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

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.