摘要
目的通过比较不同细胞类型之间胰腺十二指肠同源盒1（Pdx-1）、配对盒基因4（Pax4）、MafA（mast cell function associated antigen）和Nkx6．1等胰岛组织特异性基因其转录起始区的H3K4m3和H3K9m3修饰的差异，探讨H3K4nd和H3K9m3修饰对胰岛组织特异性基因表达的作用。方法采用染色质免疫共沉淀．实时定量聚合酶链反应（PCR）法检测小鼠胚胎干细胞（mES，1×10^7）、小鼠成纤维细胞株NIH3T3细胞（1×10^7）和小鼠B细胞株NIT-1细胞（1×10^7）三者中的胰岛组织特异性基因、Oct4基因和MLH1基因转录起始区H3K4rrd和H3K9m3修饰的状况。同时采用实时定量逆转录（RT）-PCR检测上述3种细胞各基因mRNA表达水平。分析H3K4m3和H3K9m3修饰改变与基因表达之间的关系。结果NIT-1细胞中Pdx-1、Pax4、MafA、Nkx6．1等胰岛组织特异性基因转录起始区的H3K4m的修饰水平分别为：（4．84±0．05）％、（9．91±1．33）％、（10．64±0．87）％、（0．23±0．03）％，与mES细胞比较明显增高（P〈0．05），基因表达；NIH3T3细胞中Pdx-1、Pax4、MafA、Nkx6．1等胰岛组织特异性基因转录起始区的H3Kgm3的修饰水平分别为：（0．64±0．21）％、（7．04±1．29）％、（0．39±0．10）％、（2．35±0．81）％，与mES细胞比较明显增高（P〈0．05），基因不表达。结论mK4n13与H3K9m3修饰能相互协调，共同调控胰岛组织特异性基因的表达。
Objective To explore the regtdaiion of pancreatic islet-specific genes expression by histone modification （H3K4m3 and H3Kgm3）, through comparing the difference of H3K4m3 and H3Kgm3 modification in the transcription modification site of several pancreatic islet-specific genes （Pdx-1, Pax4, MafA, Nkx6.1, etc） among different cell types. Methods Chromatin immunoprecipitation-real time quantitative polymerase chain reaction （PCR） methods were used to study H3K4m3 and H3K9m3 modification in the transcription initiation site of pancreatic specific-islet genes, Oct4 and MLH1 genes, among mouse embryonic stem ceils （1×10^7）, NIH3T3 ceils （1 ×10^7） and NIT-1 cells （1×10^7）. At the same time, the mRNA levels of these genes were quantified among the above three ceil types using reverse transcription （RT）-PCR. The relationship between the differences in H3K4m3 and H3Kgm3 modification and gene expression was analyzed. Results （1） In N1T-1 cells, the levels of H3K4m3 modification in the transcription-initiation-site of Pdx-1, Pax4, MafA, Nkx6.1 were （4.84 ±0.05）%, （9.91 ± 1.33）%, （10.64 ±0. 87）%, and （0.23 ±0.03）% respectively. Compared with mES ceils, NIT-1 cells had a sigificantly higher level of H3K4m modification （P 〈0. 05）, with a sigificant level of gene expression; （2） In NIH3T3 ceils, the levels of H3K9m3 modification in the transcriptioninitiation-site of Pdx-1, Pax4, MafA, NEx6. 1 were （0.64 ± 0. 21 ）%, （7.04 ± 1.29）%, （0. 39 ± 0. 10）%, （2.35 ±0. 81）% respectively. Compared with ruES cells, NIH3T3 cell had a sigificanfly higher level of H3K9m3 modification （P 〈0.05）, with no gene expression. Conclusion H3K4m3 and H3K9m3 modification can regulate each other, and synergtically regulate the expression of pancreatic islet-specific genes.
出处
《中华实验外科杂志》
CAS
CSCD
北大核心
2010年第4期474-477,共4页
Chinese Journal of Experimental Surgery
关键词
H3K4m3
H3K9m3
基因表达
胰岛
分化
Histone 3 lysine 4 trimethylation
Histone 3 lysine 9 trimethylation
Gene expression
Islet
Differentiation
