信[2] 

机构地区：[1]中国医科大学基础医学院病原微生物教研教室,沈阳110001 [2]中国医科大学基础医学院生物化学与分子生物学教研室,沈阳110001 [3]辽宁省金秋医院神经内科,沈阳110016

出　　处：《细胞生物学杂志》2006年第1期71-75,共5页Chinese Journal of Cell Biology

基　　金：国家自然科学基金资助项目(No.39870384)~~

摘　　要：WEB2基因参与酿酒酵母S期检查点调控机制,而RNR3基因位于该调控通路末端,DNA损伤或合成阻断时,S期检查点通路诱导RNR3过度表达。因此,通过确定WEB2在该检查点通路上是否参与调控RNR3基因的表达,将有助于进一步明确WEB2基因在检查点通路上的工作位点,了解WEB2基因如何发挥检查点调控功能。构建RNR3-LacZ基因融合质粒,用于检测酵母细胞内RNR3基因的诱导性。诱导性可以通过测定β-半乳糖苷酶的活性而得知。利用DNA损伤药物甲磺酸甲酯(MMS)及DNA合成阻断剂羟基脲(HU)处理酵母细胞,测定WEB2基因突变株和野生株细胞内RNR3基因的诱导性。结果,WEB2突变株细胞中诱导活性分别增加(8.27±0.38)倍和(9.55±0.24)倍,而野生株分别增加了(83.32±2.42)倍和(124.67±2.87)倍。反映RNR3基因在WEB2突变株中的诱导性低于野生株。同RAD53突变株相比,后者的RNR3基因的诱导性更低,仅为(2.37±0.18)倍和(2.91±0.13)倍。说明WEB2基因突变影响S期检查点通路的信号传递至RNR3基因,所以在酿酒酵母S期检查点通路上,WEB2工作在RNR3基因上游,参与调控RNR3的表达,但调控能力不如RAD53基因强。Previous works suggest the role of WEB2 in S checkpoint regulation, it is necessary to make sure its location and interactions with other checkpoint genes in this signal transduction pathway. RNR3 gene is an important effector at the end of this pathway. The transcription of RNR3 gene is induced in response to DNA damage or DNA replication block. So we detect if WEB2 gene involves in RNR3 induction .The induction of RNR3 gene was investigated in S.cerevisiae using RNR3-LacZ fusion plasmid. Gene induction was monitored by measur- ing the β-galactosidase activity. When WEB2 mutant and wild type were exposed to hydroxyurea （HU） or methyl methanesulfonate （MMS）, induction of RNR3 were observed. The β-galactosidase activity is 8.27±0.38 or 9.55± 0.24 fold to basal levels in WEB2 mutant, but is far lower than in wild type, which are 83.32±2.42 or 124.67±2.87 ford. RAD53 mutant has lower induction of RNR3, 2.37±0.18 and 2.91 ±0.13 fold, respectively. The results suggest that WEB2 gene mutation blocks the checkpoint signal transduction to RNR3 gene. WEB2 gene acts upstream of RNR3 gene, less important than RAD53, may function together with other genes in regulation of RNR3 expression.

关 键 词：WEB2基因 S期检查点 RNR3基因 酿酒酵母 

分 类 号：Q932[生物学—微生物学]