OBJECTIVE: To determine how polymorphism of transporter associated with antigen processing 1 and 2 (TAP1 and 2) alleles contributed to the pathogenesis of systemic lupus erythematosus (SLE) in Japanese patients. METHODS: TAP1 and TAP2 typing was carried out in 52 Japanese patients with SLE and 95 normal subjects by the PCR-RFLP (restriction fragment length polymorphism) method. HLA-DR typing and HLA-DRB1*15 genotyping were carried out by the PCR method and PCR-SSCP (single stranded DNA conformation polymorphism) method, respectively. RESULTS: No particular TAP 1 allele was associated with Japanese SLE or with immunological subgroup of SLE. TAP2H showed a tendency towards increased frequency in SLE (5.8% v 0% in control), but the corrected P value was not significant. No other particular association of TAP2 allele was observed. Furthermore, these was no evidence for linkage disequilibrium between any TAP1/TAP2 alleles and HLA-DRB1*1501--which is reported to be weakly but significantly association with Japanese SLE--in either the normal control or the SLE patient group. CONCLUSIONS: Neither the TAP1 nor the TAP2 gene appears to determine disease susceptibility to SLE in Japanese, and these results are in keeping with those reported in Caucasian SLE patients.
OBJECTIVES--To determine whether the TAP2 transporter gene, which lies between HLA-DP and HLA-DQ, is involved in determining susceptibility to systemic lupus erythematosus (SLE). METHODS--TAP2 types were determined by ARMS-PCR in 89 white patients with SLE and 156 control subjects. RESULTS--No particular TAP2 dimorphism or allele was associated with SLE or with any clinical/immunological subgroup of SLE. Furthermore, there was no evidence for significant linkage disequilibrium between TAP2 and HLA-DQ/DR in SLE. CONCLUSIONS--These data suggest that TAP2 is not a disease susceptibility gene for SLE and that the disease-predisposing haplotypes do not extend as far as TAP2. This indicates that any HLA-DP association with SLE must be independent of other class II (DQ/DR) associations.
A substantial genetic contribution to systemic lupus erythematosus (SLE) risk is conferred by major histocompatibility complex (MHC) gene(s) on chromosome 6p21. Previous studies in SLE have lacked statistical power and genetic resolution to fully define MHC influences. We characterized 1,610 Caucasian SLE cases and 1,470 parents for 1,974 MHC SNPs, the highly polymorphic HLA-DRB1 locus, and a panel of ancestry informative markers. Single-marker analyses revealed strong signals for SNPs within several MHC regions, as well as with HLA-DRB1 (global p = 9.99×10−16). The most strongly associated DRB1 alleles were: *0301 (odds ratio, OR = 2.21, p = 2.53×10−12), *1401 (OR = 0.50, p = 0.0002), and *1501 (OR = 1.39, p = 0.0032). The MHC region SNP demonstrating the strongest evidence of association with SLE was rs3117103, with OR = 2.44 and p = 2.80×10−13. Conditional haplotype and stepwise logistic regression analyses identified strong evidence for association between SLE and the extended class I, class I, class III, class II, and the extended class II MHC regions. Sequential removal of SLE–associated DRB1 haplotypes revealed independent effects due to variation within OR2H2 (extended class I, rs362521, p = 0.006), CREBL1 (class III, rs8283, p = 0.01), and DQB2 (class II, rs7769979, p = 0.003, and rs10947345, p = 0.0004). Further, conditional haplotype analyses demonstrated that variation within MICB (class I, rs3828903, p = 0.006) also contributes to SLE risk independent of HLA-DRB1*0301. Our results for the first time delineate with high resolution several MHC regions with independent contributions to SLE risk. We provide a list of candidate variants based on biologic and functional considerations that may be causally related to SLE risk and warrant further investigation.
Systemic lupus erythematosus (SLE) is an autoimmune disease characterized by autoantibody production and involvement of multiple organ systems. Although the cause of SLE remains unknown, several lines of evidence underscore the importance of genetic factors. As is true for most autoimmune diseases, a substantial genetic contribution to disease risk is conferred by major histocompatibility complex (MHC) gene(s) on chromosome 6. This region of the genome contains a large number of genes that participate in the immune response. However, the full contribution of this genomic region to SLE risk has not yet been defined. In the current study we characterize a large number of SLE patients and family members for approximately 2,000 MHC region variants to identify the specific genes that influence disease risk. Our results, for the first time, implicate four different MHC regions in SLE risk. We provide a list of candidate variants based on biologic and functional considerations that may be causally related to SLE risk and warrant further investigation.
OBJECTIVE—To study the influence MHC class II and TAP2 alleles exert on systemic lupus erythematosus (SLE) susceptibility and on the clinical and serological manifestations of the disease, in a cohort of Spanish patients.
METHODS—HLA-DR serological typing and HLA-DQA, DQB, and TAP2 DNA sequence specific oligotyping, were carried out in 85 unrelated Spanish SLE patients and 186 healthy controls. Autoantibodies detection was carried out by indirect immunofluorescence and counter immunoelectrophoresis.
RESULTS—Total SLE group: the frequency of HLA-DR3 and HLA-DQA1*0501 is significantly increased in this group (pc<0.005, δ=0.34 and pc<0.005, δ= 0.45, respectively) although the highest δ value (δ=0.87) is obtained when the TAP2*01 alleles are considered. No DQB allele shows significant deviation from the control group. Renal damage: it mainly occurs in HLA-DR3 patients (pc<0.0005 and δ=0.72). HLA-DQA1*0501 (pc<0.05, δ=0.57) and DQB1*0201 (pc NS, δ=0.56) are weaker susceptibility factors. Ro+ (but not La) group: this autoantibody response is associated with TAP2*01 alleles in homozygosity (p<0.05, δ=0.81). Ro/La+ group: it has a different genetic background as HLA-DQA1*0501 (δ=1) and HLA-DQB1*0201 (δ=1) are the main susceptibility factors.
CONCLUSIONS—A differential association between HLA-DR, DQA1, and DQB1 alleles and SLE or its clinical and serological manifestations are found. Furthermore, the associations are different to the ones reported in other ethnic groups. Finally, TAP2*01 group of alleles are associated with the highest susceptibility to SLE (higher than HLA-DR3) and may influence Ro (but not La) autoantibodies production, whereas HLA-DQA1*0501 and DQB1*0201 mediates concomitant Ro and La production.
Keywords: TAP2 alleles; HLA class II alleles; systemic lupus erythematosus
Transporter associated with antigen processing (TAP), a member of the ATP-binding cassette transporter super family, is composed of two integral membrane proteins, TAP-1 and TAP-2. The TAP gene product is involved in the processing of endogenous peptides that bind to MHC class I molecules. Mutations and/or polymorphism within these genes could alter the efficacy of the immune response which might be relevant for the development of autoimmune diseases and cancer.
DNA was isolated from peripheral blood sample of 200 patients with cervical cancer and 200 healthy controls. TAP1 and TAP2 allele polymorphism were determined by polymerase chain reaction.
Significant protective OR (OR = 0.22 95% CI = 0.09–0.51, P < 0.001-OR = 0.47, 95% CI = 0.24–0.92, P = 0.02) was observed for GG and combined AG+GG genotypes of TAP2 in patients with SCC respectively. Similarly, such genotypes (GG, AG+GG) appeared same OR for patient with cervical cancer in study group (OR = 0.12, 95% CI = 0.04–0.39-P < 0.001-OR = 0.5 ,95% CI = 0.25–0.95-P = 0.03). There was decrease risk of cervical cancer in user of oral contraceptive with AG and GG genotypes of TAP2 (OR = 0.55, 95% Cl = 0.41–0.73, P = 0.002, OR = 0.09, 95% CI = 0.02–0.36, P < 0.001) respectively. In case of TAP1 gene all allelic polymorphisms showed a decrease OR in patients with cervical cancer in passive smokers and user of oral contraceptives, though, no significant
Thus, TAP1 and TAP2 genes polymorphism are not linked to cervical carcinoma, since no association was found between a particular genotype and the disease.
Cervical cancer; Polymorphism; TAP1; TAP2; Gene
OBJECTIVES--To determine whether HLA-DP genes are involved in determining susceptibility to systemic lupus erythematosus (SLE). METHODS--HLA-DPA1 and DPB1 genes were amplified by PCR of DNA samples from a panel of patients with SLE and normal controls. Amplified DNA was blotted on to nylon filters and probed with sequence-specific oligonucleotide (SSO) probes. RESULTS--No DPA1 or DPB1 allele was significantly associated with SLE, or with any immunological or clinical subset of SLE. Evidence was found for only limited linkage disequilibrium between HLA-DP and HLA-DQ/DR variants, and none between HLA-DP and the TAP2 gene. CONCLUSIONS--These data indicate that HLA-DP genes do not contribute towards determining susceptibility to SLE.
Multidrug resistance (MDR) to anti-cancer drugs has been associated with the overexpression of P-glycoprotein (P-gp) and the multidrug resistance-associated protein (MRP), both being members of the ATP-binding cassette (ABC) superfamily of transporters. We investigated whether in addition to P-gp and MRP, another ABC transporter, the transporter associated with antigen processing (TAP), is associated with MDR. TAP plays a major role in MHC class I-restricted antigen presentation by mediating peptide translocation over the endoplasmic reticulum membrane. TAP1 and P-gp share a significant degree of homology among their transmembrane domains, which are thought to be the primary determinants of substrate specificity, and both can apparently mediate the translocation of peptides. Using immunocytochemistry and Western blot, TAP was overexpressed in parallel with MHC class I in several MDR human cancer cell lines. TAP was overexpressed more frequently in MRP-positive MDR cell lines (three out of three) than in P-gp positive MDR cells (two out of five). Reversal of resistance resulted in a decrease in TAP levels. Transfection of the TAP genes into TAP-deficient lymphoblastoid T2 cells conferred mild resistance to etoposide, vincristine and doxorubicin (2- to 2.5-fold). Furthermore, etoposide and vincristine inhibited TAP-dependent peptide translocation to the endoplasmic reticulum. Collectively, our results suggest that TAP may modestly contribute to the MDR phenotype, in particular in MRP- overexpressing MDR cells. Further insight into the role of TAP in MDR will require the study of other transfectants, as well as the investigation of TAP expression in P-gp and MRP-negative MDR cancer cell lines.
The major histocompatibility complex (MHC) class II transactivator gene (CIITA) encodes an important transcription factor required for HLA class II MHC-restricted antigen presentation. MHC genes, including the HLA class II DRB1*03:01 allele, are strongly associated with systemic lupus erythematosus (SLE). Recently the rs4774 CIITA missense variant (+1632G/C) was reported to be associated with susceptibility to multiple sclerosis. In the current study, we investigated CIITA, DRB1*03:01 and risk of SLE using a multi-stage analysis. In stage 1, 9 CIITA variants were tested in 658 cases and 1,363 controls (N = 2,021). In stage 2, rs4774 was tested in 684 cases and 2,938 controls (N = 3,622). We also performed a meta-analysis of the pooled 1,342 cases and 4,301 controls (N = 5,643). In stage 1, rs4774*C was associated with SLE (odds ratio [OR] = 1.24, 95% confidence interval [95% CI] = 1.07–1.44, P = 4.2 × 10−3). Similar results were observed in stage 2 (OR = 1.16, 95% CI = 1.02–1.33, P = 8.5×10−3) and the meta-analysis of the combined dataset (OR = 1.20, 95% CI = 1.09–1.33, Pmeta = 2.5×10−4). In all three analyses, the strongest evidence for association between rs4774*C and SLE was present in individuals who carried at least one copy of DRB1*03:01 (Pmeta= 1.9×10−3). Results support a role for CIITA in SLE, which appears to be stronger in the presence of DRB1*03:01.
systemic lupus erythematosus; autoimmunity; major histocompatibility complex; HLA; CIITA; MHC2TA
Background: The transporter associated with antigen processing (TAP) is an ABC transporter whose experimental structure is not known.
Results: We have modeled TAP based on the crystal structures of three related ABC transporters.
Conclusion: We identified a possible peptide binding conformation in the transport cycle of TAP.
Significance: These models help interpret experimental data and give information about the transport cycle of ABC transporters in general.
The human transporter associated with antigen processing (TAP) is a member of the ATP binding cassette (ABC) transporter superfamily. TAP plays an essential role in the antigen presentation pathway by translocating cytosolic peptides derived from proteasomal degradation into the endoplasmic reticulum lumen. Here, the peptides are loaded into major histocompatibility class I molecules to be in turn exposed at the cell surface for recognition by T-cells. TAP is a heterodimer formed by the association of two half-transporters, TAP1 and TAP2, with a typical ABC transporter core that consists of two nucleotide binding domains and two transmembrane domains. Despite the availability of biological data, a full understanding of the mechanism of action of TAP is limited by the absence of experimental structures of the full-length transporter. Here, we present homology models of TAP built on the crystal structures of P-glycoprotein, ABCB10, and Sav1866. The models represent the transporter in inward- and outward-facing conformations that could represent initial and final states of the transport cycle, respectively. We described conserved regions in the endoplasmic reticulum-facing loops with a role in the opening and closing of the cavity. We also identified conserved π-stacking interactions in the cytosolic part of the transmembrane domains that could explain the experimental data available for TAP1-Phe-265. Electrostatic potential calculations gave structural insights into the role of residues involved in peptide binding, such as TAP1-Val-288, TAP2-Cys-213, TAP2-Met-218. Moreover, these calculations identified additional residues potentially involved in peptide binding, in turn verified with replica exchange simulations performed on a peptide bound to the inward-facing models.
ABC Transporter; Molecular Docking; Molecular Dynamics; Molecular Modeling; Multidrug Transporters; Peptide Binding; Transporter Associated with Antigen Processing
The transporter associated with antigen processing (TAP) supplies cytosolic peptides into the endoplasmic reticulum for binding to major histocompatibility complex (MHC) class I molecules. Its specificity therefore influences the repertoire of peptides presented by MHC molecules. Compared to human TAP, murine TAP's binding specificity has not been characterized as well, even though murine systems are widely used for basic studies of antigen processing and presentation.
We performed a detailed experimental analysis of murine TAP binding specificity by measuring the binding affinities of 323 peptides. Based on this experimental data, a computational model of murine TAP specificity was constructed. The model was compared to previously generated data on human and murine TAP specificities. In addition, the murine TAP specificities for known epitopes and random peptides were predicted and compared to assess the impact of murine TAP selectivity on epitope selection.
Comparisons to a previously constructed model of human TAP specificity confirms the well-established differences for peptide substrates with positively charged C-termini. In addition these comparisons show that several residues at the N-terminus of peptides which strongly influence binding to human TAP showed little effect on binding to murine TAP, and that the overall influence of the aminoterminal residues on peptide affinity for murine TAP is much lower than for the human transporter. Murine TAP also partly prefers different hydrophobic amino acids than human TAP in the carboxyterminal position. These species-dependent differences in specificity determined in vitro are shown to correlate with the epitope repertoire recognized in vivo. The quantitative model of binding specificity of murine TAP developed herein should be useful for interpreting epitope mapping and immunogenicity data obtained in humanized mouse models.
Genetic susceptibility to systemic lupus erythematosus (SLE) is well established, with the HLA class II DRB1 and DQB1 loci demonstrating the strongest association. However, HLA may also influence SLE through novel biologic mechanisms in addition to genetic transmission of risk alleles. Evidence for increased maternal–offspring HLA class II compatibility in SLE and differences in maternal versus paternal transmission rates (parent-of-origin effects) and nontransmission rates (noninherited maternal antigen [NIMA] effects) in other autoimmune diseases have been reported. Thus, we investigated maternal–offspring HLA compatibility, parent-of-origin effects, and NIMA effects at DRB1 in SLE.
The cohort comprised 707 SLE families and 188 independent healthy maternal–offspring pairs (total of 2,497 individuals). Family-based association tests were conducted to compare transmitted versus nontransmitted alleles (transmission disequilibrium test) and both maternally versus paternally transmitted (parent-of-origin) and nontransmitted alleles (using the chi-square test of heterogeneity). Analyses were stratified according to the sex of the offspring. Maternally affected offspring DRB1 compatibility in SLE families was compared with paternally affected offspring compatibility and with independent control maternal–offspring pairs (using Fisher’s test) and was restricted to male and nulligravid female offspring with SLE.
As expected, DRB1 was associated with SLE (P < 1 × 10−4). However, mothers of children with SLE had similar transmission and nontransmission frequencies for DRB1 alleles when compared with fathers, including those for the known SLE risk alleles HLA–DRB1*0301, *1501, and *0801. No association between maternal–offspring compatibility and SLE was observed.
Maternal–offspring HLA compatibility, parent-of-origin effects, and NIMA effects at DRB1 are unlikely to play a role in SLE.
Selective peptide transport by the transporter associated with antigen processing (TAP) represents one of the main candidate mechanisms that may regulate the presentation of antigenic peptides to HLA class I molecules. Because TAP-binding preferences may significant impact T-cell epitope selection, there is great interest in applying computational techniques to systematically discover these elements.
We describe TAP Hunter, a web-based computational system for predicting TAP-binding peptides. A novel encoding scheme, based on representations of TAP peptide fragments and composition effects, allows the identification of variable-length TAP ligands using SVM as the prediction engine. The system was rigorously trained and tested using 613 experimentally verified peptide sequences. The results showed that the system has good predictive ability with area under the receiver operating characteristics curve (AROC) ≥0.88. In addition, TAP Hunter is compared against several existing public available TAP predictors and has showed either superior or comparable performance.
TAP Hunter provides a reliable platform for predicting variable length peptides binding onto the TAP transporter. To facilitate the usage of TAP Hunter to the scientific community, a simple, flexible and user-friendly web-server is developed and freely available at http://datam.i2r.a-star.edu.sg/taphunter/.
OBJECTIVE—To perform an exploratory analysis of the relative contribution of single MHC genes to the pathogenesis of systemic lupus erythematosus (SLE) in a homogenous white population.
METHODS—MHC class II alleles and C4 allotypes were determined in 64 SLE patients and in ethnically matched controls. HLA-DR and DQ typing was performed by polymerase chain reaction amplification with sequence specific primers. C4 allotypes were determined by agarose gel electrophoresis.
RESULTS—The frequency of C4A*Q0 was significantly higher in patients than in controls (46.9% v 25.3%, p=0.002). HLA-DRB1, DQA1, and DQB1 alleles in the whole group of SLE patients were not significantly different from those of controls. On the other hand increase in DRB1*03 was observed in the group of patients with C4A*Q0, as compared with patients with other C4A allotypes (p=0.047). There was no significant correlation between severe and mild disease, as judged by the SLEDAI, and HLADR, DQ alleles and comparing the patients with C4A*Q0 with those with other C4A allotypes there was no significant difference regarding clinical manifestations.
CONCLUSION—The results are consistent with the argument that C4A deficiency contributes independently to susceptibility and the pathogenesis of SLE. C4A*Q0 in SLE patients in Iceland shows weaker linkage disequilibrium with DR3 genes than reported in most other white populations and emphasises the role of ethnicity.
Keywords: systemic lupus erythematosus; HLA; C4 allele; disease associations
OBJECTIVE--To determine if susceptibility to Behçet's disease (BD) is associated with polymorphism of HLA-DRB1, HLA-DQB1, DQB1, and TAP1 and TAP2 genes. METHODS--Fifty eight Spanish BD patients and 116 ethnically matched unrelated healthy subjects were typed at the HLA-DRB1 and HLA-DQB1 loci using polymerase chain reaction/sequence specific oligotyping (PCR/SSO). TAP1 and TAP2 alleles were assigned using amplification refractory mutation system-PCR. RESULTS--TAP1C was absent in BD patients, but was found in 12.1% of control subjects (pcorr < 0.05; relative risk = 0.06). Additionally, a linkage disequilibrium between HLA-DQB1*0501 and TAP2B was observed in BD patients (delta = 0.095, pcorr < 0.02), but not in the control group (delta = -0.0031, p > 0.05). CONCLUSIONS--The complete absence of TAP1C alleles in BD patients may indicate that TAP1 polymorphism is not without some significance in the development of BD. Furthermore, the existence of a linkage disequilibrium between HLA-DQB1*0501 and TAP2B in our patients suggests that the gene conferring susceptibility for BD is inherited as an extended haplotype in the population studied.
OBJECTIVE—The aim of this study was to investigate the relation between the polymorphism of large molecular weight proteasome (LMP) (LMP2-LMP7) and transporter associated with antigen processing (TAP) (TAP1-TAP2) genes and rheumatoid arthritis (RA).
METHODS—Sixty RA patients and 102 ethnically matched unrelated healthy subjects were typed for LMP, TAP, and disease associated HLA-DRB1 alleles by using a new strategy based on polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) with amplification created restriction sites.
RESULTS—The polymorphism of LMP (LMP2-LMP7) and TAP (TAP1-TAP2) genes was examined in shared epitope positive and negative RA patients and controls. No significant differences in the LMP or TAP allele frequencies were observed between the total patient and control groups or the patients and controls positive or negative for the shared epitope.
CONCLUSION—The data suggest that the polymorphisms of LMP and TAP genes do not have an important influence in the pathogenesis of RA, although larger studies will be needed to provide more conclusive evidence on the role of these genes in RA. A new, highly reliable strategy for typing LMP alleles is also described.
Keywords: large molecular weight proteasome; transporter assoicated with antigen processing; rheumatoid arthritis
OBJECTIVE--To assess the associations of HLA class II antigens with systemic lupus erythematosus (SLE) in black South Africans. METHODS--HLA-DRB1 genotype frequencies assigned by polymerase chain reaction (PCR) amplification and sequence specific oligonucleotide probes were compared between 49 black SLE patients from Baragwanath Hospital and 87 ethnically matched controls. HLA-DQA1 and -DQB1 genotypes were also assigned in 45 of the SLE patients and 74 controls by PCR using sequence specific primers. RESULTS--HLA-DRB1*02 was increased in the patients compared with controls (odds ratio = 3.67; 95% confidence interval = 1.49 to 9.02; p < 0.005). HLA-DQB1*0201 was not associated with development of the disease itself, but was associated with the presence of Ro antibodies (p = 0.01). HLA-DRB1*03 was less strongly linked to DQB1*02 in this population than in white populations and was not associated with SLE. CONCLUSIONS--In black South Africans there is evidence for a locus on DR2 haplotypes contributing to SLE. Another gene, possibly HLA-DQB1*02, not linked to DR2 is involved in the subset of patients exhibiting Ro antibodies.
The transporter associated with antigen processing (TAP) is a critical component of the major histocompatibility complex (MHC) class I antigen processing and presentation pathway. TAP transports antigenic peptides into the endoplasmic reticulum where it loads them into the binding groove of MHC class I molecules. Because peptides must first be transported by TAP in order to be presented on MHC class I, TAP binding preferences should impact significantly on T-cell epitope selection.
PREDTAP is a computational system that predicts peptide binding to human TAP. It uses artificial neural networks and hidden Markov models as predictive engines. Extensive testing was performed to valid the prediction models. The results showed that PREDTAP was both sensitive and specific and had good predictive ability (area under the receiver operating characteristic curve Aroc>0.85).
PREDTAP can be integrated with prediction systems for MHC class I binding peptides for improved performance of in silico prediction of T-cell epitopes. PREDTAP is available for public use at .
Systemic lupus erythematosus (SLE) is a chronic inflammatory disease typically diagnosed by a combination of physical findings and clinical laboratory testing. Several decades ago, the diagnosis of lupus included the lupus erythematosus (LE) cell assay. SLE is associated with pleuropulmonary manifestations in well over 50% of cases. Although pleural effusion is common but very rarely is the initial manifestation of disease. There are very few reports of SLE diagnosed in a cytopathology laboratory. We report an unusual case of SLE in a 16-year-old female who presented with acute shortness of breath, fever and cough. Her chest radiograph showed bilateral pleural effusion. This effusion was tapped and sent to the cytopathology laboratory. The cytological examination of the pleural fluid revealed numerous LE cells and led to the diagnosis of SLE. Autoimmune serology techniques such as anti-nuclear antibody staining have replaced the LE cell assay. However, as presented in this report and found in a review of the literature, the in vivo finding of LE cells by cytopathology can provide an important clue to the diagnosis of SLE, especially when associated with an uncommon presentation.
LE cells; pleural effusion; systemic lupus erythematosus; tart cells
Although systemic lupus erythematosus (SLE) is a multigenic autoimmune disorder, HLA-D is the most dominant genetic susceptibility locus. This study was undertaken to investigate the hypothesis that microbial peptides bind HLA-DR3 and activate T cells reactive with lupus autoantigens. Using HLA-DR3 transgenic mice and lupus-associated autoantigen SmD protein, SmD79–93 was identified to contain a dominant HLA-DR3 restricted T cell epitope. This T cell epitope was characterized by using a T-T hybridoma, C1P2, generated from SmD immunized HLA-DR3 transgenic mouse. By pattern search analysis, 20 putative mimicry peptides (P2–P21) of SmD79–93, from microbial and human origin were identified. C1P2 cells responded to SmD, SmD79–93 and a peptide (P20) from Vibro cholerae. Immunization of HLA-DR3 mice with P20 induced T cell responses and IgG antibodies to SmD that were not cross-reactive with the immunogen. A T-T hybridoma, P20P1, generated from P20 immunized mice, not only responded to P20 and SmD79–93, but also to peptides from Streptococccus agalactiae (P17) and human-La related protein (P11). These three T cell mimics (P20, P11 and P17) induced diverse and different autoantibody response profiles. Our data demonstrates for the first time molecular mimicry at T cell epitope level between lupus-associated autoantigen SmD and microbial peptides. Considering distinct autoreactive T cell clones, activated by different microbial peptides, molecular mimicry at T cell epitope level can be an important pathway for the activation of autoreactive T cells resulting in the production of autoantibodies. In addition, the novel findings reported herein may have significant implications in the pathogenesis of SLE.
Epitope; HLA-DR3; Mice; Molecular Mimicry; Systemic Lupus; Erythematosus; SmD
The ATP-binding cassette family of transporter proteins, subfamily B (MDR/TAP), member 1 (ABCB1) (P-glycoprotein) transporter is a key component of the blood–brain barrier. Many antidepressants are subject to ABCB1 efflux. Functional polymorphisms of ABCB1 may influence central nervous system bioavailability of antidepressants subject to efflux. Single-nucleotide polymorphisms (SNPs) at rs1045642 (C3435T) of ABCB1 have been associated with efflux pump efficiency. This may explain part of the interindividual variation in antidepressant dose needed to remit. Individuals (N=113) with DSM-IV (Diagnostic and Statistical Manual of Mental Disorders, Fourth Edition) major depressive disorder (MDD) were treated with escitalopram (ESC) or venlafaxine (VEN) over 8 weeks. The17-item Hamilton Depression Rating Scale was assessed serially, blind to genotype. SNP rs1045642 of ABCB1 along with two SNPs previously reported to be in linkage disequilibrium with it (rs2032582 and rs1128503) were genotyped. Demographic features, clinical features, P450 metabolizer status and 5-HTTLPR (serotonin-transporter-linked promoter region) genotype were controlled for. Carriers of rs1045642 TT needed on average 11 mg of ESC to remit, whereas TC and CC carriers required 24 and 19 mg, respectively (P=0.0001). This equates to a 2.0- (95% confidence interval=1.5–3.4; P<0.001) fold greater ESC dose needed to remit for C carriers compared with TT carriers at rs1045642. Of VEN-treated subjects carrying TT genotype at rs1045642, 73.3% remitted compared with 12.5% for CC genotype (odds ratio=6.69; 95% confidence interval=1.72–25.9, P=0.006). These data suggest that antidepressant dose needed to remit can be predicted by an ABCB1 SNP. This has the potential clinical translation implications for dose selection and remission from MDD.
ABCB1; pharmacogenetics; antidepressant; major depression; blood–brain barrier; P-glycoprotein
The human cytomegalovirus (HCMV) protein US6 inhibits the transporter associated with antigen processing (TAP). Since TAP transports antigenic peptides into the endoplasmic reticulum for binding to major histocompatibility class I molecules, inhibition of the transporter by HCMV US6 impairs the presentation of viral antigens to cytotoxic T lymphocytes. HCMV US6 inhibits ATP binding by TAP, hence depriving TAP of the energy source it requires for peptide translocation, yet the molecular basis for the interaction between US6 and TAP is poorly understood. In this study we demonstrate that residues 89 to 108 of the HCMV US6 luminal domain are required for TAP inhibition, whereas sequences that flank this region stabilize the binding of the viral protein to TAP. In parallel, we demonstrate that chimpanzee cytomegalovirus (CCMV) US6 binds, but does not inhibit, human TAP. The sequence of CCMV US6 differs from that of HCMV US6 in the region corresponding to residues 89 to 108 of the HCMV protein. The substitution of this region of CCMV US6 with the corresponding residues from HCMV US6 generates a chimeric protein that inhibits human TAP and provides further evidence for the pivotal role of residues 89 to 108 of HCMV US6 in the inhibition of TAP. On the basis of these observations, we propose that there is a hierarchy of interactions between HCMV US6 and TAP, in which residues 89 to 108 of HCMV US6 interact with and inhibit TAP, whereas other parts of the viral protein also bind to TAP and stabilize this inhibitory interaction.
Dysfunction in various parts of immune defence, such as immune response, immune complex clearance, and inflammation, has an impact on pathogenesis in systemic lupus erythematosus (SLE). We hypothesised that combinations of common variants of genes involved in these immune functions are associated with susceptibility to SLE. The following variants were analysed: HLA DR3, HLA DQ2, C4AQ0, Fcγ receptor IIa (FcγRIIa) genotype R/R, Fcγ receptor IIIa (FcRγIIIa) genotype F/F, mannan-binding lectin (MBL) genotype conferring a low serum concentration of MBL (MBL-low), and interleukin-1 receptor antagonist (IL-1Ra) genotype 2/2. Polymorphisms were analysed in 143 Caucasian patients with SLE and 200 healthy controls. HLA DR3 in SLE patients was in 90% part of the haplotype HLA DR3-DQ2-C4AQ0, which was strongly associated with SLE (odds ratio [OR] 2.8, 95% CI 1.7–4.5). Analysis of combinations of gene variants revealed that the strong association with SLE for HLA DR3-DQ2-C4AQ0 remained after combination with FcγRIIa R/R, FcγRIIIa F/F, and MBL-low (OR>2). Furthermore, the combination of the FcγRIIa R/R and IL-1Ra 2/2 genotypes yielded a strong correlation with SLE (OR 11.8, 95% CI 1.5–95.4). This study demonstrates that certain combinations of gene variants may increase susceptibility to SLE, suggesting this approach for future studies. It also confirms earlier findings regarding the HLA DR3-DQ2-C4AQ0 haplotype.
Fcγ receptor; HLA; interleukin-1 receptor antagonist; mannan-binding lectin; systemic lupus erythematosus
Systemic lupus erythematosus (SLE) is a multi-system disorder resulting from interaction of susceptibility genes and environmental factors. SLE has protean clinical presentations at the initial diagnosis and relapses. SLE-related autoantibodies have unique patterns of diversification to linked proteins such as the snRNP particle and the diversification takes years before clinical diagnosis. There are both clinical and experimental evidence to indicate that separate genes contribute to autoimmunity and end organ damage and these genes are independent and interactive. Among the numerous susceptibility genes, HLA-D complex is dominant. Results from the authors’ laboratories led us to postulate a unified hypothesis for SLE pathogenesis. This hypothesis states that SLE-autoantibodies are initiated by environmental T cell epitope mimics of the SLE-related autoantigens in hosts with susceptible HLA-D alleles. These autoantibodies diversify over a period of years due the accumulation of cross-reactive T cells. This process ultimately leads to the generation of organ specific autoantibodies and autoreactive effector T cells due to the polyreactive nature of T and B cell receptors from hosts with susceptibility genes to end organ damage, resulting in protean clinical presentations. This hypothesis accounts for most of the features unique to SLE and has clinical implications as to how patients should be treated.
Systemic lupus erythematosus Pathogenesis; Epitope spreading; Cross-reactive T and B cell epitopes; HLA-D and end organ damage
We hypothesize that over-expression of transporters associated with antigen processing (TAP1 and TAP2), components of the major histocompatibility complex (MHC) class I antigen-processing pathway, enhances antigen-specific cytotoxic activity in response to viral infection. An expression system using recombinant vaccinia virus (VV) was used to over-express human TAP1 and TAP2 (VV-hTAP1,2) in normal mice. Mice coinfected with either vesicular stomatitis virus plus VV-hTAP1,2 or Sendai virus plus VV-hTAP1,2 increased cytotoxic lymphocyte (CTL) activity by at least 4-fold when compared to coinfections with a control vector, VV encoding the plasmid PJS-5. Coinfections with VV-hTAP1,2 increased virus-specific CTL precursors compared to control infections without VV-hTAP1,2. In an animal model of lethal viral challenge after vaccination, VV-hTAP1,2 provided protection against a lethal challenge of VV at doses 100-fold lower than control vector alone. Mechanistically, the total MHC class I antigen surface expression and the cross-presentation mechanism in spleen-derived dendritic cells was augmented by over-expression of TAP. Furthermore, VV-hTAP1,2 increases splenic TAP transport activity and endogenous antigen processing, thus rendering infected targets more susceptible to CTL recognition and subsequent killing. This is the first demonstration that over-expression of a component of the antigen-processing machinery increases endogenous antigen presentation and dendritic cell cross-presentation of exogenous antigens and may provide a novel and general approach for increasing immune responses against pathogens at low doses of vaccine inocula.
The development of protective vaccines against infectious diseases such as AIDS, SARS, and West Nile virus has become a societal priority but remains a scientific challenge. In recent years, the threat of bioterrorism agents such as anthrax and smallpox has heightened the need for the rapid development of effective new vaccines. One of the major stumbling blocks to the implementation of any vaccine is the toxic side effects on the vaccine candidate. For example, a significant number of doses of a new vaccine against smallpox have been commissioned, but approximately 20% of the individuals targeted to be inoculated will suffer toxicity due to vaccination. Furthermore, an additional difficulty in the production of vaccines is the creation of sufficient doses to vaccinate a large population. The authors have identified a novel approach that appears to address these issues. They demonstrate that the inclusion, in low doses of vaccines, of a normal component of the antigen-processing pathway, the transporter associated with antigen processing (TAP), confers protective immunity against lethal viral loads during viral challenges. This new paradigm is shown to be applicable to many viruses, including poxviruses, and could significantly advance the creation of new vaccines and improve those that already exist.
Antigen presentation by major histocompatibility complex (MHC) class I molecules requires peptide supply by the transporters associated with antigen processing (TAPs), which select substrates in a species- and, in the rat, allele-specific manner. Conflicts between TAPs and MHC preferences for COOH-terminal peptide residues in rodent cells strongly reduce the efficiency of MHC class I antigen presentation. Although human TAP is relatively permissive, some peptide ligands for human histocompatibility leukocyte antigen class I molecules are known to possess very low TAP affinities; the significance of these in vitro findings for cellular antigen presentation is not known. We studied two naturally immunodominant viral epitopes presented by HLA-A2 that display very low affinities for human TAP. Low TAP affinities preclude minimal epitope access to the endoplasmic reticulum (ER) and assembly with HLA-A2 in vitro, as well as presentation by minigene-expressing cells to cytotoxic T lymphocytes. However, NH2-terminally but not COOH-terminally extended epitope variants with higher TAP affinities assemble in vitro and are presented to cytotoxic T lymphocytes with high efficiency. Thus, human TAP can influence epitope selection and restrict access to the ER to epitope precursors. Analysis of TAP affinities of a panel of viral epitopes suggests that TAP selection of precursors may be a common phenomenon for HLA-A2–presented epitopes. We also analyzed HLA-A2–eluted peptides from minigene-expressing cells and show that an NH2-terminally extended variant with low A2 binding affinity undergoes ER processing, whereas another with high affinity is presented unmodified. Therefore, the previously reported aminopeptidase activity in the ER can also act on TAP-translocated peptides.
antigen presentation; ABC transporters; immunodominant epitopes; HLA class I; aminopeptidases