Ribosomal composition affects the noncanonical translation and toxicity of polyglycine-containing proteins in fragile X-associated conditions

Expansion of CGG repeats (CGGexp) in the 5’ untranslated region (5’UTR) of theFMR1gene underlies the fragile X-associated conditions including tremor/ataxia syndrome (FXTAS), a late-onset neurodegenerative disease. One pathomechanism of FXTAS is the repeat-associated non-AUG-initiated (RAN) translation of CGG repeats of mutantFMR1mRNA, resulting in production of FMRpolyG, a toxic protein containing long polyglycine tract. To identify novel modifiers of RAN translation we used an RNA-tagging system and mass spectrometry-based screening. It revealed proteins enriched on CGGexp-containingFMR1RNA in cellulo, including a ribosomal protein RPS26, a component of the 40S subunit. We demonstrated that RPS26, together with its chaperone TSR2, modulates FMRpolyG production and its toxicity. We also found that the number of proteins produced via RPS26-sensitive translation was limited, and 5’UTRs of mRNAs encoding these proteins were guanosine and cytosine-rich. Moreover, the silencing of another component of the 40S subunit, the ribosomal protein RPS25, also induced repression of FMRpolyG biosynthesis. Results of this study suggest that the composition of the 40S subunit plays important role in noncanonical CGGexp-related RAN translation.