title: Regulation of neutrophil and eosinophil secondary granule gene expression by transcription factors C/EBP epsilon and PU.1.
abstract: In the bone marrow of C/EBP epsilon(-/-) mice, expression of neutrophil secondary and tertiary granule mRNAs is absent for lactoferrin (LF), neutrophil gelatinase (NG), murine cathelin-like protein (MCLP), and the cathelin B9; it is severely reduced for neutrophil collagenase (NC) and neutrophil gelatinase-associated lipocalin (NGAL). In addition, the expression of eosinophil granule genes, major basic protein (MBP), and eosinophil peroxidase (EPX) is absent. These mice express C/EBP alpha, C/EBP beta, and C/EBP delta in the bone marrow at levels similar to those of their wild-type counterparts, suggesting a lack of functional redundancy among the family in vivo. Stable inducible expression of C/EBP epsilon and C/EBP alpha in the murine fibroblast cell line NIH 3T3 activated expression of mRNAs for B9, MCLP, NC, and NGAL but not for LF. In transient transfections of C/EBP epsilon and C/EBP alpha, B9 was strongly induced with weaker induction of the other genes. C/EBP beta and C/EBP delta proteins weakly induced B9 expression, but C/EBP delta induced NC expression more efficiently than the other C/EBPs. The expression of MBP was inefficiently induced by C/EBP epsilon alone and weakly induced with C/EBP epsilon and GATA-1, but the addition of PU.1 resulted in a striking cooperative induction of MBP in NIH 3T3 cells. Mutation of a predicted PU.1 site in the human MBP promoter-luciferase reporter construct abrogated the response to PU.1. Gel-shift analysis demonstrated binding of PU.1 to this site. MBP and EPX mRNAs were absent in a PU.1-null myeloid cell line established from the embryonic liver of PU.1(-/-) mice. Restitution of PU.1 protein expression restored MBP and EPX protein expression. This study demonstrates that C/EBP epsilon is essential and sufficient for the expression of a particular subset of neutrophil secondary granule genes. Furthermore, it indicates the importance of PU.1 in the cooperative activation of eosinophil granule genes.
authors.affiliation: Division of Hematology/Oncology, Cedars-Sinai Medical Center, Los Angeles, CA 90048, USA. gombarta@csmc.edu
authors.forename: Adrian F
authors.initials: AF
authors.lastname: Gombart
authors.forename: Scott H
authors.initials: SH
authors.lastname: Kwok
authors.forename: Karen L
authors.initials: KL
authors.lastname: Anderson
authors.forename: Yuji
authors.initials: Y
authors.lastname: Yamaguchi
authors.forename: Bruce E
authors.initials: BE
authors.lastname: Torbett
authors.forename: H Phillip
authors.initials: HP
authors.lastname: Koeffler
mesh_terms: D016475:3T3 Cells / Q000378:metabolism; D000209:Acute-Phase Proteins; D000818:Animals; D001798:Blood Proteins / Q000096:biosynthesis / Q000235:genetics; D022763:CCAAT-Enhancer-Binding Protein-alpha / Q000378:metabolism; D022782:CCAAT-Enhancer-Binding Protein-beta / Q000378:metabolism; D051743:CCAAT-Enhancer-Binding Protein-delta; D022762:CCAAT-Enhancer-Binding Proteins / Q000172:deficiency / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D002352:Carrier Proteins / Q000096:biosynthesis / Q000235:genetics; D017364:Collagenases / Q000096:biosynthesis / Q000235:genetics; D003594:Cytoplasmic Granules / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000502:physiology; D047091:Eosinophil Granule Proteins; D047209:Eosinophil Peroxidase; D004804:Eosinophils / Q000502:physiology*; D050981:Erythroid-Specific DNA-Binding Factors; D050982:GATA1 Transcription Factor; D018093:Gelatinases / Q000096:biosynthesis / Q000235:genetics; D005786:Gene Expression Regulation / Q000502:physiology*; D006801:Humans; D007781:Lactoferrin / Q000096:biosynthesis / Q000235:genetics; D000071068:Lipocalin-2; D054834:Lipocalins; D051379:Mice; D018345:Mice, Knockout; D009504:Neutrophils / Q000201:enzymology / Q000502:physiology*; D015513:Oncogene Proteins; D010544:Peroxidases / Q000096:biosynthesis / Q000235:genetics; D011518:Proto-Oncogene Proteins / Q000235:genetics / Q000502:physiology*; D012333:RNA, Messenger / Q000096:biosynthesis; D011993:Recombinant Fusion Proteins / Q000502:physiology; D012045:Regulatory Sequences, Nucleic Acid; D012260:Ribonucleases; D015534:Trans-Activators / Q000235:genetics / Q000502:physiology*; D014157:Transcription Factors / Q000502:physiology; D014158:Transcription, Genetic; D000099232:Proto-Oncogene Protein Spi-1
chemical_list: D000209:Acute-Phase Proteins; D001798:Blood Proteins; D022763:CCAAT-Enhancer-Binding Protein-alpha; D022782:CCAAT-Enhancer-Binding Protein-beta; D051743:CCAAT-Enhancer-Binding Protein-delta; D022762:CCAAT-Enhancer-Binding Proteins; D002352:Carrier Proteins; D017364:Collagenases; D004268:DNA-Binding Proteins; D047091:Eosinophil Granule Proteins; D047209:Eosinophil Peroxidase; D050981:Erythroid-Specific DNA-Binding Factors; D050982:GATA1 Transcription Factor; D018093:Gelatinases; D007781:Lactoferrin; D000071068:Lipocalin-2; D054834:Lipocalins; D015513:Oncogene Proteins; D010544:Peroxidases; D011518:Proto-Oncogene Proteins; D012333:RNA, Messenger; D011993:Recombinant Fusion Proteins; D012260:Ribonucleases; D015534:Trans-Activators; D014157:Transcription Factors; D000099232:Proto-Oncogene Protein Spi-1; C497252:CEBPD protein, human; C497253:Cebpd protein, mouse; C489468:Cebpe protein, mouse; C494688:GATA1 protein, human; C494689:Gata1 protein, mouse; C489506:LCN2 protein, human; C070790:CEBPE protein, human