ABSTRACT
The mitogen-dependent induction of cyclin D-dependent kinase activity is required for cells to enter the DNA synthetic (S) phase of their division cycle. Immature 32Dcl3 myeloid cells (32D) proliferating in the presence of interleukin-3 (IL-3) normally express cyclins D2 and D3, which assemble into binary holoenzyme complexes with their catalytic subunits, CDK4 and CDK6. When 32D cells are switched to medium containing granulocyte colony-stimulating factor (G-CSF) instead of IL-3, D-type cyclins are degraded and, in the absence of their associated kinase activity, the cells arrest in the first gap phase (G1) of the cell cycle and differentiate to neutrophils. We derived 32D cells in which the expression of p19INK4d, a specific polypeptide inhibitor of CDK4 and CDK6, is regulated by the heavy metal-inducible sheep metallothionein promoter. Induction of p19INK4d in response to zinc prolonged cell survival in the absence of growth factor treatment. When maintained in medium containing both IL-3 and zinc, these cells lost cyclin D-dependent kinase activity, underwent G1 phase arrest, and acquired certain morphologic, antigenic, and functional properties of mononuclear phagocytes. Cells induced to express p19INK4d did not synthesize receptors for macrophage colony-stimulating factor (M-CSF/CSF-1) and reverted to an immature myeloid phenotype when shifted back into medium containing IL-3 alone. These cells exhibited accelerated differentiation to neutrophils in response to G-CSF but also gave rise to macrophage-like cells when maintained in medium containing both G-CSF and zinc. Therefore, the acquisition of macrophage properties in response to zinc treatment neither depended upon IL-3 nor upon G1 phase arrest per se and instead reflects some ability of p19INK4d, and presumably cyclin D-dependent kinases, to affect myeloid differentiation.
Subject(s)
Carrier Proteins/biosynthesis , Cell Cycle Proteins , Cell Lineage/genetics , Cyclin-Dependent Kinase Inhibitor p16 , Cyclin-Dependent Kinases/antagonists & inhibitors , Macrophages/cytology , Carrier Proteins/genetics , Cell Differentiation/genetics , Cell Line , Cyclin-Dependent Kinase Inhibitor p19 , Enzyme Inhibitors , Gene Expression , Gene Expression Regulation, Enzymologic , Gene Transfer Techniques , Humans , Macrophages/metabolismABSTRACT
The progeny of individual macrophage precursors from mouse spleen were examined for their ability to constitutively process and present native pigeon cytochrome c or a peptide fragment of this antigen to naive CD4+ T-cells from mice transgenic for a V alpha 11/V beta 3 TCR that recognizes an epitope in the antigen fragment. The results show that constitutive Ag processing and presentation is a stable characteristic restricted to the progeny of approximately 20% of splenic macrophage precursors. This property does not appear to be randomly acquired, but to reflect the ability of certain macrophages to produce IL-12.
