A cytotoxic T-cell line, CTLL-2 cells, showed spreading after adhering to extracellular matrix proteins such as fibronectin (FN), laminin (LN) and hyarulonic acid (HA). The adhesion of CTLL-2 cells to LN was mediated by very late activation antigen-6 (VLA-6). Expression of interferon-γ (IFN-γ) mRNA was enhanced in CTLL-2 cells, also when they adhered to extracellular matrix proteins; and the enhanced IFN-γ mRNA expression by adhering to LN was blocked by anti-αa6 antibody. Calphostin C, a protein kinase C (PKC) inhibitor, markedly inhibited the enhancement of IFN-γ mRNA expression in a dose-dependent manner, which suggested that PKC acted as a second messenger in the IFN-γ mRNA expression mediated by the interaction of VLA-6 with LN in CTLL-2 cells. Furthermore, confocal laser-microscopic analysis and Western blot analysis revealed that PKC-α was activated after CTLL-2 cells adhered to LN. PKC activity translocated from the cytosol fraction to the particulate fraction, after CTLL-2 cells adhered to LN. Altogether, we suggest that PKC plays an important role in the signal transduction for IFN-γ mRNA expression after cytotoxic T cells adhere to LN.