TranscriptAccuracyAD · Leveraging long-read RNA-sequencing to understand the pathophysiology of familial and complex Alzheimer’s disease

Mutations in Presenilin 1 (PSEN1) represent the most common cause of autosomal dominant familial Alzheimer’s disease (fAD), resulting in a severe early-onset phenotype. Two pathogenic PSEN1 variants, the intron 4 deletion (int4del) and the missense mutation R278I, have been associated with fAD. However, their transcriptional consequences remain remarkably understudied.I hypothesise that int4del and R278I (i) perturb PSEN1 splicing, thereby compromising diverse PSEN1 functions in neuronal pathways that are central to Alzheimer’s pathogenesis, with the potential to exacerbate pathology; and (ii) can reveal molecular mechanisms shared with sporadic forms of the disease, paving the way for novel therapeutic strategies.To test this hypothesis, I propose analysing a novel long-read RNA-sequencing dataset formed by 12 induced pluripotent stem cell (iPSC)-derived samples from two fAD patients. These include triplicate neuronal lines harbouring the int4del and R278I mutations, respectively, alongside isogenic CRISPR-Cas9-corrected controls. Using cutting-edge techniques developed at Prof. Ana Conesa’s lab (I2SysBio, Valencia, Spain), I will: (i) characterise the complex transcriptional landscape associated with each mutation; (ii) predict the disruption of cis-acting regulators and related consequences on transcript structure; (iii) evaluate downstream cellular pathways compromised by altered transcripts; and (iv) cross-evaluate the fAD transcriptional signatures detected against published datasets from sporadic AD (sAD) patients. During my secondment at the Institute of Neurosciences (Alicante, Spain), I will be excellently trained in wet-lab assays to acquire practical skills in shRNA delivery and cell line manipulation within the context of neurodegeneration.This work will provide critical insights into the transcriptional mechanisms underlying fAD pathogenesis, aiming to inform the identification of novel RNA therapeutic strategies with applications in sAD.