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首页> 外文期刊>Molecular and Cellular Biology >Wild-type human p53 and a temperature-sensitive mutant induce Fas/APO-1 expression.
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Wild-type human p53 and a temperature-sensitive mutant induce Fas/APO-1 expression.

机译：野生型人p53和温度敏感突变体诱导Fas / APO-1表达。

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Fas/APO-1 is a cell surface protein known to trigger apoptosis upon specific antibody engagement. Because wild-type p53 can activate transcription as well as induce apoptosis, we queried whether p53 might upregulate Fas/APO-1. To explore this possibility, we examined human p53-null (H358 non-small-cell lung adenocarcinoma and K562 erythroleukemia) and wild-type p53-containing (H460 non-small-cell lung adenocarcinoma) cell lines. When H358 or H460 cells were transduced with a replication-deficient adenovirus expression construct containing the human wild-type p53 gene but not with vector alone, a marked upregulation (approximately a three-to fourfold increase) of cell surface Fas/APO-1 was observed by flow cytometry. Similarly, K562, cells stably transfected with a plasmid vector containing the temperature-sensitive human p53 mutant Ala-143 demonstrated a four- to sixfold upregulation of Fas/APO-1 by flow-cytometric analysis at the permissive temperature of 32.5 degrees C. Temperature-sensitive upregulation of Fas/APO-1 in K562 Ala-143 cells was verified by immunoprecipitation and demonstrated to result from enhanced mRNA production by nuclear run-on and Northern (RNA) analyses. Stably transfected K562 cells expressing temperature-insensitive, transcriptionally inactive p53 mutants (His-175, Trp-248, His-273, or Gly-281) failed to upregulate Fas/APO-1 at either 32.5 degrees or 37.5 degrees C. The temperature-sensitive transcription of Fas/APO-1 occurred in the presence of cycloheximide, indicating that de novo protein synthesis was not required and suggested a direct involvement of p53. Collectively, these observations argue that Fas/APO-1 is a target gene for transcriptional activation by p53.
机译：Fas / APO-1是一种细胞表面蛋白，已知可在特异性抗体参与后触发凋亡。由于野生型p53可以激活转录并诱导细胞凋亡，因此我们询问p53是否可能上调Fas / APO-1。为了探索这种可能性，我们检查了人p53无效（H358非小细胞肺腺癌和K562红白血病）和野生型含p53（H460非小细胞肺腺癌）细胞系。当用含有人野生型p53基因的复制缺陷型腺病毒表达构建体转导H358或H460细胞，但不单独用载体转导H358或H460细胞时，细胞表面Fas / APO-1明显上调（大约增加了三到四倍）。通过流式细胞仪观察。类似地，用含有温度敏感型人p53突变体Ala-143的质粒载体稳定转染的K562细胞在32.5摄氏度的允许温度下通过流式细胞术分析显示Fas / APO-1上调了4至6倍。免疫沉淀法验证了K562 Ala-143细胞中Fas / APO-1的敏感上调，并证明是通过核电泳和Northern（RNA）分析提高了mRNA产生的结果。表达温度不敏感，无转录活性的p53突变体（His-175，Trp-248，His-273或Gly-281）的稳定转染的K562细胞在32.5摄氏度或37.5摄氏度下均无法上调Fas / APO-1。 Fas / APO-1的敏感转录在环己酰亚胺的存在下发生，这表明不需要从头合成蛋白质，表明p53的直接参与。总的来说，这些发现认为Fas / APO-1是p53转录激活的靶基因。

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