SPT5 affects the rate of mRNA degradation and physically interacts with CCR4 but does not control mRNA deadenylation.
SPT5 affects the rate of mRNA degradation and physically interacts with CCR4 but does not control mRNA deadenylation.
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DOI：
10.4236/ajmb.2012.21002
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发表时间：
2012-01-01
期刊：
影响因子：
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通讯作者：
Denis, Clyde L
中科院分区：
文献类型：
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作者：
Cui, Yajun;Chiang, Yueh-Chin;Denis, Clyde L
CCR4- not复合物已被证明在mRNA代谢中具有多种作用，包括转录延伸、mRNA运输和核外泌体功能，但CCR4和CAF1的主要功能是细胞质mRNA的死烯化和降解。由于先前的遗传分析支持SPT5（已知参与转录延伸）与CCR4之间的相互作用，因此研究了SPT5与CCR4的物理关联。利用CCR4的deadenylase结构域作为诱饵的双杂交筛选发现SPT5是一个潜在的相互作用蛋白。生理浓度的SPT5可以免疫沉淀CCR4和CAF1，体外纯化的SPT5可以特异性结合CAF1和CCR4的死烯酶结构域。我们还证明，SPT5突变或spt4缺失减慢了mRNA降解的速度，这是一种与CCR4 mRNA死烯化酶复合物缺陷相关的表型。然而，与ccr4和caf1缺失不同，spt5和spt4缺陷对死基化率的影响很小。它们也不影响mRNA的脱帽或5‘ - 3’降解。这些结果表明，SPT5/SPT4与CCR4- not复合物之间的相互作用可能是涉及核事件的影响的结果，而不涉及CCR4在mRNA死基化和转换中的主要作用。
The CCR4-NOT complex has been shown to have multiple roles in mRNA metabolism, including that of transcriptional elongation, mRNA transport, and nuclear exosome function, but the primary function of CCR4 and CAF1 is in the deadenylation and degradation of cytoplasmic mRNA. As previous genetic analysis supported an interaction between SPT5, known to be involved in transcriptional elongation, and that of CCR4, the physical association of SPT5 with CCR4 was examined. A two-hybrid screen utilizing the deadenylase domain of CCR4 as a bait identified SPT5 as a potential interacting protein. SPT5 at its physiological concentration was shown to immunoprecipitate CCR4 and CAF1, and in vitro purified SPT5 specifically could bind to CAF1 and the deadenylase domain of CCR4. We additionally demonstrated that mutations in SPT5 or an spt4 deletion slowed the rate of mRNA degradation, a phenotype associated with defects in the CCR4 mRNA deadenylase complex. Yet, unlike ccr4 and caf1 deletions, spt5 and spt4 defects displayed little effect on the rate of deadenylation. They also did not affect decapping or 5' - 3' degradation of mRNA. These results suggest that the interactions between SPT5/SPT4 and the CCR4-NOT complex are probably the consequences of effects involving nuclear events and do not involve the primary role of CCR4 in mRNA deadenylation and turnover.