旨在克隆猪作用于RNA的腺苷脱氨酶2基因(ADAR2)全长cDNA序列,同时对该基因在猪不同组织中的表达规律进行探索。利用RACE(rapid-amplification of cDNA ends)对大白猪ADAR2基因mRNA全长序列进行克隆,并进行生物信息学分析;用荧光定量PCR...旨在克隆猪作用于RNA的腺苷脱氨酶2基因(ADAR2)全长cDNA序列,同时对该基因在猪不同组织中的表达规律进行探索。利用RACE(rapid-amplification of cDNA ends)对大白猪ADAR2基因mRNA全长序列进行克隆,并进行生物信息学分析;用荧光定量PCR方法检测35日龄大白猪心、肝、肺、肾、脾、脑、小肠、背最长肌和背部脂肪9种组织中ADAR2的表达水平。结果表明,猪ADAR 2基因cDNA全长6305 bp,共包含12个外显子,编码704个氨基酸,与人、黑猩猩、猕猴、长臂猿、黄牛、山羊和绵羊的CDS区核酸序列和氨基酸序列的一致性均在84%以上。该基因编码的蛋白含有2个双链RNA结合基序和一个脱氨酶结构域。猪ADAR2在检测的各组织中均表达,其中在肺中的表达量最高。综上所述,本研究成功克隆了猪ADAR2基因全长cDNA序列,并且发现其在猪体内广泛表达,为深入研究ADAR2的功能奠定了良好的基础。展开更多
OBJECTIVE To observe the effect of phenylacetate on the expression of RNA editing deaminase ADAR2 mRNA in glioma cells. METHODS Primary glial cells from human brain tissue and glioma U-251MG cells were cultured. The e...OBJECTIVE To observe the effect of phenylacetate on the expression of RNA editing deaminase ADAR2 mRNA in glioma cells. METHODS Primary glial cells from human brain tissue and glioma U-251MG cells were cultured. The expression of ADAR2 mRNA was detected by a semiquantitative reverse transcription-polymerase chain reaction (RT-PCR). The levels of ADAR2 mRNA expression before and after treatment with phenylacetate were tested by RT -PCR and image analysis. The level of ADAR2 gene expression was presented as the ratio expression rate (RER) of ADAR2 gene/β-actin based on computer image analysis. RESULTS The ADAR2 mRNA displayed mild expression in brain glial cells, and a high expression level in high-grade malignant U-251MG glioma cells. Computer image analysis showed that the RERs of the ADAR2 gene in the U-251MG cells before and after treatment with 4.0, and 5.0 mM phenylacetate for 8 h were 100.0, 73.5, 60.3, respectively. The expression of ADAR2 mRNA was decreased by phenylacetate in glioma U-251MG cells. CONCLUSION Phenylacetate can decrease the expression of ADAR2 mRNA in glioma cells, suggesting that phenylacetate, as a drug, may act on the course of RNA editing in gliomas.展开更多
The effect and mechanism of phenylacetic acid on the proliferation of pancreatic carcinoma cells were investigated in cultured pancreatic carcinoma BXPC-3 cells by means of 3-（4,5-dimethylthiazol-2-yl）-2,5-diphenylt...The effect and mechanism of phenylacetic acid on the proliferation of pancreatic carcinoma cells were investigated in cultured pancreatic carcinoma BXPC-3 cells by means of 3-（4,5-dimethylthiazol-2-yl）-2,5-diphenyltetrazolium bromide assay and flow cytometry assay.The results show that the treatment of pancreatic carcinoma cells with phenylacetic acid significantly inhibited the cell proliferation in time-dependent and dose-dependent manners.The proliferation of BXPC-3 cells was inhibited at the stage of S phase,the cells at the end stage of S phase were accumulated abundantly,and thus DNA synthesis could not be accomplished entirely.In addition,the expression of adenosine deaminases acting on RNA（ADARs） mRNA in BXPC-3 cells and pancreatic carcinoma specimen were detected by RT-PCR.Having been treated with phenylacetic acid,ADAR2 mRNA in BXPC-3 cells was significantly decreased,the differences were of statistical significance（P0.01）.Taken together,these results suggest that phenylacetic acid may likely regulate the proliferation of pancreatic carcinoma cells through the regulation of ADAR2 mRNA expression.展开更多
基金This study was supported by the National Nature Science Foundation of China (No. 30271334).
文摘OBJECTIVE To observe the effect of phenylacetate on the expression of RNA editing deaminase ADAR2 mRNA in glioma cells. METHODS Primary glial cells from human brain tissue and glioma U-251MG cells were cultured. The expression of ADAR2 mRNA was detected by a semiquantitative reverse transcription-polymerase chain reaction (RT-PCR). The levels of ADAR2 mRNA expression before and after treatment with phenylacetate were tested by RT -PCR and image analysis. The level of ADAR2 gene expression was presented as the ratio expression rate (RER) of ADAR2 gene/β-actin based on computer image analysis. RESULTS The ADAR2 mRNA displayed mild expression in brain glial cells, and a high expression level in high-grade malignant U-251MG glioma cells. Computer image analysis showed that the RERs of the ADAR2 gene in the U-251MG cells before and after treatment with 4.0, and 5.0 mM phenylacetate for 8 h were 100.0, 73.5, 60.3, respectively. The expression of ADAR2 mRNA was decreased by phenylacetate in glioma U-251MG cells. CONCLUSION Phenylacetate can decrease the expression of ADAR2 mRNA in glioma cells, suggesting that phenylacetate, as a drug, may act on the course of RNA editing in gliomas.
基金Supported by the National Natural Science Foundation of China(Nos.30801354 and 30970791)the Jilin Provincial Science & Technology Department,China(No.20080154)
文摘The effect and mechanism of phenylacetic acid on the proliferation of pancreatic carcinoma cells were investigated in cultured pancreatic carcinoma BXPC-3 cells by means of 3-（4,5-dimethylthiazol-2-yl）-2,5-diphenyltetrazolium bromide assay and flow cytometry assay.The results show that the treatment of pancreatic carcinoma cells with phenylacetic acid significantly inhibited the cell proliferation in time-dependent and dose-dependent manners.The proliferation of BXPC-3 cells was inhibited at the stage of S phase,the cells at the end stage of S phase were accumulated abundantly,and thus DNA synthesis could not be accomplished entirely.In addition,the expression of adenosine deaminases acting on RNA（ADARs） mRNA in BXPC-3 cells and pancreatic carcinoma specimen were detected by RT-PCR.Having been treated with phenylacetic acid,ADAR2 mRNA in BXPC-3 cells was significantly decreased,the differences were of statistical significance（P0.01）.Taken together,these results suggest that phenylacetic acid may likely regulate the proliferation of pancreatic carcinoma cells through the regulation of ADAR2 mRNA expression.
