Description

The CD81 Knockout HMY2-CIR Cell Line is a CRISPR/Cas9-edited knockout cell line in which human CD81 has been disrupted. This model provides a defined loss-of-function system to study CD81 functions in tetraspanin-enriched microdomain organization, cellular adhesion, signal transduction, and hepatitis C virus entry. The knockout is generated in the HMY2-CIR B-lymphoblastoid background, enabling mechanistic studies without the pleiotropic effects of pharmacological inhibitors or RNA interference.

The parental HMY2-CIR is an EBV-transformed B-lymphoblastoid cell line deficient in MHC class II expression and antigen processing. These cells retain humoral immune features including antibody secretion but cannot present exogenous antigens via MHC class II. The background is widely used to study alternative antigen-presentation pathways and B-cell signaling independently of classical MHC restriction. EBV transformation confers continuous proliferation and latent gene expression, including the viral protein LMP1, which influences host signaling.

CD81 encodes a tetraspanin that scaffolds membrane microdomains and interacts with integrins ??4??1 and ??6??1, and tetraspanins CD9, CD63, and CD82. In B cells, CD81 associates with CD19 and CD21 to co-modulate BCR signaling, activating downstream Akt, ERK1/2, NF-??B, and Rac1. Upstream regulators include IL-4, STAT6, TGF-??, and EBV LMP1. CD81 signaling proceeds through FAK, PI3K, and Src kinases. As the HCV entry receptor, CD81 binds viral E2 glycoprotein and cooperates with claudin-1 for internalization.

In HMY2-CIR, CD81 knockout abrogates contributions to membrane organization and signal integration. The MHC class II deficiency emphasizes tetraspanin-mediated co-stimulation and adhesion, making this line valuable for studying CD81-dependent immune synapse formation and integrin signaling without MHC II interference. Loss of CD81 likely alters CD19/CD21 clustering, impairing BCR-mediated PI3K/Akt and MAPK/ERK activation. Additionally, these cells resist HCV pseudoparticle entry, serving as a negative control for viral assays.

Applications include HCV pseudoparticle entry assays, co-immunoprecipitation of tetraspanin-enriched microdomains, and cell adhesion/migration assays. The knockout line is suitable for flow cytometry and western blotting to confirm CD81 loss and assess changes in CD9 and CD63. Phospho-signaling analysis can dissect CD81-dependent kinase activation downstream of BCR or integrins. These tools support research in virology, cancer metastasis, immunodeficiency, and autoimmunity. For further details, contact Ascent Research.