This product consists of a CRISPR/Cas9-edited polyclonal knockout cell population in which the DNAH5 gene has been disrupted within Raji cells through genome-wide gene disruption methods. The polyclonal format provides a heterogeneous pool of edited cells, each carrying distinct genomic modifications at the targeted locus, enabling robust loss-of-function studies without clonal isolation. By disrupting the endogenous DNAH5 locus, researchers can model the absence of functional axonemal dynein heavy chain 5, a component essential for outer dynein arm assembly and ciliary motility, in a human B lymphocyte background. This gene-edited population is produced via CRISPR/Cas9-mediated gene targeting and is supplied as a ready-to-use polyclonal pool suitable for immediate downstream applications, including protein expression analysis, interaction studies, and functional screening.
The Raji host cell line is an Epstein-Barr virus (EBV)-positive B lymphocyte cell line originally derived from a Burkitt lymphoma patient. As a suspension-adapted lymphoid cell model, Raji cells retain critical features of antigen-presenting cells and are widely used in immunology, virology, and cancer biology. They endogenously express key components of the major histocompatibility complex (MHC) class I and class II pathways, supporting their role in antigen presentation and T cell activation studies. The EBV-transformed nature of these cells also provides a tractable system for studying viral latency and lymphomagenesis. Importantly, Raji cells do not normally express motile cilia, so DNAH5 knockout in this context creates a unique platform for investigating ciliary protein functions outside of the ciliated cell environment, including potential non-ciliary roles and protein interaction networks.
DNAH5 encodes a heavy chain subunit of the outer dynein arm, a large motor protein complex that powers ciliary and flagellar beating. The DNAH5 protein interacts directly with other dynein components such as DNAI1, DNAI2, and DNAL1 to assemble a functional outer dynein arm along the axonemal microtubules. Its expression and assembly are transcriptionally regulated by upstream factors including FOXJ1, RFX2, and RFX3, which are master regulators of ciliogenesis and motile cilia differentiation. Notch signaling also modulates ciliogenesis pathways that ultimately converge on genes like DNAH5. Downstream, functional DNAH5 is required for proper ciliary beat frequency and efficient mucociliary clearance, processes that depend on the coordinated movement of multiple dynein arm components. In the Raji knockout model, disruption of DNAH5 protein expression abolishes its interactions with DNAI1 and other outer dynein arm partners, making it an effective tool for dissecting assembly pathways and protein?Cprotein interactions outside the context of intact cilia.
Despite the Raji cell line??s lack of motile cilia, the DNAH5 knockout model holds significant value for studying primary ciliary dyskinesia (PCD) and related syndromes. Because many ciliary proteins, including DNAH5, have expression patterns and interacting partners that extend beyond ciliated tissues, this B lymphocyte model allows investigation of DNAH5??s biochemical properties, post-translational modifications, and binding interactions in a well-characterized, easily manipulated host. The model can be used to validate antibodies raised against DNAH5 or other outer dynein arm proteins, ensuring specificity in a null background. Moreover, the Raji system supports high-throughput screening for small molecules or genetic modifiers that may influence DNAH5 expression or stability, a context particularly relevant for disorders like Kartagener syndrome and chronic respiratory infections where DNAH5 mutations are causative.
Researchers can employ this polyclonal knockout population in a variety of experimental workflows, from co-immunoprecipitation and mass spectrometry-based interactomics to immunofluorescence validation of commercial antibodies. Western blotting using lysates from these cells serves as a clear negative control for detecting DNAH5 protein expression, while flow cytometry permits rapid assessment of any surface markers potentially affected by DNAH5-related pathways. The combination of a knockout B lymphocyte host and DNAH5 disruption is especially suited for studying humoral immune function and antigen presentation when ciliary protein expression may play an underappreciated role. Additional applications include probing the influence of upstream regulators like FOXJ1 or RFX2 on DNAH5 downstream targets, and comparative proteomics to identify novel binding partners. For further details or custom requests, please contact Ascent Research.
