Drosophila Pur-α binds to trinucleotide-repeat containing cellular RNAs and translocates to the early oocyte
Drosophila Pur-α binds to trinucleotide-repeat containing cellular RNAs and translocates to the early oocyte
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DOI：
10.4161/rna.19760
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发表时间：
2012-05-01
期刊：
影响因子：
4.1
通讯作者：
Foerstemann, Klaus
中科院分区：
文献类型：
--
作者：
Aumiller, Verena;Graebsch, Almut;Foerstemann, Klaus
Pur-alpha被鉴定为对单链PUR基序（GGN）具有高亲和力的DNA结合蛋白（n）。与DNA结合，Pur-alpha可以激活和抑制转录。此外，Pur-alpha与RNA结合，可能参与核RNA输出以及细胞质神经元mRNP颗粒的转运。遗传性三核苷酸重复扩增疾病脆性X相关性震颤和共济失调综合征（FXTAS）导致Pur-alpha与突变的异常长r（CGG）（n）延伸的相互作用，这似乎将蛋白质从其生理mRNA靶标滴定到核RNA-蛋白质聚集体中。我们研究了果蝇Pur-alpha的功能，并证明该蛋白质在卵子发生早期在生长的卵母细胞中积累。共纯化蛋白质揭示，Pur-alpha是转运mRNP复合物的一部分，类似于其在神经细胞中的作用。我们分析了突变GFP-Pur-α融合蛋白的亚细胞定位，其中核酸结合或二聚化或两者都被阻止。我们建议，与mRNA的协会发生在细胞核中，是所需的复杂的核出口。此外，有效地易位到卵母细胞中还需要RNA结合以及二聚化。RNA结合试验表明，重组果蝇Pur-alpha可以以比以前认为的更高的亲和力结合r（CGG）（4）。相关序列，如r（CAG）（4）和opa重复序列r（CAG）（3）CAA的共有序列，也可以在体外和体内与Pur-alpha结合。因此，Pur-alpha的mRNA靶谱可能比之前预期的要大。
Pur-alpha was identified as a DNA-binding protein with high affinity for the single-stranded PUR-motif (GGN)(n). Bound to DNA, Pur-alpha can both activate and repress transcription. In addition, Pur-alpha binds to RNA and may participate in nuclear RNA export as well as transport of cytoplasmic neuronal mRNP granules. The heritable trinucleotide-repeat expansion disease fragile X associated tremor and ataxia syndrome (FXTAS) leads to interaction of Pur-alpha with mutant, abnormally long r(CGG)(n) stretches, which appears to titrate the protein away from its physiologic mRNA targets into nuclear RNA-protein aggregates. We examined the function of Drosophila Pur-alpha and demonstrate that the protein accumulates in the growing oocyte early in oogenesis. Co-purifying proteins reveal that Pur-alpha is part of transported mRNP complexes, analogous to its reported role in nerve cells. We analyzed the subcellular localization of mutant GFP-Pur-alpha fusion proteins where either nucleic acid binding or dimerization, or both, were prevented. We propose that association with mRNAs occurs in the nucleus and is required for nuclear export of the complex. Furthermore, efficient translocation into the oocyte also requires RNA binding as well as dimerization. RNA binding assays demonstrate that recombinant Drosophila Pur-alpha can bind r(CGG)(4) with higher affinity than previously thought. Related sequences, such as r(CAG)(4) and the consensus sequence of the opa-repeat r(CAG)(3) CAA, can also associate with Pur-alpha in vitro and in vivo. The mRNA target spectrum of Pur-alpha may therefore be larger than previously anticipated.