The organization of LETS protein on the surface of NIL8 hamster cells has been examined by immunofluorescence staining. The distribution of LETS protein was found to depend on the culture conditions; in subconfluent, low-serum arrested cultures the LETS protein is predominantly located at the cell-substrate interface and also in regions of cell-cell contact, whereas in dense cultures the cells are surrounded by a network of LETS protein fibrils. Transformed derivatives of these cells exhibit only sporadic staining for LETS protein, in the form of short intercellular bridges. Agents that cause alterations in cell shape and cytoplasmic filaments have been used to explore the relationship of LETS protein to the internal cytoskeletal elements. Reciprocally, perturbations of the cell surface were examined for their effects on internal filaments. The arrangement of microtubules seems to be unrelated to the presence of LETS protein in the cells studied. Actin microfilament bundles and LETS protein respond in a coordinate fashion to some perturbants but independently with respect to others. The patterns of staining for LETS protein are consistent with an involvement in cell-to-cell and cell-to-substrate adhesion.