Gfi-1 inhibits the expression of eosinophil major basic protein (MBP) during G-CSF-induced neutrophilic differentiation

Abstract

The zinc finger transcriptional repressor Gfi-1 has been shown to play a critical role in early granulopoiesis; however, its role in late neutrophilic development is poorly understood. We report here that forced expression of a dominant negative Gfi-1 mutant, N382S, resulted in augmented mRNA levels of eosinophil major basic protein (MBP) in myeloid cells induced with G-CSF to undergo terminal neutrophilic differentiation. MBP is a cytotoxic protein that is abundantly expressed in eosinophils, but not in neutrophils. Ectopic expression of MBP inhibited the proliferation and survival of differentiating myeloid cells in response to G-CSF. Significantly, while GFI-1 is upregulated during neutrophilic differentiation, it is rapidly downregulated upon induction of eosinophilic differentiation, which was associated with increased MBP expression. Knockdown of GFI-1 in eosinophilic cells also led to increased level of MBP mRNA. These results indicate that Gfi-1 functions to inhibit the expression of MBP and aberrant expression of MBP as a result of loss of Gfi-1 function may cause premature apoptosis of differentiating neutrophils. In contrast, the rapid downregulation of Gfi-1 during eosinophilic development may allow for abundant expression of MBP, a hallmark of eosinophilic differentiation.

Transcriptional Heterogeneity and Lineage Commitment in Myeloid Progenitors

“…Irf8 (Kurotaki et al, 2014;Wang et al, 2014) was highly enriched in monocyte clusters (C14 and C15 in addition to specific enrichment in C11 as discussed below). Cebpe and Gfi1 were enriched in the neutrophil and eosinophil clusters (C16-C17 and C18), consistent with their known role in neutrophil differentiation and repression of the basophil lineage (Liu and Dong, 2012). Lmo4 and Runx1 were enriched in the basophil clusters (C12 and C13), suggesting that these TFs are major drivers of this lineage.…”

The Ontogeny of a Neutrophil: Mechanisms of Granulopoiesis and Homeostasis

“…By negatively regulating the expression of c-Myc, a helix-loop-helix leucine zipper protein that locates to the promoters of certain genes with important regulatory control of the cell cycle, C/EBP-␣ induces early myeloid precursors to enter CMP differentiation pathways (55,58). Whereas C/EBP-␣, PU.1, and Irf8 induce CMPs to differentiate into monocytes and macrophages, C/EBPand Gfi-1 generate neutrophils and eosinophils (46,48,59,60). It is the acetylation of C/EBP-at specific lysines (K121 and K198) and the lack of expression of GATA-1, however, that cause early CMPs to ultimately differentiate into neutrophils instead of eosinophils (48,54,61).…”

“…In murine models, fetal knockout of Runx1 results in the loss of myeloid hematopoiesis (94), while the deletion of this gene in adult mice leads to primary monopoiesis at the expense of granulopoiesis (95). Gfi-1 deletion blocks neutrophil maturation at the promyelocyte stage, with cells demonstrating developmental failures of gelatinase granules and secretory vesicles (59,90,96). C/EBP-controls the promyelocyte-to-myelocyte transition in granulocytes and is essential for the formation of specific and gelatinase granules (97,98).…”

Gfi-1 is the transcriptional repressor ofSOCS1in acute myeloid leukemia cells

“…To knock down the Gfi-1 in U937 cells, shRNA against human Gfi-1 (clone TRCN0000020467; Open Biosystems, Huntsville, AL, USA) [19] was used. Lentiviral particles were produced by cotransfecting 293T cells with the shRNA construct along with the packaging plasmids (Delta 8.9 and vesicular stomatitis virus G glycoprotein), using Arrest-In transfection reagent (Open Biosystems).…”