The PIM-1 serine kinase prolongs survival and inhibits apoptosis-related mitochondrial dysfunction in part through a bcl-2-dependent pathway

We have examined potential mechanisms by which the Pim-1 kinase acts as a h ematopoietic cell survival factor. Enforced expression of the wild type 33 kd (FD/hpim33) and 44 kd (FD/mpim44) Pim-1 proteins in murine factor-depend ent FDCP1 cells prolonged survival after withdrawal of IL-3, while expressi on of a dominant negative Pim-1 protein (FD/pimNT81) shortened survival, Fo llowing removal of IL-3 FDCP1 cells exhibited loss of mitochondrial transme mbrane potential and production of reactive oxygen species, as determined b y how cytometry analysis. The wild type Pim-1 proteins decreased these chan ges while the dominant negative protein enhanced mitochondrial dysfunction, The anti-apoptotic activity of the kinases could not be attributed to modu lation of glutathione, catalase, or superoxide dismutase activities, Both t he FD/hpim33 and FD/mpim44 cells maintained expression of bcl-2 mRNA follow ing cytokine removal, while a substantial decrease was seen in FD/neo cells . To modulate Bcl-2 protein levels, a bcl-2 antisense RNA construct was coe xpressed with the wild type pim-1 cDNAs, FD/hpim33 cells with low cellular Bcl-2 protein levels had shortened cytokine-independent survival compared w ith FD/hpim33 clones with high Bcl-2 expression. However survival of FD/mpi m44 cells after IL-3 withdrawal was substantially independent of cellular B cl-2 protein levels. The 33 kd protein delayed, and the 44 kd protein compl etely prevented enhanced cell death associated with enforced expression of human Bar protein however. Our results suggest that the 33 kd Pim-1 kinase may enhance cell survival through cooperation with and regulation of bcl-2, In addition the 44 kd kinase may regulate the expression or activity of ot her pro- and anti-apoptotic members of the bcl-2 family.