Translation of the poly(GR) frame in C9ORF72-ALS/FTD is regulated by cis-elements involved in alternative splicing

Ribosome profiling reveals novel regulation of C9ORF72 GGGGCC repeat-containing RNA translation

“…This alteration significantly increased nano activity in the poly(GP) frame but not firefly luciferase activity. These data indicate that the CUG in ribosome footprint 3 is the start site for poly(GA) frame translation, as has been noted by others ( Green et al 2017 ; Sonobe et al 2018 ; Tabet et al 2018 ; Almeida et al 2019 ; Lampasona et al 2021 ). The observation that mutation of this CUG also up-regulates poly(GP) frame nano translation is paradoxical but could be explained if the reading frame was shifted within the G 4 C 2 repeat ( Tabet et al 2018 ).…”

“…We could detect only a very low amount of translation in the poly(GR) frame under the same conditions as the other two reading frames and thus it was not pursued further. However, intronic sequence requirements for poly(GR)-frame translation have been examined ( Lampasona et al 2021 ).…”

“…Studies based on an in vitro translation system primed with C9ORF72 pre-mRNA and reporter constructs indicate that poly(GR) and poly(GP) protein synthesis is promoted by a near-cognate CUG initiation codon in the GA frame followed by ribosome frameshifting on G 4 C 2 RNA ( Tabet et al 2018 ). However, recent work indicates that intronic sequences 5′ to the repeat regulate poly(GR) synthesis ( Lampasona et al 2021 ).…”

Translation of dipeptide repeat proteins inC9ORF72-ALS/FTD through unique and redundant AUG initiation codons

“…In the latter case, it is thought that repeat-associated non-AUG (RAN) translation of poly-GA and poly-GR occurs via non-canonical CUG and AGG codons, respectively 9, 10, 11, 12, 14, 21 . However, this model of RAN translation for poly-GA and poly-GR has been recently challenged, as translation of these DPRs does not depend on the presence of GGGGCC repeats 13, 14, 15, 21 . Nevertheless, our findings merged with those of previous studies suggest that DPR synthesis involves at least two different modes of translation: near-cognate start codon (e.g., CUG, AGG) dependent translation for poly-GA and poly-GR from sense GGGGCC transcripts, as well as conventional AUG dependent translation for poly-PR and poly-PG synthesis from antisense CCCCGG transcripts.…”

“…As previously shown, non-canonical codons (viz., CUG for poly-GA, AGG for poly-GR) initiate DPR synthesis from the sense strand, suggesting an unconventional form of translation, i.e., repeat-associated non-AUG (RAN) translation 6 . However, deletion analysis of cis -regulatory elements upstream of the GGGGCC repeats and ribosome profiling revealed that translation of the poly-GA and poly-GR frames is independent of the presence of G 4 C 2 repeats 13, 14, 15 . Moreover, a recent study reported that a canonical AUG initiation codon (194 nucleotides upstream of the repeat) is used for poly-PG synthesis from the antisense CCCCGG transcript, suggesting conventional translation is involved in the synthesis of at least one DPR.…”