Human RGS6 gene structure, complex alternative splicing, and role of N terminus and G protein γ-subunit-like (GGL) domain in subcellular localization of RGS6 splice variants
Human RGS6 gene structure, complex alternative splicing, and role of N terminus and G protein γ-subunit-like (GGL) domain in subcellular localization of RGS6 splice variants
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DOI：
10.1074/jbc.m212687200
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发表时间：
2003-08-08
影响因子：
4.8
通讯作者：
Fisher, RA
中科院分区：
文献类型：
--
作者：
Chatterjee, TK;Liu, ZY;Fisher, RA
RGS蛋白的定义是存在一个半保守的RGS结构域，该结构域赋予这些蛋白对某些Galpha亚基的gtpase激活活性。RGS6是RGS蛋白亚家族的成员，其特征是存在DEP和GGL结构域，后者是gbeta5相互作用结构域。在这里，我们报告了36种不同的人类RGS6转录本的鉴定，这些转录本是由RGS6基因异常复杂的加工产生的，RGS6基因跨越人类14号染色体的630千碱基对的基因组DNA，被19个内含子打断。这些转录本通过使用两个替代转录位点和复杂的替代剪接机制产生，并编码具有长或短n端结构域、完整或不完整GGL结构域、7个不同的c端结构域和一个发现RGS结构域的共同内部结构域的蛋白质。在表达多种RGS6剪接变异体蛋白的COS-7细胞中，研究了RGS6剪接变异体n端和GGL结构域的结构多样性在其与Gbeta5相互作用和亚细胞定位中的作用，以及Gbeta5在RGS6蛋白定位中的作用。具有完整GGL结构域的RGS6剪接变异体与Gbeta5相互作用，与n端结构域的类型无关，而缺乏完整GGL结构域的RGS6剪接变异体与Gbeta5不相互作用。RGS6蛋白变异表现出亚细胞分布模式，从细胞质独占到核/核仁独占，Gbeta5的共表达促进了RGS6蛋白的核定位。分析结果表明，RGS6蛋白的长n端和GGL结构域序列具有胞质保留序列的功能，可防止其核/核仁积累。这些发现为GGL结构域的Gbeta5独立功能和Gbeta5在RGS蛋白定位中的作用提供了第一个证据。该研究揭示了人类RGS6基因加工的异常复杂性，并为RGS6蛋白家族的结构多样性如何参与其在细胞中的定位和可能的功能提供了新的见解。
RGS proteins are defined by the presence of a semi-conserved RGS domain that confers the GTPase-activating activity of these proteins toward certain Galpha subunits. RGS6 is a member of a subfamily of RGS proteins distinguished by the presence of DEP and GGL domains, the latter a Gbeta5-interacting domain. Here we report identification of 36 distinct transcripts of human RGS6 that arise by unusually complex processing of the RGS6 gene, which spans 630 kilobase pairs of genomic DNA in human chromosome 14 and is interrupted by 19 introns. These transcripts arise by use of two alternative transcription sites and complex alternative splicing mechanisms and encode proteins with long or short N-terminal domains, complete or incomplete GGL domains, 7 distinct C-terminal domains and a common internal domain where the RGS domain is found. The role of structural diversity in the N-terminal and GGL domains of RGS6 splice variants in their interaction with Gbeta5 and subcellular localization and of Gbeta5 on RGS6 protein localization was examined in COS-7 cells expressing various RGS6 splice variant proteins. RGS6 splice variants with complete GGL domains interacted with Gbeta5, irrespective of the type of N-terminal domain, while those lacking a complete GGL domain did not. RGS6 protein variants displayed subcellular distribution patterns ranging from an exclusive cytoplasmic to exclusive nuclear/nucleolar localization, and co-expression of Gbeta5 promoted nuclear localization of RGS6 proteins. Analysis of our results show that the long N-terminal and GGL domain sequences of RGS6 proteins function as cytoplasmic retention sequences to prevent their nuclear/nucleolar accumulation. These findings provide the first evidence for Gbeta5-independent functions of the GGL domain and for a role of Gbeta5 in RGS protein localization. This study reveals extraordinary complexity in processing of the human RGS6 gene and provides new insights into how structural diversity in the RGS6 protein family is involved in their localization and likely function(s) in cells.