The OPTN Knockout Raji Polyclonal Cells are a CRISPR/Cas9-edited polyclonal knockout cell population in which the OPTN (optineurin) gene has been disrupted to create a loss-of-function model. This polyclonal population, derived from the human Raji B lymphocyte line, provides a genetically heterogeneous knockout background that allows researchers to study OPTN-dependent functions without clonal bias. CRISPR/Cas9-mediated gene disruption targets the OPTN locus, abrogating expression of the full-length optineurin protein and its associated cellular activities.
The Raji host cell line is an Epstein-Barr virus (EBV)-positive B lymphocyte model originally isolated from a Burkitt lymphoma patient. These cells retain hallmarks of mature B cells, including robust antibody production and engagement of adaptive immune signaling pathways. The Raji background is widely employed in immunology and cancer research due to its well-characterized B cell receptor signaling, NF-??B activation, and susceptibility to lytic viral reactivation, making it a relevant system for dissecting immune regulatory mechanisms and lymphomagenesis.
OPTN encodes an autophagy receptor that bridges ubiquitinated cargo to autophagosomes via its C-terminal LC3-interacting region (LIR) and UBAN domain, respectively. It is a key substrate of TANK-binding kinase 1 (TBK1), which phosphorylates OPTN at Ser172 to enhance its autophagic activity. OPTN functions as a negative regulator of NF-??B signaling by interacting with components of the IKK complex, including TANK, TRAF3, and NEMO, and is implicated in the selective removal of damaged mitochondria (mitophagy) and intracellular pathogens (xenophagy). Upstream triggers such as TNF-?? and interferon-?? converge on TBK1-mediated OPTN activation, while downstream OPTN effectors include the LC3/GABARAP family, p62/SQSTM1, and NF-??B p65. The mechanistic interplay places OPTN at a critical node where ubiquitin-dependent degradation intersects with inflammatory signaling.
In Raji B lymphocytes, OPTN disruption impairs the selective autophagic clearance of damaged mitochondria, leading to accumulation of depolarized organelles and altered metabolic homeostasis. Concurrently, the loss of OPTN removes constitutive inhibition of the IKK complex, resulting in dysregulated NF-??B activity that may affect B cell survival, differentiation, and cytokine production. This dual perturbation of autophagy and NF-??B pathways creates a powerful model to examine how these processes coordinate adaptive immune responses and contribute to lymphomagenesis when misregulated.
This knockout polyclonal cell population is suited for a range of investigations, including mechanistic studies of mitophagy and xenophagy in B cells, elucidation of OPTN-dependent NF-??B regulatory circuits, and disease modeling of primary open-angle glaucoma, amyotrophic lateral sclerosis, and inflammatory disorders. Functional assays such as Western blotting for LC3-II and p62, immunofluorescence analysis of LC3 puncta, flow cytometry-based mitophagy measurement using MitoTracker, and NF-??B dual-luciferase reporter assays can be employed to assess autophagy flux and signaling output. Additional approaches include co-immunoprecipitation of endogenous TBK1-OPTN complexes, cytokine multiplex profiling, and cell viability assessments under proteotoxic or mitochondrial stress. For further details or to discuss custom applications, please contact Ascent Research.
