RNA-protein interactions promote asymmetric sorting of the ASH1 mRNA ribonucleoprotein complex
RNA-protein interactions promote asymmetric sorting of the ASH1 mRNA ribonucleoprotein complex
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作者：
Gonsalvez, GB;Lehmann, KA;Long, RM
在酿酒酵母中，ASH 1 mRNA在细胞周期后期定位于子细胞的顶端。ASH 1 mRNA定位依赖于四个顺式作用定位元件以及Myo 4p、She 2 p和She 3 p。假设Myo 4p、She 2 p和She 3 p形成异源三聚体蛋白复合物，其直接将ASH 1 mRNA转运至子细胞。She 2 p是一种RNA结合蛋白，直接与ASH 1顺式作用定位元件相互作用，并与She 3 p结合。在这里，我们报告了七个She 2 p突变体的鉴定-N36 S，R43 A，R44 A，R52 A，R52 K，R63 A和R63 K-导致ASH 1 mRNA的离域。这些突变体的RNA结合活性是有缺陷的，但保留了与She 3 p相互作用的能力，表明功能性的She 2 p RNA结合结构域不是与She 3 p结合的先决条件。此外，N36 S和R63 K She 2 p突变体的核/胞质分布没有改变，表明She 2 p的核/胞质运输不依赖于RNA结合活性。使用N36 S和R63 K She 2 p突变体，我们观察到在不存在She 2 p RNA结合活性的情况下，Myo 4p和She 3 p都不对称地分选到子细胞。然而，在不存在She 2 p的情况下，Myo 4p和She 3 p可以通过人工将mRNA拴系到She 3 p而不对称地分离到子细胞，这意味着Myo 4p/She 3 p复合物的转运和/或锚定依赖于相关mRNA的存在。
In Saccharomyces cerevisiae, ASH1 mRNA is localized to the tip of daughter cells during anaphase of the cell cycle. ASH1 mRNA localization is dependent on four cis-acting localization elements as well as Myo4p, She2p, and She3p. Myo4p, She2p, and She3p are hypothesized to form a heterotrimeric protein complex that directly transports ASH1 mRNA to daughter cells. She2p is an RNA-binding protein that directly interacts with ASH1 cis-acting localization elements and associates with She3p. Here we report the identification of seven She2p mutants-N36S, R43A, R44A, R52A, R52K, R63A, and R63K-that result in the delocalization of ASH1 mRNA. These mutants are defective for RNA-binding activity but retain the ability to interact with She3p, indicating that a functional She2p RNA-binding domain is not a prerequisite for association with She3p. Furthermore, the nuclear/cytoplasmic distribution for the N36S and R63K She2p mutants is not altered, indicating that nuclear/cytoplasmic trafficking of She2p is independent of RNA-binding activity. Using the N36S and R63K She2p mutants, we observed that in the absence of She2p RNA-binding activity, neither Myo4p nor She3p is asymmetrically sorted to daughter cells. However, in the absence of She2p, Myo4p and She3p can be asymmetrically segregated to daughter cells by artificially tethering mRNA to She3p, implying that the transport and/or anchoring of the Myo4p/She3p complex is dependent on the presence of associated mRNA.