Macrophage Activation and Differentiation Signals Regulate Schlafen-4 Gene Expression: Evidence for Schlafen-4 as a Modulator of Myelopoiesis

Abstract

BackgroundThe ten mouse and six human members of the Schlafen (Slfn) gene family all contain an AAA domain. Little is known of their function, but previous studies suggest roles in immune cell development. In this report, we assessed Slfn regulation and function in macrophages, which are key cellular regulators of innate immunity.Methodology/Principal FindingsMultiple members of the Slfn family were up-regulated in mouse bone marrow-derived macrophages (BMM) by the Toll-like Receptor (TLR)4 agonist lipopolysaccharide (LPS), the TLR3 agonist Poly(I∶C), and in disease-affected joints in the collagen-induced model of rheumatoid arthritis. Of these, the most inducible was Slfn4. TLR agonists that signal exclusively through the MyD88 adaptor protein had more modest effects on Slfn4 mRNA levels, thus implicating MyD88-independent signalling and autocrine interferon (IFN)-β in inducible expression. This was supported by the substantial reduction in basal and LPS-induced Slfn4 mRNA expression in IFNAR-1−/− BMM. LPS causes growth arrest in macrophages, and other Slfn family genes have been implicated in growth control. Slfn4 mRNA levels were repressed during macrophage colony-stimulating factor (CSF-1)-mediated differentiation of bone marrow progenitors into BMM. To determine the role of Slfn4 in vivo, we over-expressed the gene specifically in macrophages in mice using a csf1r promoter-driven binary expression system. Transgenic over-expression of Slfn4 in myeloid cells did not alter macrophage colony formation or proliferation in vitro. Monocyte numbers, as well as inflammatory macrophages recruited to the peritoneal cavity, were reduced in transgenic mice that specifically over-expressed Slfn4, while macrophage numbers and hematopoietic activity were increased in the livers and spleens.Conclusions Slfn4 mRNA levels were up-regulated during macrophage activation but down-regulated during differentiation. Constitutive Slfn4 expression in the myeloid lineage in vivo perturbs myelopoiesis. We hypothesise that the down-regulation of Slfn4 gene expression during macrophage differentiation is a necessary step in development of this lineage.

The DNA-binding factor Ctcf critically controls gene expression in macrophages

“…Since we showed that the deletion of Ctcf does not significantly affect the expression levels of TLR4, TLR8 and TLR9 mRNA, and since these genes are not direct targets of Ctcf, the observed effect may have been indirectly caused through type I IFN. 42 Regulation of TNF expression by Ctcf is in agreement with the literature. 43 Additionally, we show diminished IL-10 production by macrophages as a result of Ctcf depletion, similarly to low IL-10 production upon Ctcf deletion in Th2 cells as we previously observed.…”

“…A set of genes downregulated during macrophage differentiation but induced upon TLR stimulation are the Schlafen (Slfn) genes, especially Slfn4. 42 In our data set, the expression levels of Slfn1, Slfn4 and Slfn9 genes were lower after LPS stimulation in LysM-Cre Ctcf f/f than in control macrophages (ratio: 0.21, 0.38 and 0.38, respectively). Since we showed that the deletion of Ctcf does not significantly affect the expression levels of TLR4, TLR8 and TLR9 mRNA, and since these genes are not direct targets of Ctcf, the observed effect may have been indirectly caused through type I IFN.…”

Expression and Regulatory Effects of Murine Schlafen (Slfn) Genes in Malignant Melanoma and Renal Cell Carcinoma

“…Several mouse Slfn genes, including Slfn1 (4), Slfn 2 (9), and Slfn3 (13, 14, 16) have been shown to play key regula- tory roles in control of cell proliferation and/or differentiation. Mouse Slfn2 has been implicated in immune cell quiescence (10), whereas Slfn4 has been implicated in the control of myelopoiesis (17).…”

“…The only evidence for regulatory effects of Slfn3 on cell growth stems from prior work, in which ectopic expression of this Slfn member in human colon cancer cells inhibited cellular growth (16). The function of Slfn4 is not well known at this time, as there is only one report showing its involvement in macrophage activation and differentiation (17). Similarly, the functions of the remaining members of the Slfn family are largely unknown.…”

SLFN14 mutations underlie thrombocytopenia with excessive bleeding and platelet secretion defects

“…Her mother (I:2) had a platelet count in whole blood of 83 × 10 9 /l, with a less severe bleeding history. The platelet count in patient cell-cycle regulation, proliferation, and differentiation (10)(11)(12)(13)(14). Recently, data have been published suggesting an important function for SLFN14 as an endoribonuclease, regulating rRNA and ribosome-associated mRNA cleavage and translational control in rabbit reticulocytes (15).…”