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Alternative splicing involving the 3' tandem splice site NAGNAG sequence may play a role in the structure-function diversity of proteins. However, how 3' tandem splice site utilization is determined is not well understood. We previously demonstrated that 3' NAGNAG-based wobble splicing occurs mostly in a tissue- and developmental stage-independent manner. Bioinformatic analysis reveals that the nucleotide preceding the AG dinucleotide may influence 3' splice site utilization; this is also supported by an in vivo splicing assay. Moreover, we found that the intron sequence plays an important role in 3' splice site selection for NAGNAG wobble splicing. Mutations of the region between the branch site and the NAGNAG 3' splice site, indeed, affected the ratio of the distal/proximal AG selection. Finally, we found that single nucleotide polymorphisms around the NAGNAG motif could affect the splice site choice, which may lead to a change in mRNA patterns and influence protein function. We conclude that the NAGNAG motif and its upstream region to the branch point sequence are required for 3' tandem splice site selection.

Wobble Splicing Reveals the Role of the Branch Point Sequence-to-NAGNAG Region in 3′ Tandem Splice Site Selection