The DNAAF5 Knockout HeLa Polyclonal Cells product is a CRISPR/Cas9-mediated gene-disrupted polyclonal population targeting DNAAF5 in HeLa cells. This polyclonal format provides heterogeneous loss-of-function alleles, suitable for population-level studies without clonal selection. It enables investigation of DNAAF5??s role in cytoplasmic dynein arm assembly within a human epithelial background.
HeLa cells, derived from cervical adenocarcinoma, are a widely used epithelial cell line with robust growth and high transfection efficiency. Although normally non-ciliated, they can be induced to form primary cilia upon serum starvation, retaining the cytoplasmic machinery required for preassembly of dynein components. This versatility makes HeLa an effective host for studying early ciliogenic events and for examining the molecular machinery of dynein assembly.
DNAAF5 is a cytoplasmic co-chaperone essential for preassembly of axonemal dynein arms, interacting with PIH1D3 and HSP70 to stabilize dynein heavy chain subunits such as DNAH5 and DNAH9. Its expression is transcriptionally regulated by RFX3 and FOXJ1, and it operates downstream of Notch signaling. DNAAF5 functions within the R2TP complex together with other dynein assembly factors like DNAAF1, DNAAF2, and DNAAF3. Loss of DNAAF5 prevents proper formation of both outer and inner dynein arms, leading to immotile cilia characteristic of primary ciliary dyskinesia (PCD) and Kartagener syndrome, which involve chronic respiratory infections, laterality defects, and infertility.
In HeLa cells, DNAAF5 knockout generates a model for investigating the cytoplasmic phase of dynein arm assembly independent of complete ciliogenesis. This facilitates biochemical dissection of preassembly complexes and permits assessment of dynein subunit trafficking to the ciliary base. The lack of mature motile cilia under standard culture conditions concentrates investigations on early assembly events, offering a simplified platform for compound screening to restore dynein formation.
Typical research applications include ciliogenesis induction assays with immunofluorescence detection of ciliary markers ARL13B and acetylated tubulin, western blotting of dynein subunits DNAH5 and DNAI1 to monitor assembly, co-immunoprecipitation to characterize DNAAF5 interactions with PIH1D3 and HSP70, and RT-qPCR profiling of ciliary gene expression. The polyclonal knockout population is well suited for PCD disease modeling, dynein assembly mechanistic studies, and high-throughput drug screening for ciliopathies. For additional information, please contact Ascent Research.
