The homeobox transcription factor VentX controls human macrophage terminal differentiation and proinflammatory activation

Abstract

Macrophages are critical players in both innate and adaptive immunity. While the exogenous signaling events leading to the terminal differentiation of macrophages from monocytes have been studied extensively, the underlying intracellular transcriptional mechanisms remain poorly understood. Here we report that the homeobox transcription factor VentX plays a pivotal role in human macrophage terminal differentiation and proinflammatory function. Our study showed that VentX expression was upregulated upon human primary monocyte-to-macrophage differentiation induced by cytokines such as M-CSF, GM-CSF, and IL-3. Moreover, ablation of VentX expression in primary monocytes profoundly impaired their differentiation to macrophages, and ectopic expression of VentX in a myeloid progenitor cell line triggered its differentiation with prominent macrophage features. Further analysis revealed that VentX was pivotal for the proinflammatory response of terminally differentiated macrophages. Mechanistically, VentX was found to control expression of proteins key to macrophage differentiation and activation, including M-CSF receptor. Importantly, preliminary analysis of gene expression in leukocytes from patients with autoimmune diseases revealed a strong correlation between levels of VentX and those of proinflammatory cytokines. Our results provide mechanistic insight into the crucial roles of VentX in macrophage differentiation and proinflammatory activation and suggest that dysregulation of VentX may play a role in the pathogenesis of autoimmune diseases.

Homeobox Transcription Factor VentX Regulates Differentiation and Maturation of Human Dendritic Cells

“…All the MO oligonucleotides were ordered from Gene Tools (Philomath, OR). LPDCs were transfected with siRNA targeting VentX as described in our previous study (17). Knockdown of IB␣ expression in THP1 cells was achieved through electroporation transfection of SignalSilence IB␣ siRNA I from Cell Signaling (Danvers, MA).…”

“…The doxycycline inducible retroviruses expressing GFP.VentX or GFP have been described in our previous study (17). The THP1 cell line conditionally expressing GFP.VentX was generated through co-transduction of pRetroX-GFP.VentX and pRetroX-Tet-On Advanced retroviruses and GFP.VentX-positive cells were sorted by FACSAria high-speed sorter (BD Bioscience, San Jose, CA) after incubation with 1.0 g/ml doxycycline for 24 h (Dana-Farber Cancer Institute Flow Cytometry Core Facility).…”

“…Real time PCR was performed on a LightCycler (480 Real-Time PCR System; Roche). The primers used for the detection of mRNA levels of VentX, IL6, IL12, p21, c-myc, CCL2, CCL3, CCL5, CCL19, CXCL1, CXCL12, CD1a, TNF␣, TLR2, and TLR4 have been shown previously (17). Western blotting analysis was conducted as described in our prior study (17).…”

“…VentX is a human homologue of the Xenopus homeobox gene Xom of the BMP4 signaling pathway, and has been defined as a novel hematopoietic transcriptional factor controlling proliferation and differentiation of hematopoietic and immune cells (15)(16)(17). Initially identified as a novel LEF/TCF associated antagonist of the canonical Wnt signaling through reverse genetic modeling of Xenopus embryogenesis, VentX was found to be a transcriptional activator of the p53/p21 and p16 ink4a /Rb tumor suppression pathways (18,19).…”

“…Initially identified as a novel LEF/TCF associated antagonist of the canonical Wnt signaling through reverse genetic modeling of Xenopus embryogenesis, VentX was found to be a transcriptional activator of the p53/p21 and p16 ink4a /Rb tumor suppression pathways (18,19). The critical role of VentX in hematopoietic cells development was indicated by its function in controlling the proliferation and differentiation of CD34 ϩ cells and monocyte to macrophage terminal differentiation (16,17).…”