Synergistic induction of intercellular adhesion molecule-1 by the human cytomegalovirus transactivators IE2p86 and pp71 is mediated via an Sp1-binding site

Abstract

Human cytomegalovirus (HCMV) infection of transplant recipients is frequently associated with allograft vasculopathy and rejection. One potential mechanism is vascular injury from HCMVtriggered, immunologically mediated processes. HCMV infection has been shown to increase the expression of intercellular adhesion molecule-1 (ICAM-1). The objective of this study was to determine the molecular basis of HCMV-enhanced ICAM-1 gene expression. Transient transfection experiments identified the IE2p86 protein as a potent activator of the ICAM-1 promoter. The tegument protein pp71 showed a strong synergistic effect on IE2p86-mediated ICAM-1 promoter activation. Mutagenesis experiments defined a DNA element from 2110 to 242 relative to the transcription start site as responsive for IE2p86. Further point mutations within this DNA element identified an Sp1-binding site that was essential for strong synergistic activation by IE2p86 and pp71. To confirm the activation of ICAM-1 gene expression, human fibroblasts (HFF) as well as endothelial cells (HUVEC) were infected with recombinant IE2p86-and pp71-expressing baculoviruses, respectively. In FACS analysis, a synergistic induction of ICAM-1 was detectable when cells were co-infected with the two recombinant baculoviruses. These findings clearly demonstrate that IE2p86 and pp71 are crucial regulatory factors for HCMV-induced ICAM-1 upregulation. INTRODUCTIONHuman cytomegalovirus (HCMV), a b-herpesvirus, is a ubiquitous human pathogen with a prevalence of infection in the adult population of 50-90 %. Although HCMV rarely causes severe disease on primary infection in immunocompetent people, it represents a highly pathogenic agent for immunosuppressed patients such as transplant recipients, as well as newborns, and establishes lifelong persistence in the infected host (Alford & Britt, 1990). So far, the exact site of latency is still uncertain; monocytes, haematopoietic precursor cells and endothelial cells have been discussed as potential reservoirs for latent HCMV (Fish et al., 1995;Sindre et al., 1996).Recent evidence suggests that HCMV not only causes acute infection in transplanted patients but may also be associated with extensive transplant vasculopathy, ultimately leading to chronic allograft rejection (Grattan et al., 1989;Skowronski et al., 1993;Koskinen et al., 1993;Lautenschlager et al., 1997a;Schnitzler et al., 1997;Borchers et al., 1999). One of the molecular mechanisms underlying the acceleration of vascular disease processes by HCMV infection may be enhancement of transplant immunogenicity. Several studies indicate that this involves the HCMV-induced upregulation of cellular adhesion molecules, which augments the adherance and infiltration of inflammatory cells that are capable of promoting vascular disease (Einsele et al., 1994;Lemstrom et al., 1995;Steinhoff et al., 1995;Koskinen et al., 1996;Yilmaz et al., 1996; Martelius et al., 1998;Waldman et al., 1998;Lautenschlager et al., 1999;The et al., 2001). For instance, cell culture experiments have demonstrated that...

NF-κB regulates thrombin-induced ICAM-1 gene expression in cooperation with NFAT by binding to the intronic NF-κB site in the ICAM-1 gene

“…These results further suggest that both NF-B and NFAT signaling contribute to thrombininduced ICAM-1 expression in endothelial cells. Our findings add to a growing body of evidence suggesting that inducible transcription factors interact with the intronic enhancer DNA elements and play critical roles in gene transcription (15,18,31). Another study showed that MHC class I-related neonatal Fc receptor for IgG induced by TNF-␣ is associated with NF-B binding to the intronic sequence (18).…”

“…A number of recent studies have shown that inducible transcription factors binding to intronic enhancer elements play critical role in gene transcription (15,18,31). There is, however, no evidence to indicate whether the inducible transcrip-tion factors interact with an intronic region of the ICAM-1 gene and contribute to the transcription of ICAM-1 gene.…”

Human Cytomegalovirus Tegument Protein pp71 Directs Long-Term Gene Expression from Quiescent Herpes Simplex Virus Genomes

“…Dependence on ATF recognition sites was also observed in cotransfection studies that investigated the response of the HCMV US11 promoter in the presence of pp71, although in this case the IE protein IE86 was also present (7). The promoter for intercellular adhesion molecule 1 was activated by pp71, in conjunction with IE86, via Sp1 recognition sites (32). Limited stimulation of endogenous intercellular adhesion molecule 1 expression by pp71 alone was also reported in these studies.…”

Human Cytomegalovirus Protein pp71 Disrupts Major Histocompatibility Complex Class I Cell Surface Expression

“…Similar to many herpesvirus gene products that have been investigated, pp71 exerts multiple roles during the cytomegalovirus life cycle. It is a transactivator of both viral and cellular genes and can synergize with the transactivating activities of IE2p86 and ppUL35 (6,17,27,29,42). pp71 also increases the infectivity of viral DNA in a manner independent from its function as a transactivator of the IE1 and IE2 genes (3).…”