Figure 5.

Expression and activation of matriptase in M- and Z-Mɸ. (A) Representative immunoblot of matriptase in cell lysates from M- and Z-Mɸ. Samples were prepared either without adding reducing agent and without boiling (nonboiled, nonreduced [NB, NR]) or with boiling and with reducing agent (B, R). Matriptase is a single polypeptide (B, R samples), and does not form complexes with AAT or other proteins (NB, NR samples). Z-Mɸ have much higher matriptase levels than do M-Mɸ (experiment was performed four times with Mɸ from different individuals). (B) Comparison of matriptase mRNA levels in M- and Z-Mɸ. Each group represents Mɸ from six individuals. (C) Matriptase was activated by incubation of Mɸ in phosphate buffer with mild acidity (pH 6.0) or with PBS (pH 7.4) as control for 20 minutes at 37°C. Shed matriptase was analyzed by immunoblot in NB, NR and B, R conditions. Buffer with pH 6.0 induced increased shedding of matriptase from Z-Mɸ cell surface as noncomplexed protein (NB, NR samples). Activated matriptase (30 kD) could be visualized in R, B samples. (D) Compared with M-Mɸ, Z-Mɸ exhibited increased activity of shed matriptase in conditioned media. Data are representative of three independent experiments.