Lysosomal function is essential for cellular homeostasis. This is highlighted by the involvement of dysfunctional lysosomes in Alzheimer’s and Batten disease. Understanding regulatory networks involved in neuronal function is key. Recently, lncRNAs have emerged as a contributor to cellular regulation including autophagy, the degradation process reliant on lysosomes. 

This project focuses on the lncRNA DANCR and its impact on neuronal lysosomes. Initially described as a regulator of differentiation, DANCR is known to regulate autophagy. However, no studies have investigated the influence of DANCR on lysosomes in human neurons. 

Transcript variants of DANCR remain unexplored. To address this, Ensembl was used to select the four most biologically relevant DANCR transcripts. This also revealed DANCR shares a transcriptional start site with the uncharacterised lncRNA LINC01618. A qPCR panel was developed to differentiate between four DANCR and four LINC01618 transcripts. In iPSCs, five out of eight transcripts were detected. CRISPRi guides were designed to target transcripts of DANCR or LINC01618. The knockdown iPSC lines showed 99.6% (P=0.0093) and 99.2% (P=0.0500) reduction in the expression of targeted DANCR and LINC01618 transcripts respectively. Interestingly, the DANCR knockdown line showed almost 2-fold increase (P=0.0378) in proliferation compared to a control line. 

Neurons derived from knockdown iPSC lines will be used to investigate the impact of DANCR and LINC01618 on differentiation and lysosomal function. RNAseq will uncover molecular pathways impacted by DANCR and LINC01618 in the knockdown lines. This will reveal the relationship between these lncRNAs and lysosomal regulation, and point towards potential therapeutic strategies.