Green KM, Glineburg MR, Kearse MG, Flores BN, Linsalata AE, Fedak SJ, Goldstrohm AC, Barmada SJ, Todd PK. RAN translation at C9orf72-associated repeat expansions is selectively enhanced by the integrated stress response. Nat Commun. 2017 Dec 8;8(1):2005. PubMed.
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I consider all three papers to be significant contributions to our understanding of the role of DPR translation in ALS. The stimulation of DPR translation by the integrated stress response is an exciting finding, and the induction of a feed-forward response in which both the stress response and DPR translation induce each other may prove to be an important step in the pathogenesis of C9ORF72-mediated disease.
While the primary discrepancy between the publications is whether DPR translation requires a 5' cap, all three publications show that cap-dependent translation is more efficient than cap-independent translation of the DPRs. It would be very interesting to see if the 5' cap could be visualized on the G4C2 transcript from IPS neurons derived from C9 patients. The three publications agree that the GA dipeptide is more efficiently translated than either the GP or GR dipeptides. Additionally, it would be interesting to know the relative translation rates of the antisense DPRs in relation to the sense DPRs and whether the antisense DPRs are also stimulated by the integrated stress response.
View all comments by Brian FreibaumNeurobiology, KU Leuven & VIB
Very interesting that three research groups investigated the intriguing mechanism behind non-ATG-mediated translation of the hexanucleotide repeats in C9ORF72, a process also known as RAN translation. We find the observation by Cheng et al. and Green et al. that stress induces a feed-forward loop to enhance RAN translation extremely interesting and it fits in a broader perspective. These insights suggest a self-sustaining mechanism by which stress, induced by arginine-containing DPRs (PR and GR), which can phase-separate, or by the repeat RNA itself (Swinnen et al., 2018), fuels a vicious circle. This stress-induced process is mediated by phosphorylation of the alpha subunit of eukaryotic initiation factor-2 (eIF2). Accordingly, the formation of DPR-induced stress granules is dependent on eIF2α phosphorylation (Boeynaems et al., 2017). The immediate consequence of stress granule formation in general is that canonical cap-dependent translation is downregulated and that non-canonical cap-independent translation (e.g., making use of internal ribosomal entry sites, or IRESes, and non-AUG start sites) is increased in order to maintain proteostasis. Despite some major discrepancies, the current papers have unraveled important aspects of this process in the context of C9ORF72. Most consistently, the C9ORF72 repeat expansion undergoes cap-independent RAN translation, however with a 70–90 percent lower efficiency compared to cap-dependent RAN translation. Also, a near-cognate CUG codon upstream of the repeat, which is in frame with poly-GA, seems to be important for RAN translation.
A number of questions remain. It still needs to be investigated how antisense repeats are translated into DPRs (PA and PR), although recent pathological data indicate that these are less abundant and that especially GR, translated from the sense strand, correlates with neurodegeneration and co-localizes with TDP-43 (Saberi et al., 2017). In addition, the exact nature of the RNA species present in patients who will evince RAN translation is still unknown. In relation to this, it is intriguing that the RAN translation process is mostly cap-dependent according to Green et al. and Tabet et al. and proposed to be cap-independent, although significantly enhanced by capping, in Cheng et al. Cap-independent translation would not only be expected based on the fact that the hexanucleotide repeats are spliced out of an intron, it would also fit better with the hypothesis that translation of non-capped RNA is still ongoing under stress conditions, while cap-dependent translation is silenced. Future research is needed to clarify this issue. Also, as only poly-GA is in frame with the near-cognate CUG codon, it is not clear how the other sense-derived DPRs are produced. One possibility is frameshifts during translation that subsequently generate poly-GP and poly-GR, which might explain why GA is the most abundant DPR in postmortem tissue. However, more complicated mechanisms could be at play as removing the CUG in the GA frame has variable effects on poly-GP and poly-GR production, depending on the cellular context.
It is clear that if a “gain-of-function” mechanism is responsible for initiating the disease, prevention of the formation of repeat-containing RNAs is the way to go. This strategy not only prevents the formation of potentially toxic RNAs, but also the translation of the different DPRs.
Ludo van den Bosch
References:
Boeynaems S, Bogaert E, Kovacs D, Konijnenberg A, Timmerman E, Volkov A, Guharoy M, De Decker M, Jaspers T, Ryan VH, Janke AM, Baatsen P, Vercruysse T, Kolaitis RM, Daelemans D, Taylor JP, Kedersha N, Anderson P, Impens F, Sobott F, Schymkowitz J, Rousseau F, Fawzi NL, Robberecht W, Van Damme P, Tompa P, Van Den Bosch L. Phase Separation of C9orf72 Dipeptide Repeats Perturbs Stress Granule Dynamics. Mol Cell. 2017 Mar 16;65(6):1044-1055.e5. PubMed.
Saberi S, Stauffer JE, Jiang J, Garcia SD, Taylor AE, Schulte D, Ohkubo T, Schloffman CL, Maldonado M, Baughn M, Rodriguez MJ, Pizzo D, Cleveland D, Ravits J. Sense-encoded poly-GR dipeptide repeat proteins correlate to neurodegeneration and uniquely co-localize with TDP-43 in dendrites of repeat-expanded C9orf72 amyotrophic lateral sclerosis. Acta Neuropathol. 2018 Mar;135(3):459-474. Epub 2017 Dec 1 PubMed.
Swinnen B, Bento-Abreu A, Gendron TF, Boeynaems S, Bogaert E, Nuyts R, Timmers M, Scheveneels W, Hersmus N, Wang J, Mizielinska S, Isaacs AM, Petrucelli L, Lemmens R, Van Damme P, Van Den Bosch L, Robberecht W. A zebrafish model for C9orf72 ALS reveals RNA toxicity as a pathogenic mechanism. Acta Neuropathol. 2018 Mar;135(3):427-443. Epub 2018 Jan 4 PubMed.
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