Edmonston measles virus prevents increased cell surface expression of peptide-loaded major histocompatibility complex class II proteins in human peripheral monocytes

Gamma interferon (IFN-gamma) induces expression of the gene products of the major histocompatibility complex (MHC), whereas IFN-alpha/beta can interfere with or suppress class II protein expression. In separate studies, measles virus (MV) was reported to induce IFN-alpha/beta and to up-regulate MHC class II proteins. In an attempt to resolve this paradox, we examined the surface expression of MHC class I and class II proteins in MV-infected peripheral monocytes in the presence and absence of IFN-alpha/beta. Infection of purified monocytes with Edmonston B MV resulted in an apparent increase in cell surface expression of HLA-A, -B, and -C class I proteins, but it had no effect on the expression of HLA-DR class II proteins. MV-infected purified monocytes expressed IFN-alpha/beta, but no measurable IFN-gamma expression was detected in supernatant fluids. Class II protein expression could be enhanced by coculture of purified monocytes with uninfected peripheral blood mononuclear cell (PBMC) supernatant. MV infection of PBMCs also did not affect expression of class II proteins, but the expression of HLA-A, -B, and -C class I proteins was increased two- to threefold in most donor cells. A direct role for IFN-alpha/beta suppression of MHC class II protein expression was not evident in monocytes since MV suppressed class II protein expression in the absence of IFN-alpha/beta. Taken together, these data suggest that MV interferes with the expression of peptide-loaded class II complexes, an effect that may potentially alter CD4(+)-T-cell proliferation and the cell-mediated immune responses that they help to regulate.     

Patterns of constitutive and IFN-γ inducible expression of HLA class II molecules in human melanoma cell lines

Major histocompatibility complex (MHC) class II proteins (HLA-DR, HLA-DP and HLA-DQ) play a fundamental role in the regulation of the immune response. The level of expression of human leukocyte antigen (HLA) class II antigens is regulated by interferon-gamma (IFN-gamma) and depends on the status of class II trans-activator protein (CIITA), a co-activator of the MHC class II gene promoter. In this study, we measured levels of constitutive and IFN-gamma-induced expression of MHC class II molecules, analysed the expression of CIITA and investigated the association between MHC class II transactivator polymorphism and expression of different MHC class II molecules in a large panel of melanoma cell lines obtained from the European Searchable Tumour Cell Line Database. Many cell lines showed no constitutive expression of HLA-DP, HLA-DQ and HLA-DR and no IFN-gamma-induced increase in HLA class II surface expression. However, in some cases, IFN-gamma treatment led to enhanced surface expression of HLA-DP and HLA-DR. HLA-DQ was less frequently expressed under basal conditions and was less frequently induced by IFN-gamma. In these melanoma cell lines, constitutive surface expression of HLA-DR and HLA-DP was higher than that of HLA-DQ. In addition, high constitutive level of cell surface expression of HLA-DR was correlated with lower inducibility of this expression by IFN-gamma. Finally, substitution A-->G in the 5' flanking region of CIITA promoter type III was associated with higher expression of constitutive HLA-DR (p<0.005). This study yielded a panel of melanoma cell lines with different patterns of constitutive and IFN-gamma-induced expression of HLA class II that can be used in future studies of the mechanisms of regulation of HLA class II expression.     

Regulation of MHC class II expression and antigen processing in murine and human mesenchymal stromal cells by IFN-gamma, TGF-beta, and cell density

Mesenchymal stromal cells (MSC) possess immunosuppressive properties, yet when treated with IFN-gamma they acquire APC functions. To gain insight into MSC immune plasticity, we explored signaling pathways induced by IFN-gamma required for MHC class II (MHC II)-dependent Ag presentation. IFN-gamma-induced MHC II expression in mouse MSC was enhanced by high cell density or serum deprivation and suppressed by TGF-beta. This process was regulated by the activity of the type IV CIITA promoter independently of STAT1 activation and the induction of the IFN regulatory factor 1-dependent B7H1/PD-L1 encoding gene. The absence of direct correlation with the cell cycle suggested that cellular connectivity modulates IFN-gamma responsiveness for MHC II expression in mouse MSC. TGF-beta signaling in mouse MSC involved ALK5 and ALK1 TGF-beta RI, leading to the phosphorylation of Smad2/Smad3 and Smad1/Smad5/Smad8. An opposite effect was observed in human MSC where IFN-gamma-induced MHC II expression occurred at the highest levels in low-density cultures; however, TGF-beta reduced IFN-gamma-induced MHC II expression and its signaling was similar as in mouse MSC. This suggests that the IFN-gamma-induced APC features of MSC can be modulated by TGF-beta, serum factors, and cell density in vitro, although not in the same way in mouse and human MSC, via their convergent effects on CIITA expression.     

Inhibition of IFN-gamma-induced class II transactivator expression by a 19-kDa lipoprotein from Mycobacterium tuberculosis: a potential mechanism for immune evasion

Mycobacterium tuberculosis (MTB) persists inside macrophages despite vigorous immune responses. MTB and MTB 19-kDa lipoprotein inhibit class II MHC (MHC-II) expression and Ag processing by a Toll-like receptor 2-dependent mechanism that is shown in this study to involve a defect in IFN-gamma induction of class II transactivator (CIITA). Exposure of macrophages to MTB or MTB 19-kDa lipoprotein inhibited IFN-gamma-induced MHC-II expression, but not IL-4-induced MHC-II expression, by preventing induction of mRNA for CIITA (total, type I, and type IV), IFN regulatory factor-1, and MHC-II. MTB 19-kDa lipoprotein induced mRNA for suppressor of cytokine signaling (SOCS)1 but did not inhibit IFN-gamma-induced Stat1 phosphorylation. Furthermore, the lipoprotein inhibited MHC-II Ag processing in SOCS1(-/-) macrophages. MTB 19-kDa lipoprotein did not inhibit translocation of phosphorylated Stat1 to the nucleus or Stat1 binding to and transactivation of IFN-gamma-sensitive promoter constructs. Thus, MTB 19-kDa lipoprotein inhibited IFN-gamma signaling independent of SOCS1 and without interfering with the activation of Stat1. Inhibition of IFN-gamma-induced CIITA by MTB 19-kDa lipoprotein may allow MTB to evade detection by CD4(+) T cells.     

A positive regulatory role for suppressor of cytokine signaling 1 in IFN-gamma-induced MHC class II expression in fibroblasts

Suppressor of cytokine signaling 1 (SOCS1) is rapidly induced following stimulation by several cytokines. SOCS1 negatively regulates cytokine receptor signal transduction by inhibiting Janus family tyrosine kinases. Lack of such feedback regulation underlies the premature death of SOCS1(-/-) mice due to unbridled IFN-gamma signaling. We used mouse embryo fibroblasts derived from SOCS1(-/-) mice to investigate the role of SOCS1 in IFN-gamma signaling pathways. SOCS1(-/-) fibroblasts were exquisitely sensitive to the IFN-gamma-mediated growth arrest and showed sustained STAT1 phosphorylation. However, SOCS1(-/-) fibroblasts were inefficient in MHC class II surface expression following IFN-gamma stimulation, despite a marked induction of the MHC class II transactivator and MHC class II gene expression. Retroviral transduction of wild-type SOCS1 relieved the growth-inhibitory effects of IFN-gamma in SOCS1(-/-) fibroblasts by inhibiting STAT1 activation. SOCS1R105K, carrying a mutation within the phosphotyrosine-binding pocket of the Src homology 2 domain, did not inhibit STAT1 phosphorylation, yet considerably inhibited IFN-gamma-mediated growth arrest. Strikingly, expression of SOCS1R105K restored the IFN-gamma-induced MHC class II expression in SOCS1(-/-) cells, indicating that expression of SOCS1 facilitates MHC class II expression in fibroblasts. Our results show that SOCS1, in addition to its negative regulatory role of inhibiting Janus kinases, has an unanticipated positive regulatory function in retarding the degradation of IFN-gamma-induced MHC class II proteins in fibroblasts.     

Regulation of MHC class II antigen expression. Opposing effects of tumor necrosis factor-alpha on IFN-gamma-induced HLA-DR and Ia expression depends on the maturation and differentiation stage of the cell

MHC class II induction by cytokines has been suggested to play a major role in the initiation and propagation of immune and autoimmune processes. TNF-alpha has been found both to enhance and also to inhibit IFN-gamma-induced MHC class II expression. In the present studies, the effect of TNF-alpha on IFN-gamma induced MHC class II expression was tested in various cell lines. On the basis of the data, we propose that, depending on the stage of differentiation and maturation of the cells, TNF-alpha might synergize or antagonize the affects of IFN-gamma on the regulation of MHC class II expression. Thus, in immature cells such as HL-60 or THP-1, TNF-alpha enhances IFN-gamma-induced class II expression. However, when differentiation was induced in these cells by TPA or IFN-gamma, the additive effect of TNF-alpha on the IFN-gamma induced DR expression was eliminated. Furthermore, TNF-alpha down-regulates the IFN-gamma-induced class II expression in differentiated cells such as human skin fibroblasts or activated macrophages. In bone marrow cells induced to differentiate in vitro, TNF-alpha decreased the IFN-gamma-induced MHC class II expression in a maturation-dependent fashion. These results provide a rational explanation for the conflicting reports regarding the effect of TNF-alpha on IFN-gamma-induced class II expression. But more importantly they may be relevant to the biologic function of TNF-alpha. Thus, we show that TNF-alpha-treated mice have reduced level of Ia expression on peritoneal macrophages and in vivo treatment with TNF-alpha antagonizes the ability of IFN-gamma to induce class II expression on these macrophages.     

Interferon-gamma-induced astrocyte class II major histocompatibility complex gene expression is associated with both protein kinase C activation and Na+ entry

Astrocytes can be induced by interferon-gamma (IFN-gamma) to express class II major histocompatibility complex (MHC) antigens. This study was undertaken to elucidate the intracellular signaling pathways involved in IFN-gamma induction of class II MHC. We examined the effects of Na+/H+ antiporter and protein kinase C (PKC) inhibitors on class II expression and Na+ influx in astrocytes. We found that amiloride and ethyl isopropylamiloride, inhibitors of Na+/H+ exchange, blocked IFN-gamma-induced class II gene expression. IFN-gamma stimulated Na+ influx, and this increased influx was inhibited by amiloride. Treatment of astrocytes with the PKC inhibitor H7 also blocked the increase in Na+ uptake induced by IFN-gamma, indicating that IFN-gamma-induced PKC activation is required for subsequent Na+ influx. IFN-gamma treatment produced an increase of total PKC activity, which was associated with a rapid translocation of PKC activity from cytosolic to particulate fraction. H7 and another PKC inhibitor, staurosporine, inhibited IFN-gamma-induced class II gene expression. However, 4 beta-phorbol 12 beta-myristate 13 alpha-acetate, a potent PKC activator, did not affect class II expression. Taken together, our data indicate that both IFN-gamma-induced PKC activation and Na+ influx are required for class II MHC expression in astrocytes but that activation of PKC alone is not sufficient for ultimate expression of this gene.     

Attenuation of MHC class II expression in macrophages infected with Mycobacterium bovis bacillus Calmette-Guérin involves class II transactivator and depends on the Nramp1 gene.

The natural resistance associated macrophage protein 1 (Nramp1) gene determines the ability of murine macrophages to control infection with a group of intracellular pathogens, including Salmonella typhimurium, Leishmania donovani, and Mycobacterium bovis bacillus Calmette-Guérin (BCG). The expression of the resistant allele of the Nramp1 gene in murine macrophages is associated with a more efficient expression of several macrophage activation-associated genes, including class II MHC loci. In this study, we investigated the molecular mechanisms involved in IFN-gamma-induced MHC class II expression in three types of macrophages: those expressing a wild-type allele of the Nramp1 gene (B10R and 129/Mphi), those carrying a susceptible form of the Nramp1 gene (B10S), and those derived from 129-Nramp1-knockout mice (129/Nramp1-KO). Previously, we published results showing that Ia protein expression is significantly higher in the IFN-gamma-induced B10R macrophages, compared with its susceptible counterpart. In this paper, we also show that the higher expression of Ia protein in B10R cells is associated with higher I-Abeta mRNA expression, which correlates with a higher level of IFN-gamma-induced phosphorylation of the STAT1-alpha protein and subsequently with elevated expression of class II transactivator (CIITA) mRNA, compared with B10S. Furthermore, we demonstrate that the infection of macrophages with M. bovis BCG results in a down-regulation of CIITA mRNA expression and, consequently, in the inhibition of Ia induction. Therefore, our data explain, at least in part, the molecular mechanism involved in the inhibition of I-Abeta gene expression in M. bovis BCG-infected macrophages activated with IFN-gamma.     

A 36-amino-acid region of CIITA is an effective inhibitor of CBP: novel mechanism of gamma interferon-mediated suppression of collagen alpha(2)(I) and other promoters

The class II transactivator (CIITA) is induced by gamma interferon (IFN-gamma) and activates major histocompatibility complex class II; however, this report shows it suppresses other genes. An N-terminal 36 amino acids of CIITA mediates suppression of the collagen alpha(2)(I) promoter via binding to CREB-binding protein (CBP). Reconstitution of cells with CBP reverts this suppression. IFN-gamma is known to inhibit collagen gene expression; to test if CIITA mediates this gene suppression, a mutant cell line defective in CIITA induction but not in the activation of STAT1/JAK/IRF-1 is studied. IFN-gamma suppression of the collagen promoter and the endogenous gene is observed in the wild-type control but not in the mutant line. Suppression is restored when CIITA is introduced. Other targets of CIITA-mediated promoter suppression include interleukin 4, thymidine kinase, and cyclin D1.