Interdependent nucleocytoplasmic trafficking and interactions of Dis3 with Rrp6, the core exosome and importin-alpha3.
Interdependent nucleocytoplasmic trafficking and interactions of Dis3 with Rrp6, the core exosome and importin-alpha3.
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DOI：
10.1111/j.1600-0854.2009.00888.x
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发表时间：
2009-05
期刊：
影响因子：
--
通讯作者：
Andrulis ED
中科院分区：
文献类型：
--
作者：
Graham AC;Davis SM;Andrulis ED
核糖核酸外切酶（RNases）的亚细胞区室化是RNA加工和降解的时空调控的重要机制。尽管在理解RNA酶底物和功能方面取得了很大进展，但我们对RNA酶如何转运和组装成功能性的亚细胞限制性复合物知之甚少。为了深入了解这个问题，我们正在研究外泌体结合蛋白Dis 3，一种进行性3'到5'核糖核酸外切酶。在这里，我们研究了果蝇Dis 3（dDis 3）蛋白的相互作用和亚细胞定位。dDis 3的N-末端结构域突变体消除了与“核心”外泌体的关联，但仅减少了与“核”外泌体相关因子dRrp 6的结合。我们发现，核定位dDis 3需要一个C-末端经典的核定位信号（NLS）。与此一致，dDis 3特异性地共沉淀NLS结合蛋白importin-α3。令人惊讶的是，缺乏或突变C-末端NLS的dDis 3构建体保留了输入蛋白-α3结合，表明相互作用是间接的。最后，我们发现内源性dDis 3和dRrp 6表现出协调的核富集或排斥，表明dDis 3，Rrp 6和importin-α3在独立于核心的复合物中相互作用。我们认为，这种复合物的运动和沉积对于外泌体核心的亚细胞区室化和调节是重要的。
Subcellular compartmentalization of exoribonucleases (RNases) is an important control mechanism in the temporal and spatial regulation of RNA processing and decay. Despite much progress towards understanding RNase substrates and functions, we know little of how RNases are transported and assembled into functional, subcellularly-restricted complexes. To gain insight into this issue, we are studying the exosome-binding protein Dis3, a processive 3' to 5' exoribonuclease. Here, we examine the interactions and subcellular localization of the Drosophila melanogaster Dis3 (dDis3) protein. N-terminal domain mutants of dDis3 abolish associations with the `core' exosome yet only reduce binding to the `nuclear' exosome-associated factor dRrp6. We show that nuclear localization of dDis3 requires a C-terminal classic nuclear localization signal (NLS). Consistent with this, dDis3 specifically co-precipitates the NLS-binding protein importin-α3. Surprisingly, dDis3 constructs that lack or mutate the C-terminal NLS retain importin-α3 binding, suggesting that the interaction is indirect. Finally, we find that endogenous dDis3 and dRrp6 exhibit coordinated nuclear enrichment or exclusion, suggesting that dDis3, Rrp6, and importin-α3 interact in a complex independent of the core. We propose that the movement and deposition of this complex is important for the subcellular compartmentalization and regulation of the exosome core.
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