Mis-spliced transcripts generatede novoproteins in TDP-43-related ALS/FTD

Abstract

Functional loss of TDP-43, an RNA-binding protein genetically and pathologically linked to ALS and FTD, leads to inclusion of cryptic exons in hundreds of transcripts during disease. Cryptic exons can promote degradation of affected transcripts, deleteriously altering cellular function through loss-of-function mechanisms. However, the possibility of de novo protein synthesis from cryptic exon transcripts has not been explored. Here, we show that mRNA transcripts harboring cryptic exons generate de novo proteins both in TDP-43 deficient cellular models and in disease. Using coordinated transcriptomic and proteomic studies of TDP-43 depleted iPSC-derived neurons, we identified numerous peptides that mapped to cryptic exons. Cryptic exons identified in iPSC models were highly predictive of cryptic exons expressed in brains of patients with TDP-43 proteinopathy, including cryptic transcripts that generated de novo proteins. We discovered that inclusion of cryptic peptide sequences in proteins altered their interactions with other proteins, thereby likely altering their function. Finally, we showed that these de novo peptides were present in CSF from patients with ALS. The demonstration of cryptic exon translation suggests new mechanisms for ALS pathophysiology downstream of TDP-43 dysfunction and may provide a strategy for novel biomarker development.

The era of cryptic exons: implications for ALS-FTD

“…Excitingly, two new pre-print publications this year have independently reported the detection of cryptic peptides in the CSF of patients with ALS [57,58]. Irwin et al demonstrated that a newly characterised monoclonal antibody, specific to a TDP-43-dependent cryptic epitope encoded by the cryptic exon found in HDGFL2, detects the cryptic peptide in C9orf72-associated ALS.…”

“…Strikingly, this includes pre-symptomatic mutation carriers [57]. In Seddighi et al, our groups first demonstrated the presence of de novo cryptic peptides in iPSC-derived neurons with TDP-43 knockdown, and then used a novel targeted proteomics assay to confirm the presence of cryptic peptides in CSF of patients with ALS-FTD [58]. Further work will be needed to assess the specificity and sensitivity of cryptic peptides as biomarkers, but, taken together, these discoveries are encouraging steps towards facilitating earlier diagnosis of ALS, and also providing a way of measuring target engagement in clinical trials for new therapies aimed at restoring TDP-43 function.…”

A panel of TDP-43-regulated splicing events verify loss of TDP-43 function in amyotrophic lateral sclerosis brain tissue

“…Diagnostic approaches include testing DEU changes or cryptic exon emergence with PCR or RNA-sequencing of biofluids (57, 88). Another emerging approach is designing aptamers or antibodies to cryptic peptides or neoepitopes that result from DEU events (89, 90). Indeed, the isoform switch of POLDIP3 from variant-1 to variant-2 due to exon skipping in POLDIP3 has also been proposed to be a measurable protein biomarker due to variant-2 being rarely detected in normal tissues (55).…”

LINE-associated cryptic splicing induces dsRNA-mediated interferon response and tumor immunity

“…Additionally, cryptic exon-containing transcripts can potentially be translated into novel proteins, which may affect cell fitness and produce neo-antigens in tumors. [64][65][66] These findings shine light on the role of hnRNPM not only as a repressor of alternatively spliced and cryptic exon inclusion, but also a guardian of transcriptome stability and proteome integrity.…”