Identification of a nuclear localization signal mediating the nuclear import of Arabidopsis splicing factor1

Abstract

The splicing factor1 protein (SF1) is involved in branch point recognition of pre-mRNA introns during the early stages of spliceosome assembly in the nucleus. In this study, we aimed to characterize the nuclear localization signal (NLS) of the Arabidopsis SF1 protein (AtSF1). There are two putative NLS sequences (RRKRRSR and RKRKSR) at the N-terminal side of the AtSF1 protein. Analysis of green fluorescence protein (GFP)-tagged AtSF1 deletion constructs indicated that the RKRKSRWADDE sequence (from the 124th to 134th amino acid residues) is necessary for GFP-tagged AtSF1 protein for the localization in the nucleus. Further analysis of the RKRKSRWADDE sequence using site-directed mutagenesis demonstrated that at least two basic amino acid residues (R and K) within the sequence is essential for the complete nuclear localization of GFP-tagged AtSF1 protein. Taken together, our findings demonstrated that only one of the two predicted NLS candidates of the AtSF1 protein is necessary for its nuclear localization, and at least two basic amino acid residues within the motif are crucial for its function. This feature of NLS may be unique in plant SF1 proteins because there is only one predicted NLS in fungal and metazoan counterparts.