¹Department of Biochemistry & Molecular Biology, The Pennsylvania State University, University Park, Pennsylvania, PA 16802, USA

²Department of Veterinary Science, The Pennsylvania State University, University Park, Pennsylvania, PA 16802, USA

³Program in Immunobiology, The Pennsylvania State University, University Park, Pennsylvania, PA 16802, USA

Correspondence to: D M Wojchowski, 115 Henning Building, The Pennsylvania State University, University Park, PA 16802, USA

^aS Zhao and JN Geiger contributed equally to this work

The hematopoietic cell S/T kinase Pim-1 was originally discovered as a target of murine leukemia provirus integration, and when expressed at increased levels is predisposing to lymphomagenesis. Recently, Pim-1 has been shown to enhance the activities of p100, c-Myb and cdc25a, and in part this might explain reported effects on mitogenesis. In the context of cytokine withdrawal, Pim-1 also can attenuate programmed cell death (PCD). Cytokine withdrawal, however, alters signaling pathways and can complicate the dissection of mitogenic vs apoptotic responses. To better study possible effects of Pim-1 on PCD, a hematopoietic cell model was developed in which proliferation was supported efficiently by SCF plus EPO in the absence of endogenous Pim-1 gene expression. This was provided by factor-dependent FDCW2 cells that express endogenous and functional c-Kit, and were transfected stably with truncated Epo receptor form mutated at a Y343 STAT5 binding site. In proliferating cells, exogenously expressed Pim-1 was observed to efficiently inhibit PCD as induced by either Co⁶⁰ or adriamycin, and the dose-dependent nature of this effect was established in several independent clones. By comparison, effects of exogenous Pim-1 on mitogenesis were nominal. In addition, in cell fractionation studies an estimated 25% of M_r 34 000 Pim-1 (but not M_r 44 000 Pim-1) was present in nuclear extracts. Thus, Pim-1 efficiently buffers hematopoietic progenitor cells against death as induced by several clinically important apoptotic agents, and may directly target nuclear effectors. Oncogene (2000) 19, 3684-3692

Pim-1 kinase; genotoxin-induced apoptosis

PCD, programmed cell death; S/T, serine/threonine; SCF, stem cell factor; EPO, erythropoietin; IL, interleukin; GM-CSF, granulocyte macrophage colony stimulating factor; Tpo, thrombopoietin; M_r, molecular weight (relative mass); ER, erythropoietin receptor; TUNEL, TdT-mediated X-dUTP nick end labeling; [³H]dT, [methyl-³H]thymidine; RTK, receptor tyrosine kinase; PCNA, proliferating cell nuclear antigen; ATM, ataxia-telangiectasia mutated; PARP, poly (ADP ribose) polymerase; JNK, c-jun N-terminal kinase; PKC, protein kinase C; ROS, reactive oxygen species; PKC, protein kinase C; PCR, polymerase chain reaction; FBS, fetal bovine serum; PBS, phosphate buffer saline; PI, propidium iodide; FITC, fluorescein isothiocyanate