Role of pICLn in methylation of Sm proteins by PRMT5.
Role of pICLn in methylation of Sm proteins by PRMT5.
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DOI：
10.1074/jbc.m109.015578
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发表时间：
2009-08-07
期刊：
影响因子：
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通讯作者：
Van Duyne GD
中科院分区：
文献类型：
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作者：
Pesiridis GS;Diamond E;Van Duyne GD
pICln是一种必需的、高度保守的26-kDa蛋白，其功能包括与人细胞的细胞质中的Sm蛋白结合，并通过存活运动神经元复合体介导细胞的RNA剪接机制的有序和受调控的组装。pICln还与PRMT 5相互作用，PRMT 5是负责在三种典型Sm蛋白的羧基末端区域上产生对称二甲基精氨酸修饰的酶。为了更好地理解pICln在这些细胞过程中的作用，我们研究了pICln和pICln·Sm复合物的性质以及pICln对PRMT 5甲基转移酶活性的影响。我们发现pICln在溶液中是一个单体，与SmD_3-SmB有很高的亲和力（Kd值约为160 nm），并与Sm蛋白和Sm蛋白亚复合物形成1：1的复合物。这些数据支持的pICln结合的封端模型，支持目前的观点，如何防止非法RNA底物Sm寡聚化pICln。我们已经发现，通过与pICln共表达，重组PRMT 5可以以可溶的活性形式产生。PRMT 5单独对多种已知底物具有混杂活性。然而，在pICln的存在下，PRMT 5甲基化的Sm蛋白被刺激，但组蛋白的甲基化被抑制。我们还发现，在pICln的突变，不影响Sm蛋白结合仍然可以有一个深刻的影响甲基转移酶的PRMT 5复合物的活性。总之，这些数据提供了对pICln功能的深入了解，并代表了PRMT 5生化分析的重要起点。
pICln is an essential, highly conserved 26-kDa protein whose functions include binding to Sm proteins in the cytoplasm of human cells and mediating the ordered and regulated assembly of the cell's RNA-splicing machinery by the survival motor neurons complex. pICln also interacts with PRMT5, the enzyme responsible for generating symmetric dimethylarginine modifications on the carboxyl-terminal regions of three of the canonical Sm proteins. To better understand the role of pICln in these cellular processes, we have investigated the properties of pICln and pICln·Sm complexes and the effects that pICln has on the methyltransferase activity of PRMT5. We find that pICln is a monomer in solution, binds with high affinity (Kd ∼ 160 nm) to SmD3-SmB, and forms 1:1 complexes with Sm proteins and Sm protein subcomplexes. The data support an end-capping model of pICln binding that supports current views of how pICln prevents Sm oligomerization on illicit RNA substrates. We have found that by co-expression with pICln, recombinant PRMT5 can be produced in a soluble, active form. PRMT5 alone has promiscuous activity toward a variety of known substrates. In the presence of pICln, however, PRMT5 methylation of Sm proteins is stimulated, but methylation of histones is inhibited. We have also found that mutations in pICln that do not affect Sm protein binding can still have a profound effect on the methyltransferase activity of the PRMT5 complex. Together, the data provide insights into pICln function and represent an important starting point for biochemical analyses of PRMT5.