Commitment to the Monocytic Lineage Occurs in the Absence of the Transcription Factor PU.1

Abstract

Mice homozygous for the disruption of the PU.1 (Spi-1) gene do not produce mature macrophages. In determining the role of PU.1 in macrophage differentiation, the present study investigated whether or not there was commitment to the monocytic lineage in the absence of PU.1. Early PU.1−/− myeloid colonies were generated from neonate liver under conditions that promote primarily macrophage and granulocyte/macrophage colonies. These PU.1−/− colonies were found to contain cells with monocytic characteristics as determined by nonspecific esterase stain and the use of monoclonal antibodies that recognize early monocyte precursors, including Moma-2, ER-MP12, ER-MP20, and ER-MP58. In addition, early myeloid cells could be grown from PU.1−/− fetal liver cultures in the presence of granulocyte-macrophage colony-stimulating factor (GM-CSF). Similar to the PU.1 null colonies, the GM-CSF–dependent cells also possessed early monocytic characteristics, including the ability to phagocytize latex beads. The ability of PU.1−/− progenitors to commit to the monocytic lineage was also verified in vivo by flow cytometry and cytochemical analysis of primary neonate liver cells. The combined data shows that PU.1 is absolutely required for macrophage development after commitment to this lineage.

Lineage plasticity and commitment in T‐cell development

“…A recent report also shows that in appropriate cells GM‐CSF stimulation can induce PU.1 expression (93). Transcription of GM‐CSF‐Rα does not depend strictly on PU.1 per se (94), but it depends on a PU.1 binding site and is usually regulated by PU.1 in normal myeloid cells (95). Thus, these different manipulations might activate each other.…”

The transcription factor PU.1 does not regulate lineage commitment but has lineage-specific effects

“…Under these conditions normal neonate liver cells produced predominantly macrophage colonies and some mixed granulocyte/macrophage and neutrophil colonies. The mixed colonies also contained predominantly macrophages [31].…”

“…The earliest identified cell of this lineage is the monoblast, which gives rise to the promonocyte and finally to the monocyte. Because unique stage-specific markers are not available for monocytes, we decided on a combination of available antibodies for initial identification of monoblasts/ promonocytes [31]. Moma-2 is specific for monocyte/macrophage lineage cells, recognizing mature macrophage subsets and bone marrow precursors [32].…”

“…We were unable to grow early myeloid cells from neonate liver or spleen in either M-CSF or GM-CSF, nor from fetal liver in M-CSF. However, early myeloid cells grew from fetal liver cultured with GM-CSF as the only added growth factor [31].…”

Monocyte-Specific Knockout of C/ebpα Results in Osteopetrosis Phenotype, Blocks Bone Loss in Ovariectomized Mice, and Reveals an Important Function of C/ebpα in Osteoclast Differentiation and Function

“…Nonspecific esterase (NSE; Sigma; 91A-1KT) staining was performed as described. (2,23,24) For X-ray analysis, radiography was performed using a high-resolution, soft X-ray system (Faxitron Model MX-20) by the UAB Small Animal Bone Phenotyping Core associated with the Center for Metabolic Bone Disease.…”