Transcriptomics of independent CRISPR-edited cell lines reveal ciliary-specific ARL13B-dependent changes
Year: 2026
Type: article
Abstract: Primary cilia coordinate signaling pathways that regulate tissue homeostasis and development, and defects in cilia contribute to numerous ciliopathies. However, the transcriptional consequences of disrupting ciliary protein localization remain poorly defined. ARL13B is a cilia-enriched regulatory GTPase required for ciliary trafficking and signaling. The ARL13BV358A variant is undetectable in cilia yet retains known biochemical functions, providing a unique model to investigate the functions of ciliary ARL13B independently of ciliogenesis. To define transcriptional programs associated with loss of ciliary ARL13B, we generated 2 independent Arl13bV358A/V358A kidney epithelial cell lines and matched rescue lines. The ARL13BV358A mutation did not affect ciliation frequency or cilia length but altered ciliary protein composition, including loss of ARL3 and INPP5E localization and increased accumulation of TULP3. RNA sequencing revealed expression changes in genes associated with ciliary biology, mechanotransduction, epithelial organization, and kidney-related phenotypes. Despite similar ciliary phenotypes, the independently derived mutant clones displayed substantial transcriptomic heterogeneity, highlighting a potential source of variation in CRISPR-based transcriptional studies. By integrating data from the independent mutant and rescue clones, we identified a high-confidence set of 131 genes whose expression reproducibly tracked with loss and restoration of ciliary ARL13B. Together, these findings demonstrate that ciliary ARL13B is required to maintain normal ciliary composition and gene expression programs and underscore the value of multiclone, rescue-based experimental designs for robust transcriptomic analyses.
Source: G3 Genes Genomes Genetics
Authors: Olivia Morrison, Tamara Caspary
Institution: Emory University
Language: English
Subfield: Genetics
Domain: Life Sciences
Open Access status: gold
Estimated APC paid: $2,709
OpenAlex ID: W7223198584