The interferon-γ-mediated inhibition of lipoprotein lipase gene transcription in macrophages involves casein kinase 2- and phosphoinositide-3-kinase-mediated regulation of transcription factors Sp1 and Sp3

Abstract

The mechanisms underlying transcriptional inhibition by interferon-γ (IFN-γ) are poorly understood despite the existence of a large number of genes that are regulated in this manner and the key role of this cytokine in inflammatory disorders such as atherosclerosis. We have previously identified a novel mechanism for transcriptional inhibition by IFN-γ that involves a reduction in the binding of transcription factors Sp1 and Sp3 to regulatory sequences in the lipoprotein lipase (LPL) gene. In the present study, we have investigated the signalling pathways that impact on the IFN-γ-mediated regulation of Sp1/Sp3 binding and LPL gene transcription in macrophages. The IFN-γ-mediated inhibition of LPL promoter activity was prevented by expression of dominant negative forms of casein kinase 2 (CK2) and protein kinase B (PKB), a key downstream component of the phosphoinositide-3-kinase (PI3K) pathway. IFN-γ activated both the catalytic subunits of CK2 without affecting their expression. CK2 interacted with both Sp1 and Sp3 and this association was increased by IFN-γ. Electrophoretic mobility shift assays showed that a CK2-mediated phosphorylation of either cellular extracts or recombinant Sp1 reduced binding to the regulatory region in the LPL gene. The action of PKB was potentially mediated through mammalian target for rapamycin proteins. Taken together, these results suggest a key role for CK2 and PI3K signalling pathways in the IFN-γ-mediated inhibition of macrophage LPL gene transcription through the regulation of Sp1/Sp3 binding.

Deciphering the modulation of gene expression by type I and II interferons combining 4sU-tagging, translational arrest and in silico promoter analysis

“…For an individual IRepG (‘lipoprotein lipase’), negative regulation by SP1 and SP3 has been described (55). Our data imply that such regulation is more widespread than commonly accepted and thus deserves a thorough investigation concerning the underlying receptor-proximal events and contribution to antiviral, anti-pathogenic and anti-tumorigenic IFNγ functions.…”

The Phosphoinositide 3‐Kinase Signaling Pathway is Involved in the Control of Modified Low‐Density Lipoprotein Uptake by Human Macrophages

“…The LPL enzyme expressed by macrophages has been shown to contribute to foam cell formation because of its ability to bind to both the cell surface and lipoproteins via different regions that cause the accumulation of the latter and their subsequent uptake by the cells (bridging action) [3, 4]. Interestingly, our previous studies had shown that PI3K is involved, at least in part, in the IFN-γ-mediated regulation of LPL expression [23] and this study extends this to its constitutive expression in human macrophages. In addition to receptor-mediated uptake of LDL, fluid-phase pinocytosis, particularly macropinocytosis, has recently been found to play an important role in the control of macrophage foam cell formation [5, 6].…”

ERK Is Integral to the IFN-γ–Mediated Activation of STAT1, the Expression of Key Genes Implicated in Atherosclerosis, and the Uptake of Modified Lipoproteins by Human Macrophages

“…Differentiation of THP-1 monocytes into macrophages was performed for 24 h using 0.16 mM PMA, unless otherwise stated. Treatment of macrophages with IFN-g or the pharmacological inhibitors was as previously described (4,5).…”

“…Several candidate kinases have been found to phosphorylate Ser 727 (1,3,4), although the exact roles of these remain poorly understood, particularly in macrophages in relation to atherosclerosis. Our previous studies have provided novel insights into IFN-g signaling and revealed critical roles for casein kinase 2 (CK2) and PI3K in the IFN-g-regulated expression in mouse macrophages of several key genes implicated in atherosclerosis (4,5). The action of both of these kinases was mediated, at least in part, through the regulation of STAT1 Ser 727 phosphorylation (4).…”