TDP-43 loss induces extensive cryptic polyadenylation in ALS/FTD

Abstract

Nuclear depletion and cytoplasmic aggregation of the RNA binding protein TDP-43 is the hallmark of ALS, occurring in over 97% of cases. A key consequence of TDP-43 nuclear loss is the de-repression of cryptic exons. Whilst TDP-43 regulated cryptic splicing is increasingly well catalogued, cryptic alternative polyadenylation (APA) events, which define the 3' end of last exons, have been largely overlooked, especially when not associated with novel upstream splice junctions. We developed a novel bioinformatic approach to reliably identify distinct APA event types: alternative last exons (ALE), 3'UTR extensions (3'Ext) and intronic polyadenylation (IPA) events. We identified novel neuronal cryptic APA sites induced by TDP-43 loss of function by systematically applying our pipeline to a compendium of publicly available and in house datasets. We find that TDP-43 binding sites and target motifs are enriched at these cryptic events and that TDP-43 can have both repressive and enhancing action on APA. Importantly, all categories of cryptic APA can also be identified in ALS and FTD post mortem brain regions with TDP-43 proteinopathy underlining their potential disease relevance. RNA-seq and Ribo-seq analyses indicate that distinct cryptic APA categories have different downstream effects on transcript and translation. Intriguingly, cryptic 3'Exts occur in multiple transcription factors, such as ELK1, SIX3, and TLX1, and lead to an increase in wild-type protein levels and function. Finally, we show that an increase in RNA stability leading to a higher cytoplasmic localisation underlies these observations. In summary, we demonstrate that TDP-43 nuclear depletion induces a novel category of cryptic RNA processing events and we expand the palette of TDP-43 loss consequences by showing this can also lead to an increase in normal protein translation.

Loss of TDP-43 induces synaptic dysfunction that is rescued by UNC13A splice-switching ASOs

“…TDP-43 cytoplasmic aggregation and nuclear loss of function (LoF) occur in approximately 97% of ALS patients 4 . Loss of nuclear TDP-43 leads to numerous changes in RNA processing, including the appearance of hundreds of novel exon inclusion, exon skipping, and polyadenylation events that are not observed under normal conditions, hence termed “cryptic” 11,23 . Many such events result in the loss of transcripts and encoded proteins.…”

“…Similar to this patient, transcranial magnetic stimulation studies have detected cortical hyperexcitability at an early stage in ALS patients that worsens with disease progression 22 . Moreover, one of the few drugs that extend survival in ALS, riluzole, causes a reduction in cortical hyperexcitability, suggesting this phenotype may be relevant for the survival of affected people 23 . Cortical hyperexcitability has also been shown for individuals with mutations in STX1B and STXBP1 (ref.…”

“…Moreover, one of the few drugs that extend survival in ALS, riluzole, causes a reduction in cortical hyperexcitability, suggesting this phenotype may be relevant for the survival of affected people 23 .…”

Reduced STMN2 and pathogenic TDP-43, two hallmarks of ALS, synergize to accelerate motor decline in mice

“…TDP-43 regulates the splicing and polyadenylation of many transcripts (Arnold et al, 2024;Bryce-Smith et al, 2024;Jeong et al, 2017;J. P. Ling et al, 2015;Ma et al, 2022;Zeng et al, 2024), but this is unfortunately difficult to model in mice because the cryptic exons TDP-43 regulates are not conserved between humans and mice (J. P. Ling et al, 2015).…”

“…TDP-43 is a predominantly nuclear RNA-binding protein that regulates many aspects of RNA processing including splicing and polyadenylation (S. C. Ling et al, 2013). TDP-43 normally functions to maintain healthy levels of correctly processed mRNAs by suppressing both cryptic exon inclusion and use of cryptic polyadenylation sites (Arnold et al, 2024;Bryce-Smith et al, 2024;Jeong et al, 2017;J. P. Ling et al, 2015;Ma et al, 2022;Zeng et al, 2024).…”