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Human interleukin-3 receptor modulates bcl-2 mRNA and protein levels
through protein kinase C in TF-1 cells
MS Rinaudo, K Su, LA Falk, S Halder and RA Mufson
Holland Laboratory for Biomedical Science, American Red Cross, Rockville,
MD 20855, USA.
Upon withdrawal of interleukin-3 (IL-3) from human factor-dependent
erythroleukemic cell line TF-1, bcl-2 mRNA and protein levels decrease
within 8 to 24 hours. Accompanying this decrease is the onset of apoptosis
as determined by flow cytometric analysis of DNA degradation. By 8 to 18
hours of deprivation approximately 70% to 80% of the cells have entered
apoptosis. Downregulation of protein kinase (PK) by a 24- hour incubation
in 100 nmol/L 12-O-tetradecanoyl-phorbol-13-acetate (TPA) in the presence
of IL-3 dramatically reduced bcl-2 mRNA levels, and induced apoptosis in
the presence of IL-3. We have also found that even in the presence of IL-3,
two inhibitors of PKC, light-activated calphostin and H-7, substantially
reduced the levels of bcl-2 mRNA between 8 and 24 hours as measured by a
semi-quantitative reverse transcriptase/polymerase chain reaction assay
method; however, the cyclic nucleotide-dependent PK inhibitor HA 1004, that
is a structural analog of H-7 but a poor inhibitor of PKC, did not reduce
bcl-2 levels in the presence of IL-3. This decrease in bcl-2 mRNA was
accompanied by a decline in bcl-2 protein levels by 8 to 24 hours after
addition of light-activated calphostin. In addition to interfering with the
maintenance of bcl-2 mRNA levels, inhibition of PKC with H-7 inhibited the
induction of bcl-2 mRNA in factor-deprived TF-1 cells restimulated with
IL-3. The cyclic nucleotide-dependent PK inhibitor HA 1004 did not inhibit
IL-3-induced bcl-2 mRNA. Studies with actinomycin D showed that
transcription plays a major role in maintaining bcl-2 levels in TF-1 cells,
and it is therefore likely that IL-3 plays a role in maintaining bcl-2
transcription through activation of PKC in these cells.
Volume 86,
Issue 1,
pp. 80-88,
07/01/1995
Copyright © 1995 by The American Society of Hematology
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