RNA Gain-of-Function in Spinocerebellar Ataxia Type 8

Abstract

Microsatellite expansions cause a number of dominantly-inherited neurological diseases. Expansions in coding-regions cause protein gain-of-function effects, while non-coding expansions produce toxic RNAs that alter RNA splicing activities of MBNL and CELF proteins. Bi-directional expression of the spinocerebellar ataxia type 8 (SCA8) CTG CAG expansion produces CUG expansion RNAs (CUGexp) from the ATXN8OS gene and a nearly pure polyglutamine expansion protein encoded by ATXN8 CAGexp transcripts expressed in the opposite direction. Here, we present three lines of evidence that RNA gain-of-function plays a significant role in SCA8: 1) CUGexp transcripts accumulate as ribonuclear inclusions that co-localize with MBNL1 in selected neurons in the brain; 2) loss of Mbnl1 enhances motor deficits in SCA8 mice; 3) SCA8 CUGexp transcripts trigger splicing changes and increased expression of the CUGBP1-MBNL1 regulated CNS target, GABA-A transporter 4 (GAT4/Gabt4). In vivo optical imaging studies in SCA8 mice confirm that Gabt4 upregulation is associated with the predicted loss of GABAergic inhibition within the granular cell layer. These data demonstrate that CUGexp transcripts dysregulate MBNL/CELF regulated pathways in the brain and provide mechanistic insight into the CNS effects of other CUGexp disorders. Moreover, our demonstration that relatively short CUGexp transcripts cause RNA gain-of-function effects and the growing number of antisense transcripts recently reported in mammalian genomes suggest unrecognized toxic RNAs contribute to the pathophysiology of polyglutamine CAG CTG disorders.

CAG repeat RNA as an auxiliary toxic agent in polyglutamine disorders

“…13,14,[26][27][28] In DM1, SCA8 and DM2, altered alternative splicing is partially a consequence of MBNL family protein sequestration by nuclear repeat RNA foci. In FXTAS, wherein mutant CGG repeat RNA titrates, among other proteins also MBNL1, only Sam68 sequestration, which is a strong and early event, interferes with splicing regulation of its pre-mRNA targets.…”

“…The RNA gain-of-function mechanism initially described in DM1, 18,20,35 wherein repeat expansions are found to be toxic only at the RNA level, now also explains some aspects of pathogenesis in other non-coding expansion disorders, including FXTAS, SCA8, SCA31, HDL2 as well as SCA10 and DM2. 11,[13][14][15][16][17] SCA8 is a unique among these diseases because…”

“…DM1, which is caused by a 3'UTR CTG repeat expansion, is the best characterized example of a disorder in which mutant RNA is thought to induce toxicity. The involvement of other repeat-harboring transcripts in neurodegeneration via an RNA gain-of-function mechanism has been described not only for trinucleotide repeat disorders, such as FXTAS, SCA8 and HDL2, [13][14][15] but also for DM2, SCA10 and SCA31. 9,11,16,17 In the RNA gain-of-function mechanism, the expanded repeat of a flawed RNA exerts its toxic function by sequestration and subsequent loss-of-function of RNA binding proteins, such as the muscleblind-like (MBNL) family.…”

“…Resultant ribonucleoprotein complexes become trapped in the nucleus where they form microscopic bodies and become toxic. Punctate intranuclear foci that label with antibodies against muscleblind 1 (MBNL1) protein are a characteristic feature of cells expressing non-coding expansions of CUG, 14,[18][19][20] CGG or CCUG repeats. 13,16 More recently, transcripts harboring translated CAG repeat expansions have been observed to form intranuclear MBNL1-positive RNA foci in human HD fibroblasts.…”

CLIP: viewing the RNA world from an RNA-protein interactome perspective

“…Apart from Ago proteins, splicing regulators are the most intensively studied proteins by CLIP. RNA binding patterns of Nova [6,8], PTBs [85,86], FOX2 [87], SFRS1 [88], TIA [89], MBNL1 [90], Cugbp1 [91], SRSF1/2 [92] and hnRNP proteins [18,93,94] have been investigated.…”