Targeted CRISPRi screening reveals unexpected resilience across the RNA polymerase III transcriptome
- 1Amsterdam UMC, Vrije Universiteit Amsterdam
Abstract
Increased RNA polymerase III (Pol III) activity and tRNA abundance are widely linked to cancer cell growth, yet the functional requirement for individual Pol III genes and core components remains unclear, in part due to the difficulty of achieving gene-specific perturbation of highly conserved loci. Here, we developed an inducible CRISPR interference platform and a custom single-guide RNA (sgRNA) library enabling gene-specific targeting of Pol III-transcribed genes and Pol III machinery. Genome-wide screening identified several Pol III dependencies in diploid fibroblasts and HEK293T cells, including multiple initiator methionine tRNA genes among the strongest fitness dependencies. Unexpectedly, glioblastoma models remained largely insensitive to repression of both individual Pol III genes and core Pol III components, despite efficient target repression. These findings establish a general strategy for gene-specific interrogation of conserved Pol III genes and indicate that glioblastoma models tolerate extensive perturbation of Pol III genes and machinery.
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BibTeX
@misc{simeonov2026targeted,
title = {{Targeted CRISPRi screening reveals unexpected resilience across the RNA polymerase III transcriptome}},
author = {B. Simeonov and Z. Veryeri and Y. Faas and A. Gerber},
howpublished = {bioRxiv},
year = {2026},
month = sep,
doi = {10.64898/2026.09.24.754042},
url = {https://www.biorxiv.org/content/10.64898/2026.09.24.754042v1},
note = {CC BY-NC-ND},
}
RIS
TY - UNPB
TI - Targeted CRISPRi screening reveals unexpected resilience across the RNA polymerase III transcriptome
AU - B. Simeonov
AU - Z. Veryeri
AU - Y. Faas
AU - A. Gerber
PY - 2026
DA - 2026/09/24
PB - bioRxiv
DO - 10.64898/2026.09.24.754042
UR - https://www.biorxiv.org/content/10.64898/2026.09.24.754042v1
AB - Increased RNA polymerase III (Pol III) activity and tRNA abundance are widely linked to cancer cell growth, yet the functional requirement for individual Pol III genes and core components remains unclear, in part due to the difficulty of achieving gene-specific perturbation of highly conserved loci. Here, we developed an inducible CRISPR interference platform and a custom single-guide RNA (sgRNA) library enabling gene-specific targeting of Pol III-transcribed genes and Pol III machinery. Genome-wide screening identified several Pol III dependencies in diploid fibroblasts and HEK293T cells, including multiple initiator methionine tRNA genes among the strongest fitness dependencies. Unexpectedly, glioblastoma models remained largely insensitive to repression of both individual Pol III genes and core Pol III components, despite efficient target repression. These findings establish a general strategy for gene-specific interrogation of conserved Pol III genes and indicate that glioblastoma models tolerate extensive perturbation of Pol III genes and machinery.
ER -