The ability of a reconstituted cell-free system to transport mRNA as a ribonucleoprotein particle has been examined. Poly(A) messenger ribonucleoproteins (mRNPs), UV cross-linked after release from isolated liver nuclei in a cell-free system, exhibited a buoyant density of 1.33 g/cm3 in cesium sulfate and 1.47 g/cm3 in cesium chloride, values identical to those of poly(A) mRNP isolated directly from liver polysomes. Furthermore, the in vivo and in vitro transported mRNP showed a similar degree of resistance to RNase digestion and had sedimentation coefficients approximately 2.5 times that of the isolated mRNA. Release of both total mRNA and alpha 2 mu-globulin mRNA was proportional to the concentration of a specific cytoplasmic protein. Removal of the transport proteins from the cytosol with streptomycin sulfate provided a basal system incapable of supporting the active transport of alpha 2 mu-globulin mRNA. Hybridization of released RNA with a recombinant probe specific for intron 6 of alpha 2 mu-globulin showed that intron sequences were retained within the nucleus under optimal alpha 2 mu-globulin mRNA transport conditions and that the transported alpha 2 mu-globulin mRNA was of mature size.
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机译:已经检查了重组的无细胞系统将mRNA作为核糖核蛋白颗粒转运的能力。聚(A)信使核糖核蛋白(mRNP),在无细胞系统中从分离的肝核释放后通过紫外线交联,在硫酸铯中的浮力密度为1.33 g / cm3,在氯化铯中的浮力密度为1.47 g / cm3,值相同直接从肝脏多核糖体中分离出的poly(A)mRNP。此外,体内和体外转运的mRNP对RNase消化的抵抗力相似,沉降系数约为分离的mRNA的2.5倍。总mRNA和α2μ-球蛋白mRNA的释放与特定细胞质蛋白的浓度成比例。用硫酸链霉素从胞质溶胶中去除转运蛋白提供了基础系统,该系统无法支持α2 mu-globulin mRNA的主动转运。释放的RNA与对α2 mu-globulin的内含子6特异的重组探针的杂交表明,内含子序列在最佳α2 mu-globulin mRNA的运输条件下保留在细胞核内,并且运输的α2 mu-globulin mRNA的成熟尺寸。
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