Oral Presentation International Congress on Neuronal Ceroid Lipofuscinoses 2025

Targeted long-read RNA sequencing reveals the complexity of CLN3 transcription and the consequences of the most common 1-kb deletion in patients with juvenile CLN3 disease. (126118)

Christopher Minnis 1 , Hao-Yu Zhang 1 , Emil Gustavsson 1 , Jasmaine Lee 2 , Claire Anderson 3 , Ramy Saad 1 , Emily Gardner 1 , Angela Schulz 4 , Miriam Nickel 4 , Gavin Arno 5 , Neringa Jurkute 6 , Nicola Gammaldi 7 , Filippo M Santorelli 7 , Paul Gissen 1 , Phillipa Mills 1 , Mina Ryten 1 , Sara E Mole 1
  1. Great Ormond Street Institute of Child Health, University College London, London, Greater London, United Kingdom
  2. Department of Neurodegenerative Disease, Queen Square Institute of Neurology, University College London, London, Greater London, United Kingdom
  3. Department of Genetics and Genomic Medicine, Great Ormond Street Institute of Child Health, University College London, London, Greater London, United Kingdom
  4. University Children’s Hospital, University Medical Center Hamburg-Eppendorf, Hamburg, Germany
  5. UCL Institute of Ophthalmology, London, and Moorfields Eye Hospital NHS Foundation Trust, London, UK
  6. Department of Neuro-ophthalmology, The National Hospital for Neurology and Neurosurgery, University College London Queen Square Institute of Neurology, The National Hospital for Neurology and Neurosurgery, London, UK
  7. Molecular Medicine for Neurodegenerative and Neuromuscular Diseases Unit, IRCCS Stella Maris Foundation, Pisa, Italy

A growing body of work indicates a greater diversity of structural transcripts than are currently annotated for most genes in genome databases. This includes genes that underlie life-limiting diseases, such as CLN3 disease. The role of transcript diversity in classic juvenile CLN3 disease is poorly understood, and the consequences of the 1-kb deletion are unclear. Here, we report our findings from the first long-read RNA sequencing targeting CLN3 in blood samples derived from patients clinically and genetically diagnosed with juvenile CLN3 disease and control individuals. CLN3 transcription is complex, with >600 different transcripts encoding 38 different open reading frames (ORF) and no dominantly expressed transcript. The 1-kb deletion causes a total loss of some transcripts encoding protein isoforms of different lengths, including the canonical 438 amino acid protein (CLN3-438aa) and other significantly smaller isoforms such as CLN3-384aa, CLN3-421aa, and CLN3-223aa (ATG start in exon 8). The highest expressed disease transcripts are CLN3-181aa (lacking only exons 7 and 8), and novel isoforms that lack additional exons and so encode longer ORFs: CLN3-339aa (exon 5), CLN3-315aa (exons 4, 5), CLN3-311aa (exons 3, 4) and CLN3-328aa (exon 9). Notably, the disease transcript that encodes CLN3-181aa is also detected at low levels in healthy controls. Although predicted to be degraded by nonsense-mediated decay, it appears to be translated into protein, as its unique peptide sequence, ‘AQPGSPS’, is detected in mass spectrometry databases. Together, these findings confirm the complexity of transcription at the CLN3 locus, reveal the impact of the 1-kb deletion on isoform abundance, and highlight the importance of understanding the contribution of these isoforms to CLN3 function in health and disease. Moreover, they impact the future design and development of personalised therapeutics and the design and generation of disease models. Finally, they underline the importance of complete annotation for disease genes.

  1. Targeted long-read RNA sequencing reveals the complexity of CLN3 transcription and the consequences of the most common 1-kb deletion in patients with juvenile CLN3 disease. Christopher J Minnis, Hao-Yu Zhang, Emil Gustavsson, Jasmaine Lee, Claire Anderson, Emily Gardner, Angela Schulz, Miriam Nickel, Gavin Arno, Neringa Jurkute, Andrew R Webster, Nicola Gammaldi, Filippo M Santorelli, Paul Gissen, Phillipa Mills, Mina Ryten, Sara E Mole bioRxiv 2025.04.24.650398; doi: https://doi.org/10.1101/2025.04.24.650398