P03-04[C09-06]DJM-1 cells deficient in BP180/Type XVII collagen exhibit rudiment hemidesmosomes and abnormal pattern of laminin-332 matrix

Type I hemidesmosomes (HDs) are cell-to-matrix adhesions that are found mainly in stratified epithelia including epidermis. Type I HDs contain plectin and BP230 as linker proteins, and integrin alpha 6 beta 4 and BP180 as transmembrane adhesion receptors. Laminin 332 is known as a major matrix ligand for HDs. Genetic defects in the genes encoding these HD components caused various types of epidermolysis bullosa, indicating importance of HDs in the connection between epidermal basal cells and underlying basement membranes. Recently, we reported that DJM-1 cells, a human squamous cell carcinoma cell line, were able to form mature HDs in a long-term culture condition with a serum-free medium containing calcium at a low concentration. In this study, we produced DJM-1 cells deficient in BP180 by DNA editing using CRISPR/Cas9 system. Under the long-term culture condition, the DJM-1 cells lacking BP180 formed HD-like adhesions, in which BP230 was completely absent and plectin distribution was markedly reduced. The deposited laminin-332 matrix showed more diffuse pattern compared to the matrix deposited by parental cells. These phenotypes were rescued by the transfection of the cDNA encoding the full-length human BP180. The results indicate that BP180 plays an important role not only in the formation of HDs but also in the matrix assembly of laminin-332, which may underlie the pathological mechanism of junctional epidermolysis bullosa. In addition, our study demonstrated that DJM-1 is a useful cell line for producing a cell model of its epidermolysis bullosa to investigate pathological mechanism and to test therapeutic reagents.