Oral Presentation Australasian RNA Biology and Biotechnology Association 2024 Conference

A specific variant of NL02, a brain-enriched lncRNA, regulates lysosomal function in human neurons (108984)

Indranil Basak 1 2 , Kirstin O McDonald 1 2 , Alex van der Weerden 1 2 , Robert Day 1 2 , Sarah D Diermeier 1 , Stephanie M Hughes 1 2
  1. Biochemistry, University of Otago, Dunedin, Otago, New Zealand
  2. Brain Health Research Centre, University of Otago, Dunedin, Otago, New Zealand

Long non-coding RNAs (lncRNAs) are best known for their role in regulating gene transcription, mRNA degradation, translation inhibition, microRNA sponging, and protein scaffolding. LncRNA NL02 is highly expressed in healthy human brains without a known brain-specific function. NL02 may have a neurodevelopmental role, and NL02 level has been shown to be decreased in the substantia nigra of Parkinson’s disease patients' brains. Many neurodegenerative diseases, including Parkinson’s and Alzheimer’s disease, show a prominent pathological feature – lysosomal dysfunction and the link between lncRNAs and lysosomes is understudied. Therefore, we aim to discover how novel lncRNAs, like NL02, regulate lysosomal function and neuronal homeostasis.

Using human induced pluripotent stem cell (iPSC)-derived brain cells and bioinformatic analysis, we found that NL02 shows variant-specific expression in human iPSC-derived cortical and dopaminergic neurons and astrocytes. CRISPRi inhibition of two different variants of NL02 in human iPSC-derived cortical neurons showed that inhibition of only one variant (NL02-201) significantly increases Lysotracker signal (1.9 fold increase, p<0.05) compared to control neurons, indicating a change in lysosomal function. NL02-201 inhibition also showed significantly increased cell death (1.2 fold increase, p<0.05) and impaired neuronal activity (measured using calcium transient, p<0.01) compared to control neurons, suggesting a variant-specific role of NL02 in cortical neurons. Preliminary analysis showed both variants were localised in the nucleus of iPSCs. Bioinformatic analysis of the NL02-binding partners indicates that only one exon is the binding site for all the RNA interactions, which belongs to NL02-201 and not the other variant.

Combining our data, we can conclude that NL02 has an uncharacterised function associated with lysosomal function and neuronal homeostasis. Therefore, identifying NL02’s RNA and protein binding partners in different human brain cells will reveal the true lysosome- and brain-specific role of NL02.