PU.1 (Spi-1) and C/EBP alpha regulate expression of the granulocyte-macrophage colony-stimulating factor receptor alpha gene

Mol Cell Biol. 1995 Oct;15(10):5830-45. doi: 10.1128/MCB.15.10.5830.


Growth factor receptors play an important role in hematopoiesis. In order to further understand the mechanisms directing the expression of these key regulators of hematopoiesis, we initiated a study investigating the transcription factors activating the expression of the granulocyte-macrophage colony-stimulating factor (GM-CSF) receptor alpha gene. Here, we demonstrate that the human GM-CSF receptor alpha promoter directs reporter gene activity in a tissue-specific fashion in myelomonocytic cells, which correlates with its expression pattern as analyzed by reverse transcription PCR. The GM-CSF receptor alpha promoter contains an important functional site between positions -53 and -41 as identified by deletion analysis of reporter constructs. We show that the myeloid and B cell transcription factor PU.1 binds specifically to this site. Furthermore, we demonstrate that a CCAAT site located upstream of the PU.1 site between positions -70 and -54 is involved in positive-negative regulation of the GM-CSF receptor alpha promoter activity. C/EBP alpha is the major CCAAT/enhancer-binding protein (C/EBP) form binding to this site in nuclear extracts of U937 cells. Point mutations of either the PU.1 site or the C/EBP site that abolish the binding of the respective factors result in a significant decrease of GM-CSF receptor alpha promoter activity in myelomonocytic cells only. Furthermore, we demonstrate that in myeloid and B cell extracts, PU.1 forms a novel, specific, more slowly migrating complex (PU-SF) when binding the GM-CSF receptor alpha promoter PU.1 site. This is the first demonstration of a specific interaction with PU.1 on a myeloid PU.1 binding site. The novel complex is distinct from that described previously as binding to B cell enhancer sites and can be formed by addition of PU.1 to extracts from certain nonmyeloid cell types which do not express PU.1, including T cells and epithelial cells, but not from erythroid cells. Furthermore, we demonstrate that the PU-SF complex binds to PU.1 sites found on a number of myeloid promoters, and its formation requires an intact PU.1 site adjacent to a single-stranded region. Expression of PU.1 in nonmyeloid cells can activate the GM-CSF receptor alpha promoter. Deletion of the amino-terminal region of PU.1 results in a failure to form the PU-SF complex and in a concomitant loss of transactivation, suggesting that formation of the PU-SF complex is of functional importance for the activity of the GM-CSF receptor alpha promoter. Finally, we demonstrate that C/EBP alpha can also active the GM-CSF receptor alpha promoter in nonmyeloid cells. These results suggest that PU.1 and C/EBP alpha direct the cell-type-specific expression of GM-CSF receptor alpha, further establish the role of PU.1 as a key regulator of hematopoiesis, and point to C/EBP alpha as an additional important factor in this process.

Publication types

  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Amino Acid Sequence
  • B-Lymphocytes
  • Base Sequence
  • CCAAT-Enhancer-Binding Proteins
  • Cell Line
  • DNA / metabolism
  • DNA, Single-Stranded / metabolism
  • DNA-Binding Proteins / metabolism*
  • Genes, Reporter
  • Hematopoiesis / genetics
  • Humans
  • Methylation
  • Molecular Sequence Data
  • Monocytes
  • Mutation
  • Nuclear Proteins / metabolism*
  • Promoter Regions, Genetic / genetics
  • Receptors, Granulocyte-Macrophage Colony-Stimulating Factor / genetics*
  • Retroviridae Proteins, Oncogenic
  • T-Lymphocytes
  • Transcription Factors / metabolism*
  • Transcriptional Activation / genetics*


  • CCAAT-Enhancer-Binding Proteins
  • DNA, Single-Stranded
  • DNA-Binding Proteins
  • Nuclear Proteins
  • Receptors, Granulocyte-Macrophage Colony-Stimulating Factor
  • Retroviridae Proteins, Oncogenic
  • Transcription Factors
  • v-Spi-1 protein, Friend spleen focus-forming virus
  • DNA