American Society of Hematology

Figure 4

$Figure 4. The cytoplasmic tail of CD45 is proteolytically cleaved and released into the cytoplasm. (A) Inhibition of de novo protein synthesis by cycloheximide (2 and 10 μg/mL) did not block production of ct-CD45 upon monocyte stimulation with PMA or zymosan. (B) The activity of protease inhibitors AEBSF, aprotinin, EGTA, calpeptin, E64d, MG-132, and pepstatin on production of ct-CD45 in zymosan-stimulated monocytes was tested in Western blots. (C) Membrane and cytoplasmic fractions of resting monocytes and monocytes stimulated for 90 minutes with PMA were isolated and analyzed in Western blots with mAb 8-301. Presence of the serine protease inhibitor AEBSF inhibited production of membrane-bound ct-CD45. The γ-secretase inhibitor X blocked the release of membrane-bound ct-CD45 into the cytosol. Isolation of membranes was controlled with anti-gp91phox, isolation of cytoplasm with an actin antibody. Data represent at least 2 independently performed experiments.$

The cytoplasmic tail of CD45 is proteolytically cleaved and released into the cytoplasm. (A) Inhibition of de novo protein synthesis by cycloheximide (2 and 10 μg/mL) did not block production of ct-CD45 upon monocyte stimulation with PMA or zymosan. (B) The activity of protease inhibitors AEBSF, aprotinin, EGTA, calpeptin, E64d, MG-132, and pepstatin on production of ct-CD45 in zymosan-stimulated monocytes was tested in Western blots. (C) Membrane and cytoplasmic fractions of resting monocytes and monocytes stimulated for 90 minutes with PMA were isolated and analyzed in Western blots with mAb 8-301. Presence of the serine protease inhibitor AEBSF inhibited production of membrane-bound ct-CD45. The γ-secretase inhibitor X blocked the release of membrane-bound ct-CD45 into the cytosol. Isolation of membranes was controlled with anti-gp91^phox, isolation of cytoplasm with an actin antibody. Data represent at least 2 independently performed experiments.

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