Detection of TP53 mutation in ameloblastoma by the use of a yeast functional assay.
Detection of TP53 mutation in ameloblastoma by the use of a yeast functional assay.
复制标题
使用酵母功能测定法检测成釉细胞瘤中的 TP53 突变。
DOI：
10.1034/j.1600-0714.2002.00006.x
复制
发表时间：
2002
期刊：
影响因子：
--
通讯作者：
T. Moriuchi
中科院分区：
文献类型：
--
作者：
T. Shibata;D. Nakata;I. Chiba;T. Yamashita;Y. Abiko;M. Tada;T. Moriuchi
背景
TP 53的突变已在包括口腔病变在内的多种肿瘤中观察到，但在成釉细胞瘤中尚无报道。本研究的目的是使用新开发的酵母功能检测法检测成釉细胞瘤的TP 53状态，其准确性和灵敏度已被证明高于先前的基于DNA结构的方法，如单链构象多态性（SSCP）分析。
方法
采用酵母功能分析法和DNA测序法对12例经组织学诊断的成釉细胞瘤进行了TP 53基因的检测。从冷冻组织样品中提取RNA后，进行逆转录（RT）-PCR，并使用这些样品。该测定可通过蛋白质的DNA结合活性检测密码子67和347之间的p53 mRNA突变，并将其显示为红色菌落。
结果
一例47岁男性患者的红色酵母菌菌落率为17%，其他11例患者的红色酵母菌菌落率为5.4%（平均值;范围，3-8%）。为了证实这一结果，我们从17%红色菌落的情况下获得了另外9个样品，其中含有或不含有肿瘤组织，并通过测定法对它们进行了分析。7份肿瘤细胞组织学阴性的样本产生4.7%的红色菌落（平均值;范围，1-7%）。两个肿瘤细胞组织学阳性的样品产生20%或46%的红色菌落。从显示高红色菌落比率的这三个样品的红色菌落中回收p53质粒，纯化后进行测序分析。在所有这些样品中，证实了TGT（Cys）238达特（Tyr）的相同克隆突变。
结论
这些结果表明，TP 53突变可能参与分子发病机制的成釉细胞瘤的一个子集，虽然它是罕见的。
BACKGROUND
Mutations in TP53 have been observed in a variety of tumors including oral lesions, though there are no reports in ameloblastomas. The purpose of the study was to examine the TP53 status of ameloblastomas using newly developed yeast functional assay whose accuracy and sensitivity has been proven to be higher than those of the previous DNA structure-based methods such as single strand conformation polymorphism (SSCP) analysis.
METHODS
TP53 status was analyzed by yeast functional assay and DNA sequencing in 12 cases of ameloblastoma which were diagnosed histologically and represented the clinical features of a benign tumor. After the extraction of RNA from the frozen tissue samples without microdissection, reverse transcription (RT)-PCR was carried out and these samples were used. The assay can detect mutations of p53 mRNA between codons 67 and 347 by the DNA-binding activity of the protein and reveal them as red colonies.
RESULTS
One case of 47-year-old male gave 17% red colonies of yeast and the other 11 cases gave 5.4% (mean; range, 3-8%). To confirm this result, we obtained other nine samples from the case of 17% red colonies, which contained or did not contain tumor tissues, and analyzed them by the assay. Seven samples that were histologically negative for tumor cells gave 4.7% red colonies (mean; range, 1-7%). Two samples that were histologically positive for tumor cells gave 20 or 46% of red colonies. The p53 plasmids were recovered from the red colonies of these three samples showing high red colony ratios and were subjected to sequencing analysis after purification. In all these samples, the same clonal mutation of TGT (Cys) 238 TAT (Tyr) was demonstrated.
CONCLUSIONS
These results suggest that TP53 mutation may be involved in molecular pathogenesis in a subset of ameloblastomas, though it is infrequent.