Leis J P, Scheible P, Smith R E
J Virol. 1980 Sep;35(3):722-31. doi: 10.1128/JVI.35.3.722-731.1980.
We purified the p19 proteins from the Prague C strain of Rous sarcoma virus, avian myeloblastosis virus, B77 sarcoma virus, myeloblastosis-associated virus-2(0), and PR-E 95-C virus and measured their binding affinities for 60S viral RNA by the nitrocellulose filter binding technique. The apparent association constants of the p19 proteins from Rous sarcoma virus Prague C, avian myeloblastosis virus, and B77 sarcoma virus for homologous and heterologous 60S RNAs were similar (1.5 x 10(11) to 2.6 x 10(11) liters/mol), whereas those of myeloblastosis-associated virus-2(0) and PR-E 95-C virus were 10-fold lower. The sizes and relative amounts of the virus-specific polyadenylic acid-containing RNAs in the cytoplasms of cells infected with Rous sarcoma virus Prague C, myeloblastosis-associated virus-2(0), and PR-E 95-C virus were determined by fractionating the RNAs on agarose gels containing methylmercury hydroxide, transferring them to diazobenzyloxymethyl paper and hybridizing them to a 70-nucleotide complementary DNA probe. In cells infected with Rous sarcoma virus Prague C we detected 3.4 x 10(6)-, 1.9 x 10(6)-, and 1.1 x 10(6)-dalton RNAs, in PR-E 95-C virus-infected cells we detected 3.4 x 10(6)-, 1.9 x 10(6)- and 0.7 x 10(6)-dalton RNAs, and in cells infected with myeloblastosis-associated virus-2(0) we detected 3 x 10(6)- and 1.3 x 10(6)-dalton RNAs. Each of these RNA species contained RNA sequences derived from the 5' terminus of genome-length RNA, as evidenced by hybridization with the 5' 70-nucleotide complementary DNA. The ratios of subgenomic mRNA's to genome-length RNAs in cells infected with myeloblastosis-associated virus-2(0) and PR-E 95-C virus were three- to five-fold higher than the ratio in cells infected with Rous sarcoma virus Prague C. These results suggest that more processing of viral RNA in infected cells is correlated with lower binding affinities of the p19 protein for viral RNA, and they are consistent with the hypothesis that the p19 protein controls processing of viral RNA in cells.
我们从劳氏肉瘤病毒布拉格C株、禽成髓细胞瘤病毒、B77肉瘤病毒、成髓细胞瘤相关病毒2(0)以及PR-E 95-C病毒中纯化了p19蛋白，并通过硝酸纤维素滤膜结合技术测定了它们与60S病毒RNA的结合亲和力。劳氏肉瘤病毒布拉格C株、禽成髓细胞瘤病毒和B77肉瘤病毒的p19蛋白与同源和异源60S RNA的表观缔合常数相似(1.5×10¹¹至2.6×10¹¹升/摩尔)，而成髓细胞瘤相关病毒2(0)和PR-E 95-C病毒的表观缔合常数则低10倍。通过在含有氢氧化甲基汞的琼脂糖凝胶上对RNA进行分级分离，将其转移到重氮苄氧基甲基纸上，并与70个核苷酸的互补DNA探针杂交，测定了感染劳氏肉瘤病毒布拉格C株、成髓细胞瘤相关病毒2(0)和PR-E 95-C病毒的细胞胞质中病毒特异性含聚腺苷酸RNA的大小和相对含量。在感染劳氏肉瘤病毒布拉格C株的细胞中，我们检测到了3.4×10⁶、1.9×10⁶和1.1×10⁶道尔顿的RNA；在感染PR-E 95-C病毒的细胞中，我们检测到了3.4×10⁶、1.9×10⁶和0.7×10⁶道尔顿的RNA；在感染成髓细胞瘤相关病毒2(0)的细胞中，我们检测到了3×10⁶和1.3×10⁶道尔顿的RNA。通过与5'端70个核苷酸的互补DNA杂交证明，这些RNA种类中的每一种都含有源自基因组长度RNA 5'端的RNA序列。感染成髓细胞瘤相关病毒2(0)和PR-E 95-C病毒的细胞中亚基因组mRNA与基因组长度RNA的比率比感染劳氏肉瘤病毒布拉格C株的细胞中的比率高3至5倍。这些结果表明，感染细胞中病毒RNA的更多加工与p19蛋白对病毒RNA的较低结合亲和力相关，并且它们与p19蛋白控制细胞中病毒RNA加工的假说一致。
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