The expression of MHC class I molecules is crucial for the initiation and regulation of adaptive immune responses against pathogens. NOD-, LRR- and CARD-containing 5 (NLRC5) was recently identified ...as a specific transactivator of MHC class I genes (CITA). NLRC5 and the master regulator for MHC class II genes, class II transactivator (CIITA), interact with similar MHC promoter-bound factors. Here, we provide a broad overview of the molecular mechanisms behind MHC class I transcription and the role of the class I transactivator NLRC5 in MHC class I-dependent immune responses.
Uveal melanoma (UM) with an inflammatory phenotype, characterized by infiltrating leukocytes and increased human leukocyte antigen (HLA) expression, carry an increased risk of death due to ...metastases. These tumors should be ideal for T-cell based therapies, yet it is not clear why prognostically-infaust tumors have a high HLA expression. We set out to determine whether the level of HLA molecules in UM is associated with other genetic factors, HLA transcriptional regulators, or microenvironmental factors.
28 enucleated UM were used to study HLA class I and II expression, and several regulators of HLA by immunohistochemistry, PCR microarray, qPCR and chromosome SNP-array. Fresh tumor samples of eight primary UM and four metastases were compared to their corresponding xenograft in SCID mice, using a PCR microarray and SNP array.
Increased expression levels of HLA class I and II showed no dosage effect of chromosome 6p, but, as expected, were associated with monosomy of chromosome 3. Increased HLA class I and II protein levels were positively associated with their gene expression and with raised levels of the peptide-loading gene TAP1, and HLA transcriptional regulators IRF1, IRF8, CIITA, and NLRC5, revealing a higher transcriptional activity in prognostically-bad tumors. Implantation of fresh human tumor samples into SCID mice led to a loss of infiltrating leukocytes, and to a decreased expression of HLA class I and II genes, and their regulators.
Our data provides evidence for a proper functioning HLA regulatory system in UM, offering a target for T-cell based therapies.
Nonclassical HLAs, HLA-E and HLA-G, are known to affect clinical outcome in various tumor types. We examined the clinical impact of HLA-E and HLA-G expression in early breast cancer patients, and ...related the results to tumor expression of classical HLA class I. Our study population (n = 677) consisted of all early breast cancer patients primarily treated with surgery in our center between 1985 and 1995. Tissue microarray sections of arrayed tumor and normal control material were immunohistochemically stained for HLA-E and HLA-G. For evaluation of HLA-E and HLA-G and the combined variable, HLA-EG, a binary score was used. Expression of classical HLA class I molecules was determined previously. HLA-E, HLA-G, and HLA-EG on breast tumors were classified as expression in 50, 60, and 23% of patients, respectively. Remarkably, only in patients with loss of classical HLA class I tumor expression, expression of HLA-E (p = 0.027), HLA-G (p = 0.035), or HLA-EG (p = 0.001) resulted in a worse relapse-free period. An interaction was found between classical and nonclassical HLA class I expression (p = 0.002), suggestive for a biological connection. We have demonstrated that, next to expression of classical HLA class I, expression of HLA-E and HLA-G is an important factor in the prediction of outcome of breast cancer patients. These results provide further evidence that breast cancer is immunogenic, but also capable of evading tumor eradication by the host's immune system, by up- or downregulation of HLA class Ia and class Ib loci.
In brain tissues from multiple sclerosis (MS) patients, clusters of activated HLA-DR-expressing microglia, also referred to as preactive lesions, are located throughout the normal-appearing white ...matter. The aim of this study was to gain more insight into the frequency, distribution and cellular architecture of preactive lesions using a large cohort of well-characterized MS brain samples.
Here, we document the frequency of preactive lesions and their association with distinct white matter lesions in a cohort of 21 MS patients. Immunohistochemistry was used to gain further insight into the cellular and molecular composition of preactive lesions.
Preactive lesions were observed in a majority of MS patients (67%) irrespective of disease duration, gender or subtype of disease. Microglial clusters were predominantly observed in the vicinity of active demyelinating lesions and are not associated with T cell infiltrates, axonal alterations, activated astrocytes or blood-brain barrier disruption. Microglia in preactive lesions consistently express interleukin-10 and TNF-α, but not interleukin-4, whereas matrix metalloproteases-2 and -9 are virtually absent in microglial nodules. Interestingly, key subunits of the free-radical-generating enzyme NADPH oxidase-2 were abundantly expressed in microglial clusters.
The high frequency of preactive lesions suggests that it is unlikely that most of them will progress into full-blown demyelinating lesions. Preactive lesions are not associated with blood-brain barrier disruption, suggesting that an intrinsic trigger of innate immune activation, rather than extrinsic factors crossing a damaged blood-brain barrier, induces the formation of clusters of activated microglia.
Major histocompatibility complex (MHC) class I and class II molecules play critical roles in the activation of the adaptive immune system by presenting antigens to CD8+ and CD4+ T cells, ...respectively. Although it has been well known that CIITA (MHC class II transactivator), an NLR (nucleotide-binding domain, leucine-rich-repeat containing) protein, as a master regulator of MHC class II gene expression, the mechanism of MHC class I gene transactivation was unclear. Recently, another NLR protein, NLRC5 (NLR family, CARD domain-containing 5), was identified as an MHC class I transactivator (CITA). NLRC5 is a critical regulator for the transcriptional activation of MHC class I genes and other genes involved in the MHC class I antigen presentation pathway. CITA/NLRC5 plays a crucial role in human cancer immunity through the recruitment and activation of tumor killing CD8+ T cells. Here, we discuss the molecular function and mechanism of CITA/NLRC5 in the MHC class I pathway and its role in cancer.
Autosomal-recessive immunodeficiency, centromeric instability, and facial anomalies (ICF) syndrome is mainly characterized by recurrent, often fatal, respiratory and gastrointestinal infections. ...About 50% of patients carry mutations in the DNA methyltransferase 3B gene (DNMT3B) (ICF1). The remaining patients carry unknown genetic defects (ICF2) but share with ICF1 patients the same immunological and epigenetic features, including hypomethylation of juxtacentromeric repeat sequences. We performed homozygosity mapping in five unrelated ICF2 patients with consanguineous parents and then performed whole-exome sequencing in one of these patients and Sanger sequencing in all to identify mutations in the zinc-finger- and BTB (bric-a-bric, tramtrack, broad complex)-domain-containing 24 (ZBTB24) gene in four consanguineously descended ICF2 patients. Additionally, we found ZBTB24 mutations in an affected sibling pair and in one patient for whom it was not known whether his parents were consanguineous. ZBTB24 belongs to a large family of transcriptional repressors that include members, such as BCL6 and PATZ1, with prominent regulatory roles in hematopoietic development and malignancy. These data thus indicate that ZBTB24 is involved in DNA methylation of juxtacentromeric DNA and in B cell development and/or B and T cell interactions. Because ZBTB24 is a putative DNA-binding protein highly expressed in the lymphoid lineage, we predict that by studying the molecular function of ZBTB24, we will improve our understanding of the molecular pathophysiology of ICF syndrome and of lymphocyte biology in general.
Major histocompatibility complex (MHC)-I and MHC-II molecules play an essential role in the immune response to pathogens by virtue of their ability to present peptides to CD8⁺ and CD4⁺ T cells, ...respectively. Given this critical role, MHC-I and MHC-II genes are regulated in a tight fashion at the transcriptional level by a variety of transcription factors that interact with conserved cis-acting regulatory promoter elements. In addition to the activities of these regulatory factors, modification of chromatin also plays an essential role in the efficient transcription of these genes to meet with local requirement for an effective immune response. The focus of this review is on the transcription factors that interact with conserved cis-acting promoter elements and the epigenetic mechanisms that modulate induced and constitutive expression of these MHC genes.
MHC class I plays a critical role in the immune defense against viruses and tumors by presenting antigens to CD8 T cells. An NLR protein, class II transactivator (CIITA), is a key regulator of MHC ...class II gene expression that associates and cooperates with transcription factors in the MHC class II promoter. Although CIITA also transactivates MHC class I gene promoters, loss of CIITA in humans and mice results in the severe reduction of only MHC class II expression, suggesting that additional mechanisms regulate the expression of MHC class I. Here, we identify another member of the NLR protein family, NLRC5, as a transcriptional regulator of MHC class I genes. Similar to CIITA, NLRC5 is an IFN-γ–inducible nuclear protein, and the expression of NLRC5 resulted in enhanced MHC class I expression in lymphoid as well as epithelial cell lines. Using chromatin immunoprecipitation and reporter gene assays, we show that NLRC5 associates with and activates the promoters of MHC class I genes. Furthermore, we show that the IFN-γ–induced up-regulation of MHC class I requires NLRC5, because knockdown of NLRC5 specifically impaired the expression of MHC class I. In addition to MHC class I genes, NLRC5 also induced the expression of β2-microglobulin, transporter associated with antigen processing, and large multifunctional protease, which are essential for MHC class I antigen presentation. Our results suggest that NLRC5 is a transcriptional regulator, orchestrating the concerted expression of critical components in the MHC class I pathway.
The cytokine interferon‐γ (IFNγ) can induce expression of MHC class II (MHCII) on many different cell types, leading to antigen presentation to CD4+ T cells and immune activation. This has also been ...linked to anti‐tumour immunity and graft‐versus‐host disease. The extent of MHCII upregulation by IFNγ is cell type‐dependent and under extensive control of epigenetic regulators and signalling pathways. Here, we identify novel genetic and chemical factors that control this form of MHCII expression. Loss of the oxidative stress sensor Keap1, autophagy adaptor p62/SQSTM1, ubiquitin E3‐ligase Cullin‐3 and chromatin remodeller BPTF impair IFNγ‐mediated MHCII expression. A similar phenotype is observed for arsenite, an oxidative stressor. Effects of the latter can be reversed by the inhibition of HDAC1/2, linking oxidative stress conditions to epigenetic control of MHCII expression. Furthermore, dimethyl fumarate, an antioxidant used for the treatment of several autoimmune diseases, impairs the IFNγ response by manipulating transcriptional control of MHCII. We describe novel pathways and drugs related to oxidative conditions in cells impacting on IFNγ‐mediated MHCII expression, which provide a molecular basis for the understanding of MHCII‐associated diseases.
Synopsis
IFNγ‐induced expression of MHC class II molecules in non‐immune tissue is critical for inflammation and autoimmune diseases. This study identifies novel genetic and chemical factors that control MHCII expression, and potentially autoimmune diseases.
Oxidative stressors affect IFNγ‐induced MHC class II expression in non‐haematopoietic tissues.
A Keap1‐Cul3‐MYST1‐BPTF axis and HDAC1/2 control IFNγ‐induced MHC class II expression.
These pathways are targeted by arsenite but also by autoimmune suppressing drugs such as DMF.
IFNγ‐induced expression of MHC class II molecules in non‐immune tissue is critical for inflammation and autoimmune diseases. This study identifies novel genetic and chemical factors that control MHCII expression, and potentially autoimmune diseases.
Genetic P300/CBP-associated factor (PCAF) variation affects restenosis-risk in patients. PCAF has lysine acetyltransferase activity and promotes nuclear factor kappa-beta (NFκB)-mediated ...inflammation, which drives post-interventional intimal hyperplasia development. We studied the contributing role of PCAF in post-interventional intimal hyperplasia.
PCAF contribution to inflammation and intimal hyperplasia was assessed in leukocytes, macrophages and vascular smooth muscle cells (vSMCs) in vitro and in a mouse model for intimal hyperplasia, in which a cuff is placed around the femoral artery. PCAF deficiency downregulate CCL2, IL-6 and TNF-alpha expression, as demonstrated on cultured vSMCs, leukocytes and macrophages. PCAF KO mice showed a 71.8% reduction of vSMC-rich intimal hyperplasia, a 73.4% reduction of intima/media ratio and a 63.7% reduction of luminal stenosis after femoral artery cuff placement compared to wild type (WT) mice. The association of PCAF and vascular inflammation was further investigated using the potent natural PCAF inhibitor garcinol. Garcinol treatment reduced CCL2 and TNF-alpha expression, as demonstrated on cultured vSMCs and leukocytes. To assess the effect of garcinol treatment on vascular inflammation we used hypercholesterolemic ApoE*3-Leiden mice. After cuff placement, garcinol treatment resulted in reduced arterial leukocyte and macrophage adherence and infiltration after three days compared to untreated animals.
These results identify a vital role for the lysine acetyltransferase PCAF in the regulation of local inflammation after arterial injury and likely the subsequent vSMC proliferation, responsible for intimal hyperplasia.
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