Suppression of lung cancer cell growth by ribozyme-mediated modification of p53 pre-mRNA.
Suppression of lung cancer cell growth by ribozyme-mediated modification of p53 pre-mRNA.
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发表时间：
1995-09
影响因子：
6.4
通讯作者：
D. Cai;T. Mukhopadhyay;J. Roth
中科院分区：
文献类型：
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作者：
D. Cai;T. Mukhopadhyay;J. Roth
一种被设计用于切割p53前信使RNA （mRNA）的抗p53核酶（催化RNA）可以有效地降低内源性突变p53 mRNA的水平。逆转录病毒介导的双锤头核酶（Rz5a）在人类肺癌细胞系中被设计用于切割密码子5和外显子6边界附近密码子187处未剪接的p53 RNA，降低了在密码子254处含有纯合p53突变的h26br细胞中p53 RNA和蛋白的突变水平。体外实验表明，核酶对p53前mRNA有催化裂解作用，但对p53 mRNA无催化裂解作用。这种核酶对p53前mrna的切割是特异性的，因为它的催化结构域（Rz5m）的突变使其在体外失去了切割活性。Rz5a核酶的表达显著抑制H226Br细胞的生长。然而，另一种靶向p53基因密码子264靠近7号内含子和8号外显子边界的核酶（Rz7a）在体外可以切割pre-mRNA，但不抑制细胞生长。p53前mrna的修饰位点可能决定了该细胞系中核酶介导的生长抑制程度。我们发现p53前mrna可以被体内特定的核酶修饰，这表明这些药物可能在癌症基因治疗策略中发挥作用。
An anti-p53 ribozyme (catalytic RNA) designed to cleave the p53 pre-messenger RNA (mRNA) can efficiently reduce the level of endogenous mutant p53 mRNA. Retrovirus-mediated transduction of a hammerhead ribozyme (Rz5a) designed to cleave unspliced p53 RNA at codon 187 near the boundary of intron 5 and exon 6 reduced the level of mutant p53 RNA and protein in the human H226Br lung cancer cell line, which contains a homozygous p53 mutation at codon 254. The catalytic cleavage of the p53 pre-mRNA but not the p53 mRNA by the ribozyme was shown in vitro. The cleavage of the p53 pre-mRNA by this ribozyme was specific because a mutation in its catalytic domain (Rz5m) abolished the cleavage activity in vitro. Expression of the Rz5a ribozyme significantly suppressed the growth of the H226Br cells in culture. However, another ribozyme (Rz7a) targeted at codon 264 of the p53 gene near the boundary of intron 7 and exon 8 showed in vitro cleavage of the pre-mRNA but did not suppress cell growth. The site of modification in the p53 pre-mRNA may determine the degree of ribozyme-mediated growth suppression in this cell line. Our findings that p53 pre-mRNA can be modified by a specific ribozyme in vivo suggest a possible role for these agents in gene therapy strategies for cancer.