Type I interferon transcriptional network regulates expression of coinhibitory receptors in human T cells.

Although inhibition of T cell coinhibitory receptors has revolutionized cancer therapy, the mechanisms governing their expression on human T cells have not been elucidated. In the present study, we show that type 1 interferon (IFN-I) regulates coinhibitory receptor expression on human T cells, inducing PD-1/TIM-3/LAG-3 while inhibiting TIGIT expression. High-temporal-resolution mRNA profiling of IFN-I responses established the dynamic regulatory networks uncovering three temporal transcriptional waves. Perturbation of key transcription factors (TFs) and TF footprint analysis revealed two regulator modules with different temporal kinetics that control expression of coinhibitory receptors and IFN-I response genes, with SP140 highlighted as one of the key regulators that differentiates LAG-3 and TIGIT expression. Finally, we found that the dynamic IFN-I response in vitro closely mirrored T cell features in acute SARS-CoV-2 infection. The identification of unique TFs controlling coinhibitory receptor expression under IFN-I response may provide targets for enhancement of immunotherapy in cancer, infectious diseases and autoimmunity.

Author Correction: Activated ß-catenin in Foxp3+ regulatory T cells links inflammatory environments to autoimmunity.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

Activated ß-catenin in Foxp3+ regulatory T cells links inflammatory environments to autoimmunity.

Foxp3+ regulatory T cells (Treg cells) are the central component of peripheral immune tolerance. Whereas a dysregulated Treg cytokine signature has been observed in autoimmune diseases, the regulatory mechanisms underlying pro- and anti-inflammatory cytokine production are elusive. Here, we identify an imbalance between the cytokines IFN-γ and IL-10 as a shared Treg signature present in patients with multiple sclerosis and under high-salt conditions. RNA-sequencing analysis on human Treg subpopulations revealed ß-catenin as a key regulator of IFN-γ and IL-10 expression. The activated ß-catenin signature was enriched in human IFN-γ+ Treg cells, as confirmed in vivo with Treg-specific ß-catenin-stabilized mice exhibiting lethal autoimmunity with a dysfunctional Treg phenotype. Moreover, we identified prostaglandin E receptor 2 (PTGER2) as a regulator of IFN-γ and IL-10 production under a high-salt environment, with skewed activation of the ß-catenin-SGK1-Foxo axis. Our findings reveal a novel PTGER2-ß-catenin loop in Treg cells linking environmental high-salt conditions to autoimmunity.

A predominant role for the HLA class II region in the association of the MHC region with multiple sclerosis.

Genetic susceptibility to multiple sclerosis is associated with genes of the major histocompatibility complex (MHC), particularly HLA-DRB1 and HLA-DQB1 (ref. 1). Both locus and allelic heterogeneity have been reported in this genomic region. To clarify whether HLA-DRB1 itself, nearby genes in the region encoding the MHC or combinations of these loci underlie susceptibility to multiple sclerosis, we genotyped 1,185 Canadian and Finnish families with multiple sclerosis (n = 4,203 individuals) with a high-density SNP panel spanning the genes encoding the MHC and flanking genomic regions. Strong associations in Canadian and Finnish samples were observed with blocks in the HLA class II genomic region (P < 4.9 x 10(-13) and P < 2.0 x 10(-16), respectively), but the strongest association was with HLA-DRB1 (P < 4.4 x 10(-17)). Conditioning on either HLA-DRB1 or the most significant HLA class II haplotype block found no additional block or SNP association independent of the HLA class II genomic region. This study therefore indicates that MHC-associated susceptibility to multiple sclerosis is determined by HLA class II alleles, their interactions and closely neighboring variants.

Genetic mapping across autoimmune diseases reveals shared associations and mechanisms.

Autoimmune and inflammatory diseases are polygenic disorders of the immune system. Many genomic loci harbor risk alleles for several diseases, but the limited resolution of genetic mapping prevents determining whether the same allele is responsible, indicating a shared underlying mechanism. Here, using a collection of 129,058 cases and controls across 6 diseases, we show that ~40% of overlapping associations are due to the same allele. We improve fine-mapping resolution for shared alleles twofold by combining cases and controls across diseases, allowing us to identify more expression quantitative trait loci driven by the shared alleles. The patterns indicate widespread sharing of pathogenic mechanisms but not a single global autoimmune mechanism. Our approach can be applied to any set of traits and is particularly valuable as sample collections become depleted.

A ChIP-seq defined genome-wide map of vitamin D receptor binding: associations with disease and evolution.

Initially thought to play a restricted role in calcium homeostasis, the pleiotropic actions of vitamin D in biology and their clinical significance are only now becoming apparent. However, the mode of action of vitamin D, through its cognate nuclear vitamin D receptor (VDR), and its contribution to diverse disorders, remain poorly understood. We determined VDR binding throughout the human genome using chromatin immunoprecipitation followed by massively parallel DNA sequencing (ChIP-seq). After calcitriol stimulation, we identified 2776 genomic positions occupied by the VDR and 229 genes with significant changes in expression in response to vitamin D. VDR binding sites were significantly enriched near autoimmune and cancer associated genes identified from genome-wide association (GWA) studies. Notable genes with VDR binding included IRF8, associated with MS, and PTPN2 associated with Crohn's disease and T1D. Furthermore, a number of single nucleotide polymorphism associations from GWA were located directly within VDR binding intervals, for example, rs13385731 associated with SLE and rs947474 associated with T1D. We also observed significant enrichment of VDR intervals within regions of positive selection among individuals of Asian and European descent. ChIP-seq determination of transcription factor binding, in combination with GWA data, provides a powerful approach to further understanding the molecular bases of complex diseases.

Pearls & Oy-sters: Homozygous Complement Factor I Deficiency Presenting as Fulminant Relapsing Complement-Mediated CNS Vasculitis.

A 36-year-old man presented multiple times with fever, headache, alteration of mental status, and focal neurologic deficits. MRI revealed extensive white matter lesions that were partially reversed between episodes. Workup revealed persistently low complement factor C3, low factor B, and absent alternative complement pathway activity. Biopsy revealed neutrophilic vasculitis. Genetic testing revealed a homozygous variant in complement factor I (CFI), which was thought to be pathogenic. CFI regulates complement-mediated inflammation, and deficiency in this factor leads to unchecked alternative pathway activity and decrease in C3 and factor B through consumption. The patient has remained stable since starting IL-1ß inhibition. Complement factor I is a rare disorder that should be considered in patients with atypical relapsing neurologic disease associated with neutrophilic pleocytosis.

Parent-of-origin effects at the major histocompatibility complex in multiple sclerosis.

Multiple sclerosis (MS) susceptibility is characterized by maternal parent-of-origin effects and increased female penetrance. In 7796 individuals from 1797 MS families (affected individuals n = 2954), we further implicate epigenetic modifications within major histocompatibility complex (MHC) class II haplotypes as mediating these phenomena. Affected individuals with the main MS-associated allele HLA-DRB1*15 had a higher female-to-male ratio versus those lacking it (P = 0.00023). Distorted transmission of MHC haplotypes by both parent-of-origin and gender-of-affected-offspring was most evident in the maternal HLA-DRB1*15 transmission to affected female offspring (OR = 3.31, 95% CI = 2.59-4.24) contrasting with similarity among maternal transmission to affected male offspring (OR = 2.13, 95% CI = 1.44-3.14), paternal transmissions to affected female (OR = 2.14, 95% CI = 1.64-2.78) and male (OR = 2.16, 95% CI = 1.37-3.39) offspring. Significant parent-of-origin effects were observed in affected females (maternal: P = 9.33 x 10(-42); paternal: P = 1.12 x 10(-15); comparison: P = 0.0014), but not in affected males (maternal: P = 6.70 x 10(-8); paternal: P = 2.54 x 10(-6); comparison: P = 0.95). Conditional logistic regression analysis revealed further differential risk of HLA diplotypes. Risks for HLA-DRB1*15 and likely for other HLA-DRB1 haplotypes were restricted by (i) parent-of-origin, (ii) gender-of-offspring and (iii) trans epistasis in offspring. These findings may illuminate the gender bias characterizing autoimmunity overall. They raise questions about the concept of restricted antigen presentation in autoimmunity and suggest that gender-specific epigenetic interactions may be the driving forces behind the MHC haplotypic associations. Haplotype-specific epigenetic modifications at MHC class II and their decay appear to be at the heart of MS pathogenesis and inheritance of risk, providing the focus for gene-environment interactions that determine susceptibility and resistance.

Expression of the multiple sclerosis-associated MHC class II Allele HLA-DRB1*1501 is regulated by vitamin D.

Multiple sclerosis (MS) is a complex trait in which allelic variation in the MHC class II region exerts the single strongest effect on genetic risk. Epidemiological data in MS provide strong evidence that environmental factors act at a population level to influence the unusual geographical distribution of this disease. Growing evidence implicates sunlight or vitamin D as a key environmental factor in aetiology. We hypothesised that this environmental candidate might interact with inherited factors and sought responsive regulatory elements in the MHC class II region. Sequence analysis localised a single MHC vitamin D response element (VDRE) to the promoter region of HLA-DRB1. Sequencing of this promoter in greater than 1,000 chromosomes from HLA-DRB1 homozygotes showed absolute conservation of this putative VDRE on HLA-DRB1*15 haplotypes. In contrast, there was striking variation among non-MS-associated haplotypes. Electrophoretic mobility shift assays showed specific recruitment of vitamin D receptor to the VDRE in the HLA-DRB1*15 promoter, confirmed by chromatin immunoprecipitation experiments using lymphoblastoid cells homozygous for HLA-DRB1*15. Transient transfection using a luciferase reporter assay showed a functional role for this VDRE. B cells transiently transfected with the HLA-DRB1*15 gene promoter showed increased expression on stimulation with 1,25-dihydroxyvitamin D3 (P = 0.002) that was lost both on deletion of the VDRE or with the homologous "VDRE" sequence found in non-MS-associated HLA-DRB1 haplotypes. Flow cytometric analysis showed a specific increase in the cell surface expression of HLA-DRB1 upon addition of vitamin D only in HLA-DRB1*15 bearing lymphoblastoid cells. This study further implicates vitamin D as a strong environmental candidate in MS by demonstrating direct functional interaction with the major locus determining genetic susceptibility. These findings support a connection between the main epidemiological and genetic features of this disease with major practical implications for studies of disease mechanism and prevention.

Epistasis among HLA-DRB1, HLA-DQA1, and HLA-DQB1 loci determines multiple sclerosis susceptibility.

Multiple sclerosis (MS), a common central nervous system inflammatory disease, has a major heritable component. Susceptibility is associated with the MHC class II region, especially HLA-DRB5*0101-HLA-DRB1*1501-HLA-DQA1*0102-HLA-DQB1*0602 haplotypes(hereafter DR2), which dominate genetic contribution to MS risk. Marked linkage disequilibrium (LD) among these loci makes identification of a specific locus difficult. The once-leading candidate, HLA-DRB1*15, localizes to risk, neutral, and protective haplotypes. HLA-DRB1*15 and HLA-DQB1*0602, nearly always located together on a small ancestral chromosome segment, are strongly MS-associated. One intervening allele on this haplotype, viz. HLA-DQA1*0102, shows no primary MS association. Two Canadian cohorts (n = 830 and n = 438 trios) genotyped for HLA-DRB1, HLA-DQA1 and HLA-DQB1 alleles were tested for association using TDT. To evaluate epistasis involving HLA-DRB1*15, transmissions from HLA-DRB1*15-negative parents were stratified by the presence/absence of HLA-DRB1*15 in affected offspring. All 3 alleles contribute to MS susceptibility through novel epistatic interactions. HLA-DQA1*0102 increased disease risk when combined with HLA-DRB1*1501 in trans, thereby unambiguously implicating HLA-DQ in MS susceptibility. Three-locus haplotypes demonstrated that HLA-DRB1*1501 and HLA-DQB1*0602 each influence risk. Transmissions of rare morcellated DR2 haplotypes showed no interaction with HLA-DQA1*0102. Incomplete haplotypes bearing only HLA-DRB1*1501 or HLA-DQB1*0602 did not predispose to MS. Balanced reciprocal transmission distortion can mask epistatic allelic association. These findings implicate epistasis among HLA class II alleles in human immune responses generally, provide partial explanation for intense linkage disequilibrium in the MHC, have relevance to animal models, and demonstrate key roles for DR2-specific interactions in MS susceptibility. MHC disease associations may be more generally haplotypic or diplotypic.

A multiple sclerosis-protective coding variant reveals an essential role for HDAC7 in regulatory T cells.

Genome-wide association studies identifying hundreds of susceptibility loci for autoimmune diseases indicate that genes active in immune cells predominantly mediate risk. However, identification and functional characterization of causal variants remain challenging. Here, we focused on the immunomodulatory role of a protective variant of histone deacetylase 7 (HDAC7). This variant (rs148755202, HDAC7.p.R166H) was identified in a study of low-frequency coding variation in multiple sclerosis (MS). Through transcriptomic analyses, we demonstrate that wild-type HDAC7 regulates genes essential for the function of Foxp3+ regulatory T cells (Tregs), an immunosuppressive subset of CD4 T cells that is generally dysfunctional in patients with MS. Moreover, Treg-specific conditional hemizygous deletion of HDAC7 increased the severity of experimental autoimmune encephalitis (EAE), a mouse model of neuroinflammation. In contrast, Tregs transduced with the protective HDAC7 R166H variant exhibited higher suppressive capacity in an in vitro functional assay, mirroring phenotypes previously observed in patient samples. In vivo modeling of the human HDAC7 R166H variant by generation of a knock-in mouse model bearing an orthologous R150H substitution demonstrated decreased EAE severity linked to transcriptomic alterations of brain-infiltrating Tregs, as assessed by single-cell RNA sequencing. Our data suggest that dysregulation of epigenetic modifiers, a distinct molecular class associated with disease risk, may influence disease onset. Last, our approach provides a template for the translation of genetic susceptibility loci to detailed functional characterization, using in vitro and in vivo modeling.

Toward Precision Phenotyping of Multiple Sclerosis.

The classification of multiple sclerosis (MS) has been established by Lublin in 1996 and revised in 2013. The revision includes clinically isolated syndrome, relapsing-remitting, primary progressive and secondary progressive MS, and has added activity (i.e., formation of white matter lesions or clinical relapses) as a qualifier. This allows for the distinction between active and nonactive progression, which has been shown to be of clinical importance. We propose that a logical extension of this classification is the incorporation of additional key pathological processes, such as chronic perilesional inflammation, neuroaxonal degeneration, and remyelination. This will distinguish MS phenotypes that may present as clinically identical but are driven by different combinations of pathological processes. A more precise description of MS phenotypes will improve prognostication and personalized care as well as clinical trial design. Thus, our proposal provides an expanded framework for conceptualizing MS and for guiding development of biomarkers for monitoring activity along the main pathological axes in MS.

Epigenetics in multiple sclerosis susceptibility: difference in transgenerational risk localizes to the major histocompatibility complex.

Multiple sclerosis (MS) susceptibility demonstrates a complex pattern of inheritance. Haplotypes containing HLA-DRB1*1501 carry most of the genetic risk. Epidemiological evidence implicating epigenetic factors includes complex distortion of disease transmission seen in aunt/uncle-niece/nephew (AUNN) pairs. Unexpectedly, in AUNN families we found that allele frequencies for HLA-DRB1*1501 were different between the first and second generations affected. Affected aunts had significantly lower HLA-DRB1*15 frequency compared with their affected nieces (chi(2) = 9.90, P = 0.0016), whereas HLA-DRB1*15 frequency in affected males remains unaltered across the two generations (chi(2) = 0.23, P = 0.63). We compared transmissions for the HLA-DRB1*15 allele using a family-based transmission disequilibrium test approach in 1690 individuals from 350 affected sibling pair (ASP) families and 960 individuals from 187 AUNN families. Transmissions differed between the ASP and the AUNN families (chi(2) = 6.92; P = 0.0085). The risk carried by HLA-DRB1*15 was increased in families with affected second-degree relatives (AUNN: OR = 4.07) when compared with those consisting only first-degree relatives (ASP: OR = 2.17), establishing heterogeneity of risk among HLA-DRB1*15 haplotypes based on whether collateral parental relatives are affected. These observations strongly implicate gene-environment interactions in susceptibility and more specifically, that epigenetic modifications differentiate among human leukocyte antigen class II risk haplotypes and are involved in the determination of the gender bias in MS. These data strongly suggest that the female-specific increasing risk of MS is mediated through these alleles or adjacent variation. The comparison of transmission of the same allele in vertically affected pedigrees (AUNN) to collinear sibling pairs (ASP) may provide a useful screen for putative epigenetic marks.

Of mice and men: experimental autoimmune encephalitis and multiple sclerosis.

BACKGROUND: Research using experimental autoimmune encephalitis (EAE) models accounts for almost 20% of the papers. published in multiple sclerosis (MS). METHODS: We performed a literature review of papers indexed with ISI Web of Science on EAE and MS over the last 30 years and a detailed analysis of studies of molecular pathways in EAE published in 2008 and 2009. RESULTS: The impact of EAE studies declines more rapidly than other studies published on MS (EAE cited corrected half-life = 4·00 years vs. MS cited corrected half-life = 9·66 years, P < 0·0001). The pathology of EAE differs quite markedly from that observed in the human disease. EAE has implicated many different genes as important to pathogenesis but only a minority of these are supported by human studies. CONCLUSIONS: Future research should critically appraise precisely what is being modelled by EAE before drawing conclusions about human disease.

HLA class I alleles tag HLA-DRB1*1501 haplotypes for differential risk in multiple sclerosis susceptibility.

The major locus for multiple sclerosis (MS) susceptibility is located within the class II region of the Major Histocompatibility Complex (MHC). HLA-DRB1 alleles, constituting the strongest MS susceptibility factors, have been widely exploited in research including construction of transgenic animal models of MS. Many studies have concluded that HLA-DRB1*15 allele itself determines MS-associated susceptibility. If this were true, haplotypes bearing this allele would confer equal risk. If HLA-DRB1*15 bearing haplotypes differed for risk, roles for other loci in this region would be implied and further study of the fine structure of this locus would be compelling. We have tested the hypothesis comparing haplotypes stratified by HLA class I tagging. We show here that HLA-DRB1*15-bearing-haplotypes in 1970 individuals from 494 MS families are indeed heterogeneous. Some HLA-DRB1*15 haplotypes determine susceptibility while others do not. Three groups of class I tagged HLA-DRB1*15 haplotypes were not over-transmitted: (i) HLA-DRB1*15-HLA-B*08 (TR = 25, NT = 23, Odds Ratio = 1.09), (ii) -HLA-B*27 (TR = 18, NT = 17, Odds Ratio = 1.06), and (iii) rare HLA-DRB1*15 haplotypes (frequency <0.02). Rare haplotypes were significantly different from common haplotypes, and transmissions were remarkably similar to those for class-I-matched non-HLA-DRB1*15 haplotypes. These results unambiguously indicate that HLA-DRB1*15 is part of a susceptibility haplotype but cannot be the susceptibility allele itself, requiring either epistatic interactions, epigenetic modifications on some haplotypes, or nearby structural variation. These findings strongly imply that differences among HLA-DRB1*15 haplotypes will furnish the basis for MHC-associated susceptibility in MS and raise the possibility that the MHC haplotype is the fundamental unit of genetic control of immune response.

The inheritance of resistance alleles in multiple sclerosis.

Multiple sclerosis (MS) is a complex trait in which alleles at or near the class II loci HLA-DRB1 and HLA-DQB1 contribute significantly to genetic risk. HLA-DRB1*15 and HLA-DRB1*17-bearing haplotypes and interactions at the HLA-DRB1 locus increase risk of MS but it has taken large samples to identify resistance HLA-DRB1 alleles. In this investigation of 7,093 individuals from 1,432 MS families, we have assessed the validity, mode of inheritance, associated genotypes, and the interactions of HLA-DRB1 resistance alleles. HLA-DRB1*14-, HLA-DRB1*11-, HLA-DRB1*01-, and HLA-DRB1*10-bearing haplotypes are protective overall but they appear to operate by different mechanisms. The first type of resistance allele is characterised by HLA-DRB1*14 and HLA-DRB1*11. Each shows a multiplicative mode of inheritance indicating a broadly acting suppression of risk, but a different degree of protection. In contrast, a second type is exemplified by HLA-DRB1*10 and HLA-DRB1*01. These alleles are significantly protective when they interact specifically in trans with HLA-DRB1*15-bearing haplotypes. HLA-DRB1*01 and HLA-DRB1*10 do not interact with HLA-DRB1*17, implying that several mechanisms may be operative in major histocompatibility complex-associated MS susceptibility, perhaps analogous to the resistance alleles. There are major practical implications for risk and for the exploration of mechanisms in animal models. Restriction of antigen presentation by HLA-DRB1*15 seems an improbably simple mechanism of major histocompatibility complex-associated susceptibility.

Epigenetic fine-mapping: identification of causal mechanisms for autoimmunity.

Genome-wide association studies (GWAS) have identified genetic susceptibility loci for a variety of autoimmune and inflammatory diseases. These studies confirm the fundamental genetic basis of individual autoimmune diseases, and also point to shared etiological mechanisms across the spectrum of autoimmunity. While hundreds of genetic loci have been implicated in autoimmune diseases, the translation of individual susceptibility loci into specific molecular mechanisms for individual diseases remains difficult. This review highlights recent advances in the genetics of autoimmune disease, and the emerging use of epigenetic techniques to identify pathogenic cell types and causal molecular mechanisms of autoimmunity.

PRKCA and multiple sclerosis: association in two independent populations.

Multiple sclerosis (MS) is a chronic disease of the central nervous system responsible for a large portion of neurological disabilities in young adults. Similar to what occurs in numerous complex diseases, both unknown environmental factors and genetic predisposition are required to generate MS. We ascertained a set of 63 Finnish MS families, originating from a high-risk region of the country, to identify a susceptibility gene within the previously established 3.4-Mb region on 17q24. Initial single nucleotide polymorphism (SNP)-based association implicated PRKCA (protein kinase C alpha) gene, and this association was replicated in an independent set of 148 Finnish MS families (p = 0.0004; remaining significant after correction for multiple testing). Further, a dense set of 211 SNPs evenly covering the PRKCA gene and the flanking regions was selected from the dbSNP database and analyzed in two large, independent MS cohorts: in 211 Finnish and 554 Canadian MS families. A multipoint SNP analysis indicated linkage to PRKCA and its telomeric flanking region in both populations, and SNP haplotype and genotype combination analyses revealed an allelic variant of PRKCA, which covers the region between introns 3 and 8, to be over-represented in Finnish MS cases (odds ratio = 1.34, 95% confidence interval 1.07-1.68). A second allelic variant, covering the same region of the PRKCA gene, showed somewhat stronger evidence for association in the Canadian families (odds ratio = 1.64, 95% confidence interval 1.39-1.94). Initial functional relevance for disease predisposition was suggested by the expression analysis: The transcript levels of PRKCA showed correlation with the copy number of the Finnish and Canadian "risk" haplotypes in CD4-negative mononuclear cells of five Finnish multiplex families and in lymphoblast cell lines of 11 Centre d'Etude du Polymorphisme Humain (CEPH) individuals of European origin.

Parental transmission of HLA-DRB1*15 in multiple sclerosis.

Multiple sclerosis (MS) is a complex trait in which HLA-DRB1*15 bearing MHC haplotypes increase risk of MS in people of Northern European descent. In this investigation of 7,334 individuals from 1,515 MS families, the largest cohort used to study the HLA-DRB1 locus to date, we analysed the transmission of HLA-DRB1*15 haplotypes stratified by sex of transmitting parent. A significant over transmission of HLA-DRB1*15 from mothers was observed (chi (2) = 7.73, P = 0.0054), suggesting that parent of origin effects at the MHC determine susceptibility to MS.

Parental non-inherited HLA resistance alleles do not confer protection against multiple sclerosis.

Multiple sclerosis (MS) is a complex trait in which alleles at or near the class II loci HLA-DRB1 and HLA-DQB1 contribute significantly to genetic risk. HLA-DRB1*14 and DRB1*11 bearing haplotypes protect against MS and DRB1*01 and DRB1*10 interact with DRB1*15 to reduce risk of the disease. Recent work in other autoimmune diseases such as rheumatoid arthritis has suggested that maternal non-transmitted protective alleles can also confer disease resistance. In this investigation of 7093 individuals from 1432 MS families, we have analysed the transmission of HLA-DRB1*14,*11,*10 and *01 haplotypes, stratified by sex of parent. No significant transmission differences between mothers and fathers were found, suggesting that non-inherited resistance alleles do not appear to play a role in MS.