Bidirectional Transcription at the PPP2R2B Gene Locus in Spinocerebellar Ataxia Type 12

Abstract Background Spinocerebellar ataxia type 12 (SCA12) is a neurodegenerative disease caused by expansion of a CAG repeat in the PPP2R2B gene. Objective In this study, we tested the hypothesis that the PPP2R2B antisense ( PPP2R2B‐AS1 ) transcript containing a CUG repeat is expressed and contributes to SCA12 pathogenesis. Methods Expression of PPP2R2B‐AS1 transcript was detected in SCA12 human induced pluripotent stem cells (iPSCs), iPSC‐derived NGN2 neurons, and SCA12 knock‐in mouse brains using strand‐specific reverse transcription polymerase chain reaction. The tendency of expanded PPP2R2B‐AS1 ( expPPP2R2B‐AS1 ) RNA to form foci, a marker of toxic processes involving mutant RNAs, was examined in SCA12 cell models by fluorescence in situ hybridization. The apoptotic effect of expPPP2R2B‐AS1 transcripts on SK‐N‐MC neuroblastoma cells was evaluated by caspase 3/7 activity. Western blot was used to examine the expression of repeat associated non‐ATG‐initiated translation of expPPP2R2B‐AS1 transcript in SK‐N‐MC cells. Results The repeat region in the PPP2R2B gene locus is bidirectionally transcribed in SCA12 iPSCs, iPSC‐derived NGN2 neurons, and SCA12 mouse brains. Transfected expPPP2R2B‐AS1 transcripts induce apoptosis in SK‐N‐MC cells, and the apoptotic effect may be mediated, at least in part, by the RNA secondary structure. The expPPP2R2B‐AS1 transcripts form CUG RNA foci in SK‐N‐MC cells. expPPP2R2B‐AS1 transcript is translated in the alanine open reading frame (ORF) via repeat‐associated non‐ATG translation, which is diminished by single‐nucleotide interruptions within the CUG repeat and MBNL1 overexpression. Conclusions These findings suggest that PPP2R2B‐AS1 contributes to SCA12 pathogenesis and may therefore provide a novel therapeutic target for the disease. © 2023 International Parkinson and Movement Disorder Society.