Characterization of caspase proteases in cytokine-dependent myeloid progenitor cells using enzyme affinity labeling
✍ Scribed by Richard L. Cho; Daniel E. Johnson
- Publisher
- John Wiley and Sons
- Year
- 1999
- Tongue
- English
- Weight
- 148 KB
- Volume
- 73
- Category
- Article
- ISSN
- 0730-2312
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✦ Synopsis
Bone marrow-derived myeloid progenitor cells are dependent on the presence of cytokines such as interleukin-3 (IL-3) for their survival. The withdrawal of IL-3 from IL-3-dependent myeloid progenitors results in death via an apoptotic program. Previous studies have shown that IL-3 withdrawal induces the activities of caspase proteases. However, the molecular identities of myeloid progenitor caspases have not been determined. In this study, we used an affinity labeling reagent (biotin-YVAD-acyloxymethyl ketone) that binds to processed active caspase subunits, to study caspase activation in 32D and FDCP-1 myeloid progenitor cells. After IL-3 withdrawal, we detected affinity labeling of caspase subunits of 20, 17, and 16 kDa in both cell lines. Surprisingly, affinity labeling of the 20-and 17-kDa proteins, but not the 16-kDa protein, was also detected in healthy cells maintained in the presence of IL-3. By contrast, in cytokine-independent cell lines, affinity labeling of caspase subunits was detected only after treatment with an apoptotic stimulus. Immunoblotting experiments showed that caspase-3 constitutes at least a portion of the 20-and 17-kDa affinity-labeled proteins detected in the myeloid progenitor cell lines. Taken together, these data provide direct evidence of caspase activation in cytokine-dependent myeloid progenitors, and suggest that unique apoptotic pathways may exist in these cells.