A weak PPT establishes inefficient splicing of MAT2A intron 8. (A) Diagram showing the 3′ splice site of the MAT2A detained intron. The PPT is in purple, the UGUA element is bold underlined, purines interrupting the PPT are marked with asterisks, the 3′ splice site AG is underlined in bold black, and the branch point A is underlined in bold red (Mercer et al. 2015). (B) Representative northern blots (top) and quantification (bottom) of the purine-to-pyrimidine mutations in the UGUA motif in the DI. The cells were transfected and the next day were cultured in Met-depleted media for the 0–6 h prior to RNA harvesting. The quantification is presented as percent detained intron. (C) Same as panel B except pyrimidine-to-purine mutations in the UGUA were tested. (D) Same as B, except the A at −7 position relative to the 3′ splice site was mutated to U. (E) Same as B except pyrimidine-to-pyrimidine and purine-to-purine mutations were tested. Samples in B and C are referenced to the same WT control data.
