Walker K K, Levine A J
Department of Molecular Biology, Prineton University, NJ 08544, USA.
Proc Natl Acad Sci U S A. 1996 Dec 24;93(26):15335-40. doi: 10.1073/pnas.93.26.15335.
Activation of the p53 tumor suppressor protein has been demonstrated to block cell growth by inducing either a transient cell cycle arrest or programmed cell death (apoptosis). Although evidence exists linking p53's function as an activator of transcription to its ability to effect cell cycle arrest, the role of this activity in the induction of apoptosis remains unclear. To gain insight into the molecular mechanisms underlying p53-mediated antiproliferative pathways, a study was initiated to explore the functions of a putative p53 signaling domain. This region of the human p53 protein is localized between amino acids 61 and 94 (out of 393) and is noteworthy in that it contains five repeats of the sequence PXXP (where P represents proline and X any amino acid). This motif has been shown to play a role in signal transduction via its SH3 domain binding activity. A p53 cDNA deletion mutant (delta pro AE), which lacks this entire proline-rich domain (deleted for amino acids 62-91), was created and characterized for a variety of p53 functions. The entire domain has been shown to be completely dispensable for transcriptional activation. On the other hand, this deletion of the p53 proline-rich domain impairs p53's ability to suppress tumor cell growth in culture. Amino acid substitution mutations at residues 22 and 23 of p53 (eliminates transcriptional activity) also impair p53-mediated inhibition of cell growth in culture. Unlike wild-type p53, the delta proAE mutant cDNA can be stably expressed in tumor derived cell lines with few immediate detrimental effects. These cells express physiologic levels of p53 protein that are induced normally in response to DNA damage, indicating that removal of the proline-rich domain does not disrupt p53's upstream regulation by DNA damage. These data indicate that, in addition to the transcriptional activation domain, the p53 proline-rich domain plays a critical role in the transmission of antiproliferative signals down-stream of the p53 protein and may link p53 to a direct signal transduction pathway.
p53肿瘤抑制蛋白的激活已被证明可通过诱导短暂的细胞周期停滞或程序性细胞死亡（凋亡）来阻断细胞生长。尽管有证据表明p53作为转录激活因子的功能与其影响细胞周期停滞的能力有关，但其在诱导凋亡中的作用仍不清楚。为深入了解p53介导的抗增殖途径的分子机制，启动了一项研究以探索一个假定的p53信号结构域的功能。人p53蛋白的这一区域位于氨基酸61至94之间（共393个氨基酸），值得注意的是它包含序列PXXP的五个重复（其中P代表脯氨酸，X代表任何氨基酸）。该基序已被证明通过其SH3结构域结合活性在信号转导中发挥作用。构建了一个p53 cDNA缺失突变体（delta pro AE），它缺少整个富含脯氨酸的结构域（缺失氨基酸62 - 91），并对其多种p53功能进行了表征。已证明整个结构域对于转录激活是完全不必要的。另一方面，p53富含脯氨酸结构域的缺失损害了p53在培养中抑制肿瘤细胞生长的能力。p53第22和23位残基的氨基酸替代突变（消除转录活性）也损害了p53在培养中介导的细胞生长抑制。与野生型p53不同，delta proAE突变体cDNA可以在肿瘤衍生细胞系中稳定表达，几乎没有直接的有害影响。这些细胞表达的p53蛋白水平在生理范围内，并且在DNA损伤时正常诱导，表明去除富含脯氨酸的结构域不会破坏DNA损伤对p53的上游调控。这些数据表明，除了转录激活结构域外，p53富含脯氨酸的结构域在p53蛋白下游抗增殖信号的传递中起关键作用，并且可能将p53与直接的信号转导途径联系起来。
