CRISPRa-mediated activation of Tfeb in a fibril-induced alpha-synuclein aggregation model
- 1Lund University
Abstract
Accumulation of misfolded alpha-synuclein (a-syn) is a key hallmark of Parkinson's disease (PD), making it a key target for novel therapeutic strategies. Here, we use a previously described screening assay to select multiple efficient single guide RNA (sgRNA) combinations targeting transcription factor EB (TFEB) for CRISPR-activation (CRISPRa) using a nuclease-deficient Staphylococcus aureus Cas9 coupled to a truncated VP64-p53-Rta (VPR) activator complex. Selected sgRNA are applied in a rat neuroblastoma in-vitro model of fibril-induced a-syn aggregation with the goal of inducing autophagy and reduce protein aggregation. We successfully induce a-syn aggregation in this model and observe a stagnation in phosphorylated a-syn (p-a-syn) compared to non-phosphorylated a-syn across treatment groups. While we achieve robust expression of Tfeb in the exogenous screening assay and at endogenous mRNA level, we did not observe proportional increase in protein expression. Additionally, we do not observe any additive effects of CRISPRa induction combined with media treatments inducing Tfeb nuclear translocation. Taken together, these findings illustrate difficulties in epigenetic induction of Tfeb using CRISPRa and highlights the need for additional studies exploring this application.
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BibTeX
@misc{arvidsson2026crispramediated,
title = {{CRISPRa-mediated activation of Tfeb in a fibril-induced alpha-synuclein aggregation model}},
author = {E. Arvidsson and E. Pi Börjesson and E. Sabarese and L. Quintino and C. Lundberg},
howpublished = {bioRxiv},
year = {2026},
month = oct,
doi = {10.64898/2026.10.16.757246},
url = {https://www.biorxiv.org/content/10.64898/2026.10.16.757246v1},
note = {CC BY},
}
RIS
TY - UNPB
TI - CRISPRa-mediated activation of Tfeb in a fibril-induced alpha-synuclein aggregation model
AU - E. Arvidsson
AU - E. Pi Börjesson
AU - E. Sabarese
AU - L. Quintino
AU - C. Lundberg
PY - 2026
DA - 2026/10/16
PB - bioRxiv
DO - 10.64898/2026.10.16.757246
UR - https://www.biorxiv.org/content/10.64898/2026.10.16.757246v1
AB - Accumulation of misfolded alpha-synuclein (a-syn) is a key hallmark of Parkinson's disease (PD), making it a key target for novel therapeutic strategies. Here, we use a previously described screening assay to select multiple efficient single guide RNA (sgRNA) combinations targeting transcription factor EB (TFEB) for CRISPR-activation (CRISPRa) using a nuclease-deficient Staphylococcus aureus Cas9 coupled to a truncated VP64-p53-Rta (VPR) activator complex. Selected sgRNA are applied in a rat neuroblastoma in-vitro model of fibril-induced a-syn aggregation with the goal of inducing autophagy and reduce protein aggregation. We successfully induce a-syn aggregation in this model and observe a stagnation in phosphorylated a-syn (p-a-syn) compared to non-phosphorylated a-syn across treatment groups. While we achieve robust expression of Tfeb in the exogenous screening assay and at endogenous mRNA level, we did not observe proportional increase in protein expression. Additionally, we do not observe any additive effects of CRISPRa induction combined with media treatments inducing Tfeb nuclear translocation. Taken together, these findings illustrate difficulties in epigenetic induction of Tfeb using CRISPRa and highlights the need for additional studies exploring this application.
ER -