FIGURE 2.

Matriptase-HAI-1 complex accumulates in intracellular structures. Caco-2 cells grown on Transwell filters, were fixed, permeabilized, and immunolabeled with antibodies against total matriptase (M32) and HAI-1 (A–C) or matriptase-HAI-1 complex (M69) and HAI-1 (D–F). Using a confocal scanning microscope, images were taken in the XY plane showing a single section through the monolayer and the XZ plane showing a cross section of the monolayer. The position of the plane of the XY section is indicated on the XZ plane (black arrows) on the right. The scale bar represents 10 μm. A–C, Matriptase was detected on the basolateral plasma membrane of the Caco-2 cells co-localizing with HAI-1. Matriptase and HAI-1 was also co-localizing in structures near the apical membrane (white arrowheads). D–F, matriptase-HAI-1 complex was observed in structures near the apical plasma membrane (white arrowheads). Low amounts of matriptase-HAI-1 complex were detected on the basolateral plasma membrane. HAI-1 was detected both on apical and basolateral plasma membranes as well as in the apical structures co-localizing with matriptase. G and H, Caco-2 cells were treated with or without Triton X-100 prior to immunolabeling with the antibody M69. G, in the cells permeabilized with Triton X-100, a distinct detection of matriptase-HAI-1 complex was observed in apical vesicular structures. H, in cells without permeabilization, the vesicular structures with matriptase-HAI-1 complex were not detected. Only a weak basolateral signal was observed. The nuclei were visualized by DAPI staining shown in blue. Results shown are representative of three independent experiments.