Alcázar-Román Abel R, Tran Elizabeth J, Guo Shuangli, Wente Susan R
Department of Cell and Developmental Biology, Vanderbilt University Medical Center, U-3209 MRBIII, 465 21st Avenue South, Nashville, TN 37232-8240, USA.
Nat Cell Biol. 2006 Jul;8(7):711-6. doi: 10.1038/ncb1427. Epub 2006 Jun 18.
Regulation of nuclear mRNA export is critical for proper eukaryotic gene expression. A key step in this process is the directional translocation of mRNA-ribonucleoprotein particles (mRNPs) through nuclear pore complexes (NPCs) that are embedded in the nuclear envelope. Our previous studies in Saccharomyces cerevisiae defined an in vivo role for inositol hexakisphosphate (InsP6) and NPC-associated Gle1 in mRNA export. Here, we show that Gle1 and InsP6 act together to stimulate the RNA-dependent ATPase activity of the essential DEAD-box protein Dbp5. Overexpression of DBP5 specifically suppressed mRNA export and growth defects of an ipk1 nup42 mutant defective in InsP6 production and Gle1 localization. In vitro kinetic analysis showed that InsP6 significantly increased Dbp5 ATPase activity in a Gle1-dependent manner and lowered the effective RNA concentration for half-maximal ATPase activity. Gle1 alone had minimal effects. Maximal InsP6 binding required both Dbp5 and Gle1. It has been suggested that Dbp5 requires unidentified cofactors. We now propose that Dbp5 activation at NPCs requires Gle1 and InsP6. This would facilitate spatial control of the remodelling of mRNP protein composition during directional transport and provide energy to power transport cycles.
核mRNA输出的调控对于真核基因的正常表达至关重要。这一过程中的关键步骤是mRNA-核糖核蛋白颗粒（mRNP）通过嵌入核膜的核孔复合体（NPC）进行定向转运。我们之前在酿酒酵母中的研究确定了肌醇六磷酸（InsP6）和与NPC相关的Gle1在mRNA输出中的体内作用。在此，我们表明Gle1和InsP6共同作用以刺激必需的DEAD-box蛋白Dbp5的RNA依赖性ATP酶活性。DBP5的过表达特异性地抑制了在InsP6产生和Gle1定位方面存在缺陷的ipk1 nup42突变体的mRNA输出和生长缺陷。体外动力学分析表明，InsP6以Gle1依赖性方式显著增加Dbp5的ATP酶活性，并降低了达到最大ATP酶活性一半时的有效RNA浓度。单独的Gle1作用极小。InsP6的最大结合需要Dbp5和Gle1两者。有人提出Dbp5需要未确定的辅因子。我们现在提出，在NPC处Dbp5的激活需要Gle1和InsP6。这将有助于在定向转运过程中对mRNP蛋白质组成重塑进行空间控制，并为转运循环提供能量。
