The 30-kD subunit of mammalian cleavage and polyadenylation specificity factor and its yeast homolog are RNA-binding zinc finger proteins
The 30-kD subunit of mammalian cleavage and polyadenylation specificity factor and its yeast homolog are RNA-binding zinc finger proteins
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DOI：
10.1101/gad.11.13.1703
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发表时间：
1997-07-01
影响因子：
10.5
通讯作者：
Keller, W
中科院分区：
文献类型：
--
作者：
Barabino, SML;Hubner, W;Keller, W
切割和多聚腺苷酸化特异性因子（CPSF）是哺乳动物RNA 3 '端加工机制的关键组分，由160、100、73和30 kD的四个亚基组成。在这里，我们报告的分离和表征的cDNA编码的30 kD多肽。针对这种蛋白质产生的抗体抑制切割和多聚腺苷酸化，并与其他CPSF亚基共免疫沉淀。该蛋白质序列含有5个C3 H-锌指重复序列和一个假定的RNA结合锌指基序在羧基末端。与这一观察结果一致，体外翻译的30-kD蛋白结合RNA聚合物，对poly（U）具有明显的偏好。此外，还有一个必不可少的S. YTH 1基因在蛋白水平上与CPSF 30 K有40%的同源性。从条件性yth 1突变体制备的提取物具有正常的切割活性，但不能使上游切割产物聚腺苷酸化。通过加入纯化的聚腺苷酸化因子I（PF I）可以恢复有效的聚腺苷酸化活性。我们证明，Yth 1 p是RF I的一个组成部分，在体内和体外与Fip 1 p，一个已知的RF I亚基相互作用。
Cleavage and polyadenylation specificity factor (CPSF), a key component of the mammalian RNA 3'-end processing machinery, consists of four subunits of 160, 100, 73, and 30 kD. Here we report the isolation and characterization of a cDNA encoding the 30-kD polypeptide. Antibodies raised against this protein inhibit cleavage and polyadenylation and coimmunoprecipitate the other CPSF subunits. The protein sequence contains five C3H-zinc-finger repeats and a putative RNA-binding zinc knuckle motif at the carboxyl terminus. Consistent with this observation, the in vitro translated 30-kD protein binds RNA polymers with a distinct preference for poly(U). In addition, an essential S. cerevisiae gene, YTH1, was cloned which is 40% identical to CPSF 30K at the protein level. Extracts prepared from a conditional yth1 mutant have normal cleavage activity, but fail to polyadenylate the upstream cleavage product. Efficient polyadenylation activity can be restored by the addition of purified polyadenylation factor I (PF I). We demonstrate that Yth1p is a component of RF I that interacts in vivo and in vitro with Fip1p, a known RF I subunit.