Characterization of a novel MICA allele,MICA*012:05, by cloning and sequencing

A new MICA allelic variant, MICA*012:05, has been identified in a Chinese Mongolian population. Following polymerase chain reaction–sequence‐based typing (PCR‐SBT), this new allele was further confirmed by cloning and sequencing. MICA*012:05 was linked to an HLA‐A*24‐C*01‐B*55:02‐DRB1*09 haplotype. MICA*012:05 differs from MICA*012:01 by a single synonymous C to T substitution at nucleotide position 269 in exon 3.     

MICA Gene Deletion in 3411 DNA Samples from Five Distinct Populations in Mainland China and Lack of Association with Nasopharyngeal Carcinoma (NPC) in a Southern Chinese Han population

Deletion of major histocompatibility complex class I chain‐related genes A (MICA*Del) was investigated in 3,411 DNA samples from two southern Chinese Han populations (Hunan Han, HNH; Guangdong Han, GDH), two northern Chinese populations (Inner Mongolia Han, IMH; Inner Mongolia Mongol, IMM) and one southeastern Chinese Han population (Fujian Han, FJH) using an in‐house polymerase chain reaction‐sequence specific priming (PCR‐SSP) assay, which enables direct discrimination between heterozygote and homozygote for MICA*Del. MICA*Del showed a frequency ranging from 0.8% in FJH to 5.7% in IMM (P_corrected < 0.05), indicating northward increase in frequency of MICA*Del in Chinese populations. In contrast to the association reported recently in a Taiwan Chinese population and a Malaysian Chinese cohort, MICA*Del distribution did not differ between 1,120 patients with nasopharyngeal carcinoma (NPC) and 1,483 normal controls in the HNH population (1.03% in NPC cases vs 1.18% in the controls, OR (95% CI) = 0.87 (0.51‐1.47), p = 0.69). Further gender‐stratified analysis also failed to disclose any male‐specific association reported in a Taiwan Chinese population. Multi‐locus typing of the 94 samples carrying MICA*Del revealed two new haplotypes, HLA‐A*11:01‐B*13:01‐MICA*Del‐MICB*009N‐DRB1*04:06 and HLA‐B*35:01‐MICA*Del‐MICB*009N‐DRB1*15:01, in addition to HLA‐B*48‐MICA*Del. Unexpectedly, two samples with MICA*Del in the HNH population were each consistently found to have two distinct MICA alleles, indicating the existence of two MICA gene copies on certain HLA haplotypes. Based on the results from a sizeable case‐control study, our data suggest that there is no association between MICA*Del and NPC in the southern Chinese Han population.     

Characterization of a novel HLA‐B*39:01:01‐related allele,HLA‐B*39:130, by cloning and phasing

A novel HLA‐B*39:01:01‐related variant, HLA‐B*39:130, has been identified in a normal individual of Han ethnicity in Hunan province, southern China. Following Sanger polymerase chain reaction–sequence‐based typing (PCR‐SBT), this new allele was further confirmed by cloning, phasing and sequencing. Aligned with HLA‐B*39:01:01, HLA‐B*39:130 has a nonsynonymous thymine substitution at nucleotide position 94 in exon 4, resulting in amino acid change from threonine to isoleucine at codon 214 (ACA→ATA) of the mature HLA‐BmRNA molecule.     

MICA*A4 protects against ulcerative colitis,whereas MICA*A5.1 is associated with abscess formation and age of onset

Ulcerative colitis (UC) is one of the two major forms of inflammatory bowel disease, the aetiology of which remains unknown. Several studies have demonstrated the genetic basis of disease, identifying more than 130 susceptibility loci. The major histocompatibility complex class I chain‐related gene A (MICA) is a useful candidate to be involved in UC pathogenesis, because it could be important in recognizing the integrity of the epithelial cell and its response to stress. The aim of this study was to analyse the relationship between polymorphisms in the transmembrane domain of MICA and susceptibility to develop UC. A total of 340 patients with UC and 636 healthy controls were genotyped for MICA transmembrane polymorphism using a polymerase chain reaction (PCR) combined with fluorescent technology. Different MICA alleles were determined depending on the PCR product size. The allele MICA*A4 was less frequent in patients than in controls (P = 0·003; OR = 0·643), and this protective role is higher when it forms haplotype with B*27 (P = 0·002; OR = 0·294). The haplotype HLA‐B*52/MICA*A6 was also associated with UC [P = 0·001; odds ratio (OR) = 2·914]. No other alleles, genotypes or haplotypes were related with UC risk. Moreover, MICA*A5.1 is associated independently with abscesses (P = 0·002; OR = 3·096) and its frequency is lower in patients diagnosed between ages 17 and 40 years (P = 0·007; OR = 0·633), meaning an extreme age on onset. No association with location, extra‐intestinal manifestations or need for surgery was found.     

MICA genetic polymorphism and HLA-A,C,B,MICA and DRB1 haplotypic variation in a southern Chinese Han population: identification of two new MICA alleles, MICA*060 and MICA*062

In this study, 201 healthy, unrelated Han subjects in Hunan province, southern China, were investigated by sequence-based typing (SBT) for the allelic variation of the human major histocompatibility complex (MHC) class I chain-related gene A (MICA). Nineteen MICA alleles were observed, among which MICA*008:01 predominated with gene frequency of 30.35%. There was significant linkage disequilibrium (LD) of MICA*012:01 with HLA-B*54 and HLA-B*55, which was not observed in a northern Chinese Han population. Haplotype HLA-A*11-C*07-B60-MICA*008:01 (9.16%) was highly specific to this southern Chinese Han population. The most common five-locus haplotype in this population was HLA-A*02-C*01-B*46-MICA*010-DRB1*09 (8.73%). A new MICA allele, MICA*060, was identified on an HLA-A*02-C*01-B*55:02-DRB1*14 haplotype through extended family analysis. MICA*060 has probably arisen from MICA*012:01. Another new MICA allele, MICA*062, was identified by screening 1432 subjects using polymerase chain reaction-sequence-specific priming technology. MICA*062 has probably derived from MICA*010. Of particular interest is that MICA*062 was carried on an HLA-C*08-B*48:01-DRB1*14 haplotypic segment, as HLA-B*48 has been consistently shown to be primarily linked to MICA gene deletion in east Asian populations. Our results provide new insight into MICA genetic polymorphism in human populations. The findings reported here are of importance for future studies on the potential role of MICA in allogeneic organ transplantation and disease association in populations of Chinese ancestry.     

The association between MICA/MICB polymorphism and respiratory syncytial virus infection in children

MICA/MICB gene polymorphisms are related to several cancers and infectious diseases, but there are no reports on the association between MICA/MICB gene polymorphisms and respiratory syncytial virus (RSV) infection. To clarify the association between MICA/MICB gene polymorphisms and infection of RSV in children, we collected fresh blood samples from paediatric patients with and without pneumonia after RSV infection. The MICA/MICB alleles were characterized by PCR sequence‐specific primers (PCR‐SSP) and PCR sequence‐based genotyping (PCR‐SBT), and then, the frequency of the MICA/MICB alleles and haplotypes was calculated. The results showed that the frequencies of MICA*002:01 and MICA‐A9 in RSV‐infected patients were significantly lower than in controls (9% vs. 20%, pc = 0.04). The allele frequency of MICA*002:01 in pneumonia patients (8%) and nonpneumonia patients (9%) was significantly lower than in controls (20%, pc = 0.02). MICA*002:01‐MICB*008(Δrel = 0.616), MICA*009‐MICB*016 (Δrel = 0.506), and MICA*045‐MICB*014 (Δrel = 0.700) showed linkage disequilibrium in patients infected with RSV. The haplotype frequency of MICA*002:01‐MICB*005:02 in RSV‐infected patients was significantly lower than in controls (10% vs. 16%, pc = 0.033). In conclusion, allele MICA*002:01/A9 and haplotype MICA*002:01‐MICB*005:02 were negatively associated with RSV respiratory tract infections.     

MICA,MICB Polymorphisms and Linkage Disequilibrium with HLA‐B in a Chinese Mongolian Population

In this study, polymorphisms of major histocompatibility complex class I chain‐related genes A and B (MICA and MICB) and human leucocyte antigen (HLA)‐B gene were investigated for 158 unrelated Chinese Mongolian subjects recruited from central Inner Mongolia Autonomous Region, northern China, by polymerase chain reaction–sequence‐based typing (PCR‐SBT) and cloning. Collectively, 79 alleles, including 20 MICA, 12 MICB and 47 HLA‐B alleles, were identified. MICA*008:01 (21.2%), MICB*005:02 (48.1%) and HLA‐B*51:01 (7.91%) were the most common alleles. Significant global linkage disequilibrium (LD) was detected between HLA‐B and MICA, HLA‐B and MICB, and MICA and MICB loci (all P < 0.000001). The most frequent haplotypes were HLA‐B*51:01‐MICA*009:01 (7.28%), HLA‐B*58:01‐MICB*008 (6.96%), MICA*010‐MICB*005:02 (13.92%) and HLA‐B*58:01‐MICA*002:01‐MICB*008 (6.96%). HLA‐B‐MICA haplotypes such as HLA‐B*50:01‐MICA*009:02 were associated with single MICB allele. Some HLA‐B‐MICA haplotypes were associated with multiple MICB alleles, including HLA‐B*51:01‐MICA*009:01. One novel MICB allele, MICB*031, was identified, which has possibly arisen from MICB*002:01 through single mutation event. We also confirmed the existence of a recently recognized MICA allele, MICA*073, whose ethnic origin has not been previously described. Genotype distributions at MICA, MICB and HLA‐B were consistent with a neutrality model. Our results provide new insight into MIC genetic polymorphisms in Chinese ethnic groups. Findings shown here are important from an anthropologic perspective and will inform future studies of the potential role of MIC genes in allogeneic organ transplantation and HLA‐linked disease association in populations of related ancestry.     

Characterization of a novel MICA allele,MICA*047

We report the identification of a novel MICA allele, MICA*047. It was initially detected because of an unusual hybridization pattern with sequence-specific oligonucleotides (SSOP) in a normal subject of Caucasian origin. Cloning and sequencing of both strands, and comparison of the sequence with previously defined MICA alleles, revealed that the new allele is similar to MICA*041 except for one nucleotide substitution at position 811 (C-->G). It appears that this new allele could have been generated by an interallelic sequence exchange between MICA*011 and MICA*0411.     

Identification of a new MICA allele,MICA*047

The MHC Class I related (MIC) gene family has been shown to be very polymorphic with 46 different MICA alleles being officially named by the WHO Nomenclature Committee for factors of the HLA system to date. We have identified a novel MICA allele, MICA*047, in a Coya American Indian individual from the Jujuy province of north-western Argentina. The novel MICA*047 allele differs from the MICA*030 allele by a single non-synonymous substitution in exon 2, condon 26 GTA-->GGA, Valine to Glycine1.     

Association of MICA and HLA‐B alleles with leprosy in two endemic populations in Brazil

Leprosy is a prevalent disease in Brazil, which ranks as the country with the second highest number of cases in the world. The disease manifests in a spectrum of forms, and genetic differences in the host can help to elucidate the immunopathogenesis. For a better understanding of MICA association with leprosy, we performed a case–control and a family‐based study in two endemic populations in Brazil. MICA and HLA‐B alleles were evaluated in 409 leprosy patients and in 419 healthy contacts by PCR‐SSOP‐Luminex‐based technology. In the familial study, analysis of 46 families was completed by direct sequencing of all exons and 3′/5′untranslated regions, using the Ilumina MiSeq platform. All data were collected between 2006 and 2009. Statistical analysis was performed using the Chi‐square or Fisher's exact test together with a multivariate analysis. Family‐based association was assessed by transmission disequilibrium test (TDT) software FBAT 2.0.4. We found associations between the haplotype MICA*002‐HLA‐B*35 with leprosy in both the per se and the multibacillary (MB) forms when compared to healthy contacts. The MICA allele *008 was associated with the clinical forms of paucibacillary (PB). Additionally, MICA*029 was associated with the clinical forms of MB. The association of MICA*029 allele (MICA‐A4 variant) with the susceptibility to the MB form suggests this variant for the transmembrane domain of the MICA molecule may be a risk factor for leprosy. Two MICA and nine HLA‐B variants were found associated with leprosy per se in the Colônia do Prata population. Linkage disequilibrium analysis revealed perfect linkage disequilibrium (LD) between HLA‐B markers rs2596498 and rs2507992, and high LD (R² = .92) between these and the marker rs2442718. This familial study demonstrates that MICA association signals are not independent from those observed for HLA‐B. Our findings contribute the knowledge pool of the immunogenetics of Hansen's disease and reveals a new association of the MICA*029 allele.     

MICA polymorphism in a sample of the São Paulo population,Brazil

The major histocompatibility complex (MHC) class I chain‐related A (MICA) gene, located near HLA‐B, codes for protein products with structural similarities to those of classical MHC class I genes, but which neither bind β₂‐microglobulin nor present peptide. Expressed predominantly on gastrointestinal and tumour epithelial cells, they are stress‐induced and interact with C‐type lectin like receptor (NKG2D) on γδ, αβ CD8+ T cells and natural killer (NK) cells. MICA is highly polymorphic, with 54 extracellular allelic sequences described. We typed 200 healthy subjects in a sample of the São Paulo population by extended polymerase chain reaction–sequence‐specific primers (PCR‐SSP) to characterize the MICA polymorphism and analysed MICA/HLA‐B linkage disequilibrium. The MICA*008 group (g) was predominant (47%), with several HLA‐B associations. Rare combinations MICA*008g‐HLA‐B37, MICA*008g‐B72 and MICA*010‐HLA‐B52 were detected. Given the extent of this polymorphism and its possible relevance for disease association, we determined MICA and HLA‐B alleles in 33 Behçet's patients, in an attempt to clarify the associated genetic marker. Our results showed an increase of MICA*006, but not MICA*009, in the patient group (6/33) compared with controls (3/200) (18.2% vs. 1.5%; P_c = 0.005). Both alleles were always in association with HLA‐B51, suggesting that HLA‐B is indeed the primary susceptibility locus (P = 0.00008) and that MICA*006 may be an additional risk factor.     

Two novel MICA alleles, MICA*054 and MICA*056

We report the identification of two novel major histocompatibility complex (MHC) class I-related chain A (MICA) alleles. MICA*054 has a nucleotide substitution of A to G at position 871 (codon 268), encoding an amino acid change of serine to glycine in the α-3 domain. MICA*056 has a nucleotide substitution at position 758 of G to C resulting in the substitution of tryptophan for serine at codon 230, also in the α-3 domain.     

HLA class I allele lineages and haplotype frequencies in Arabs of the United Arab Emirates

The high degree of polymorphism of the HLA system provides suitable genetic markers to study the diversity and migration of different world populations and is beneficial for forensic identification, anthropology, transplantation and disease associations. Although the United Arab Emirates (UAE) population of about nine million people is heterogeneous, information is limited for the HLA class I allele and haplotype frequencies of the Bedouin ethnic group. We performed low‐resolution PCR‐SSP genotyping of three HLA class I loci at HLA‐A, ‐B and ‐C for 95 unrelated healthy Bedouins from the cities of Al Ain and Abu Dhabi in the UAE. A total of 54 HLA allele lineages were detected; the most frequent low‐resolution allele lineages at each HLA locus were A*02 (0.268), B*51 (0.163) and C*07 (0.216). The inferred estimates for the two most frequent HLA‐A and HLA‐B haplotypes were HLA‐A*02 ~ HLA‐B*50 (0.070) and HLA‐A*02 ~ HLA‐B*51 (0.051), and the most frequent 3‐locus haplotype was HLA‐A*02 ~ HLA‐B*50 ~ HLA‐C*06 (0.068). The HLA allele lineage frequencies of the UAE Arabs were compared to those previously reported for 70 other world populations, and a strong genetic similarity was detected between the UAE Arabs and the Saudi Arabians from the west with evidence of a limited gene flow between the UAE Arabs and Pakistani across the Gulf from the east, and the UAE Arabs and Omani from the south of the Gulf Peninsula.     

强直性脊柱炎患者MICA基因第2、3和4外显子的多态性及其与HLA-B抗原的连锁不平衡

目的探讨皖籍汉族人群MICA基因（major histocompatibility complex class Ⅰchain-related gene A，MICA）第2、3、4外显子的多态性，及其与HLA-B抗原的连锁不平衡在强直性脊柱炎（ankylosing spondylitis，AS）发病中的作用。方法采用聚合酶链反应-序列特异性寡核苷酸探针杂交（polymerase chain reactionsequence-specific oligonucleotide probing，PCR-SS0）技术对56例AS患者和112名正常对照人群进行MICA基因第2、3、4外显子的多态性和HLA-B抗原的检测。结果AS患者和正常对照人群的MICA等位基因分布均以MICA＊008占优势，频率分别为32．14％和30．36％。两组人群MICA＊007等位基因的分布差异有统计学意义（X^2=10．18，P〈0．05，RR=2．50）。单倍型分析显示，AS患者和正常对照人群的MICA等位基因均显示出与多个HLA-B位点的连锁不平衡现象，两组间差异有统计学意义的单倍型为MICA＊007-B27（X^2=18．46，P〈0．05，RR=7．47）。分层分析结果显示，HLA-B27阳性与AS的相关性有统计学意义（P〈0．05），但MICA＊007基因与AS的相关性无统计学意义（P〉0．05）。结论AS患者中MICA＊007等位基因频率的显著升高可能源于MICA基因与HLA-B位点间的广泛连锁不平衡。     

Characterization of a novel allelic variant in HLA‐B*40 lineage,HLA‐B*40:298:02, by cloning and sequencing

A novel allelic variant in HLA‐B*40 lineage, HLA‐B*40:298:02, has been identified in an individual of Han ethnicity afflicted with nasopharyngeal carcinoma in Hunan province, southern China. Following polymerase chain reaction–Sanger sequence‐based typing (PCR–SBT), this new variant was further confirmed by two distinct strategies of cloning and sequencing. HLA‐B*40:298:02 differs from HLA‐B*40:298:01 by a single synonymous cytosine substitution at nucleotide position 26 (T→C) in exon 3, which corresponds to codon 99 of the mature HLA‐B mRNA molecule. This new allele has an estimated frequency of 0.0002, in about 2,500 sequence‐based typed subjects from the same population.     

Characterization of a new HLA‐A allele,HLA‐A*02:07:08, by cloning and sequencing

In this report, we present a novel HLA‐A*02:07 allele, HLA‐A*02:07:08. HLA‐A*02:07:08 was identified in an individual of Han ethnicity in Hunan province, southern China. Following polymerase chain reaction‐sequence‐based typing (PCR‐SBT), this new allele was further confirmed by cloning and sequencing. HLA‐A*02:07:08 differs from HLA‐A*02:07:01 by a single synonymous C to T substitution at nucleotide position 131 in exon 3.     

Codominant expression of the polymorphic MICA alloantigens encoded by genes in the HLA region

The HLA‐related, polymorphic MHC class I‐related chain A (MICA) gene encodes a 383‐amino acid polypeptide, with three extracellular domains (α1, α2 and α3), a transmembrane region and a cytoplasmic tail. We have previously shown that freshly isolated endothelial cells, fibroblasts, keratinocytes and monocytes express MICA, while peripheral blood CD4⁺, CD8⁺ or CD19⁺ lymphocytes do not. This polymorphic MICA molecule is a target for specific alloantibodies in sera from kidney, heart and lung transplant recipients, although its possible role during graft rejection remains to be demonstrated. In this study we investigated whether there is codominance in the expression of MICA. We isolated RNA from a heterozygous cell line (HCT116), previously shown by sequencing‐based typing to be MICA*001/MICA*00902, as well as 12 clones derived from it. Thereafter, we retrotranscribed the RNA into cDNA, and performed a molecular typing using MICA‐sequence specific oligonucleotides (SSOP). Using this approach, we detected the RNA encoding MICA*001 and MICA*00902 in all the clones and in the parental cell line, indicating that MICA is codominantly expressed. This codominant expression was further confirmed by cloning and sequencing plasmids encoding these two alleles produced from the same HCT116 RNA preparation. We also produced the two recombinant MICA proteins (MICA*001 and MICA*00902). They reacted with rabbit anti‐MICA polyclonal antibodies by ELISA and Western blot, indicating that the plasmids carrying the cDNA inserts probably encode functional MICA proteins. This strongly suggests that, like the HLA class I and class II proteins, MICA is codominantly expressed. The codominant expression of the polymorphic, HLA‐like MICA alloantigens may have implications for the immune response elicited by the allograft in organ transplantation.     

Identification of three MICA alleles in the genotype of a patient with chronic lymphocytic leukemia

Major histocompatibility complex (MHC) class I-related chain A gene (MICA) sequence-based genotyping (SBT) was attempted on a peripheral blood sample collected from a patient evaluated for hematopoietic stem cell retransplant. The electropherogram pattern of MICA SBT indicated the possibility of carrying more than two MICA alleles. Subsequent cloning and sequencing of the polymerase chain reaction products revealed the presence of three distinct MICA alleles: MICA*008:01/:04 (A5.1), MICA*007:01(A4), and MICA*002:01 (A9) in the genotype of this patient. The origin of the third extra MICA allele could not be determined and would require MICA genotyping information from other family members, which is unavailable.     

MHC class I chain-like gene A (MICA) and its associations with inflammatory bowel disease and peripheral arthropathy.

MHC class I chain gene A (MICA) is a non-classical Class I gene which is expressed on the surface of epithelia without beta 2-microglobulin. The gene is found in the major histocompatibility complex (MHC) in tight linkage disequilibrium with human leucocyte antigen-B (HLA-B). Its precise function is unknown, but it interacts with gamma delta T cells of the intestinal immune system. This region of the MHC has been implicated in inflammatory bowel disease (IBD) pathogenesis by recent association mapping studies and this study was performed to examine the prevalence of MICA gene polymorphisms in IBD, in particular in type 2 peripheral arthropathy (PeA), which also has a strong HLA-B association. An assessment of the prevalence of MICA polymorphisms in IBD was made. Blood from 50 ulcerative colitis (UC) and 50 Crohn's disease controls was taken and MICA status determined using allele-specific PCR for 16 known alleles of MICA. A further 91 UC patients were recruited to confirm the results of this stage, and then the polymorphisms were studied in 52 type 1 and 45 type 2 PeA patients. The MICA status of these groups was compared with 118 blood and organ donor controls with appropriate correction for multiple comparisons. UC overall was associated with possession of MICA*007 in 32% compared to 11% of controls (P(c) = 0.017). This association was confirmed in a second cohort of 91 patients (23% versus 11%, P = 0.02). These were independent of HLA class I status. Type 2 IBD PeA was associated with MICA*008 in 98% compared to 73% of controls (P = 0.0001). MICA*007 is associated with susceptibility to UC in our population and MICA*008 with type 2 IBD PeA. Further work is now required to assess the distribution and expression of MICA throughout the gut in health and disease.