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Search for the regulatory factor of apoptosis in adult T-cell leukemia (ATL) cells

寻找成人T细胞白血病（ATL）细胞凋亡的调节因子

基本信息

批准号：
12670993
负责人：
MAEDA Takahiro
金额：
$ 2.18万
依托单位：
Nagasaki University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (C)
财政年份：
2000
资助国家：
日本
起止时间：
2000 至 2001
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-12670993/
关键词：
Fas adult T-cell leukemia ATL apoptosis genetic mutation trimer

项目摘要

1. Anti-Fas monoclonal antibody (mAb) resistant adult T-cell leukemia (ATL) cell line, RS04RS04 RS04 cells did not show apoptosis after treatment with IgM anti-Fas mAb. We found that the lactacystin, which was a specific inhibitor of the proteosome, activated the caspase 8 and induced apoptosis in RS04 cells, indicating that the apoptotic pathway in the cytoplasm in RS04 cells was intact. By sequencing analysis of Fas gene, we found the heterozygous missense mutation with the transition of A to G at exon 2, resulting in substitution of arginine for histidine. The three dimensional molecular modeling of the mutated Fas protein indicated the definitive conformational alteration around the amino acids 52 to 58. It was suggested that the trimerization of Fas molecules was interfered with the mutated Fas, resulting in the impairment of signal transduction of apoptosis by dominant negative mechanism.2. Anti-Fas mAb resistantATL cell line, OMTIn OMT cells, we found the missense point mutation at nucleotide 1132 with the transition of T to C, resulting in the substitution of threonine for isoleucine. This mutation might cause the conformational alteration of death domain and fail to associate with the FADD molecule, which located at right downstream under the Fas molecule.3. ATL cells which barely express membrane FasBecause the Fas gene mutations were detected at the relatively high rate in Fas-resistant ATL cell lines, we investigated the primary ATL cells, which barely expressed membrane Fas and were resistant to IgM anti-Fas mAb. In four of seven cases of patients with ATL, we found Fas gene mutations, three point mutations and a 2 base pair deletion. It was indicated that the various mutations of Fas gene were one of the mechanisms of preventing the apoptosis in ATL cells.

1.抗Fas单克隆抗体（mAb）耐药的成人T细胞白血病（ATL）细胞株RS04 RS04经IgM抗Fas mAb处理后未出现凋亡。我们发现蛋白酶体的特异性抑制剂lactacystin可激活caspase 8并诱导RS04细胞凋亡，表明RS04细胞胞质内的凋亡途径是完整的。通过对Fas基因测序分析，发现该基因第2外显子存在A → G的杂合错义突变，导致组氨酸被精氨酸取代。突变的Fas蛋白的三维分子建模表明，围绕氨基酸52至58的明确的构象变化。提示突变的Fas干扰了Fas分子的三聚化，通过显性负性机制阻碍了凋亡信号转导.抗Fas单克隆抗体耐药ATL细胞株OMT中，我们发现OMT细胞中第1132位核苷酸发生错义突变，由T变为C，导致异亮氨酸被苏氨酸取代。该突变可能导致死亡结构域的构象改变，不能与位于Fas分子下游的FADD分子结合.由于Fas耐药的ATL细胞系中Fas基因突变率相对较高，我们研究了原代ATL细胞，这些细胞几乎不表达膜Fas，并对IgM抗Fas mAb具有耐药性。在7例ATL患者中，我们发现4例Fas基因突变，3个点突变和2个碱基对缺失。提示Fas基因的各种突变是抑制ATL细胞凋亡的机制之一。