MICB Research Articles

The association between non‐melanoma skin cancer and a young dimorphic Alu element within the major histocompatibility complex class I genomic region

A non-melanoma skin cancer (NMSC) susceptibility locus within the major histocompatibility complex (MHC) class I region was previously identified telomeric of the HLA-C gene using high-density microsatellite markers. Here, we have extended the previous microsatellite study by using the same DNA samples obtained from 154 NMSC patients and 213 normal controls from the town of Busselton in Western Australia and examined the relationship between five polymorphic Alu insertions (POALINs) within the MHC class I region and their association with NMSC. The genotype distribution of the AluyTF insertion that is located within the NMSC susceptibility region telomeric of the HLA-C gene was significantly increased according to the Fisher's exact test in the NMSC patients, and it was not in Hardy-Weinberg equilibrium in the control group. There was no difference between the cancer patients and controls for the genotypes of the AluyMICB locus within intron 1 of the MICB gene and the other three POALINs (AluyHJ, AluyHG and AluyHF) that are located within the genomic region of the HLA-A, -G and -F gene cluster. The test for significant linkage disequilibrium for 10 pairs of POALIN loci and estimations of two locus POALIN haplotype frequencies also revealed AluyTF differences between the cases and controls. In conclusion, the MHC class I POALIN, AluyTF, that is located within the NMSC susceptibility locus and near the HLA-C gene was strongly associated with NMSC. This finding, using five different polymorphic Alu insertion markers, supports the previous microsatellite association study that one or more genes located in close proximity to the AluyTF insertion has a potential role in NMSC.

AluyMICB dimorphism within the class I region of the major histocompatibility complex is associated with asthma and airflow obstruction in the Busselton population

To examine the association between the Alu dimorphism within the first intron of the MICB gene and asthma and airflow obstruction. Background The highly polymorphic non-classical MHC class I polypeptide-related (MIC) genes, MICA and MICB, encode stress inducible glycoproteins, which are expressed on a variety of epithelial cells, including those of the lungs. AluyMICB genotyping was performed on 1109 subjects from the Busselton Health Study. From a standard questionnaire, 359 individuals indicated that they had been diagnosed by a doctor with asthma. Lung function was assessed by the forced expired volume in 1 second (FEV1) and expressed as a percent of the predicted value. Airflow obstruction was defined as FEV1<80% predicted. In men, a dominant relationship was found between the AluyMICB DD genotype and asthma (P=0.006; chi2(2)=7.65). Furthermore, multivariate analysis adjusted for age, height, weight and body mass index (BMI) showed a relationship between DD genotype and asthma in men in a dominant model (odds ratio (OR)=1.97; 95% confidence interval (CI)=1.11-3.51; P=0.021). In women, an association was found between the AluyMICB II genotype and FEV1 percent predicted as a continuous variable (P=0.001). When adjusted for age and BMI, it showed a significant relationship between AluyMICB and airflow obstruction in a dominant model (OR=14.11%, 95% CI 3.29-60.57, P<0.001). However, no association was found between the AluyMICB II genotypes and airflow obstruction in men. These findings suggest the possible involvement of a MHC class I gene in abnormal airway structure in women and airway function in men.

MHC Class I Chain-Related Gene B Promoter Polymorphisms and Celiac Disease

The possibility that susceptibility to celiac disease (CD) might be influenced by the MHC class I chain-related gene family, MICA and MICB, has been previously reported. In this study, we analyzed the MICB promoter and examined the association of the polymorphisms found within such in a group of CD patients. To study the MICB promoter we sequenced the 5′ flanking region of MICB gene in DNA from homozygous B-lymphoblastoid cell lines corresponding to the most frequent MICB alleles found in our population (MICB*00502, MICB*002, MICB*004, and MICB*008). DNA from a MICB*003 homozygous individual was also analyzed. Sequence analysis revealed six single nucleotide polymorphisms located at positions 45860 C/A, 45862 G/C, 45877 C/G, 46113 A/C, 46219 G/C, and 46286 G/C and an insertion of 2 bp --/AG at position 45944 according to the published genomic sequence. Those polymorphisms were found to be associated in four different haplotypes corresponding to different MICB alleles. Subsequently, 126 CD subjects and 117 healthy controls were typed by polymerase chain reaction using sequence-specific primers for these polymorphisms. MICB promoter polymorphism haplotypes were also found in our population and showed strong linkage disequilibrium with MICB alleles. MICB promoter polymorphism Haplotype 3, included in MICB*002 and MICB*008 alleles, was found to be overrepresented in CD patients (79.4% CD patients vs 45.3% healthy controls; p c < 0.0001; OR = 4.64; CI 95% = 2.64–8.16). Both MICB*008 and MICB*002 alleles were found as part of the CD susceptibility extended haplotypes B8/DR3/DQ2, B18/DR3/DQ2, and DR4/DQ8.

Reduced MIC Gene Repertoire Variation in West African Chimpanzees as Compared to Humans

The human major histocompatibility complex class I chain-related (MIC) genes are members of a multicopy family showing similarity to the classical HLA-A, HLA-B, and HLA-C genes. Only the MICA and MICB genes produce functional transcripts. In chimpanzees, however, only one MIC gene is expressed, showing an intermediate character, resulting from a deletion fusing the MICA and MICB gene segments together. The present population study illustrates that all chimpanzee haplotypes sampled possess the hybrid MICA/B gene. In contrast to the human situation this gene displays reduced allelic variation. The observed repertoire reduction of the chimpanzee MICA/B gene is in conformity with the severe repertoire condensation documented for Patr-B locus lineages, probably due to the close proximity of both genes.

Study on the haplotypes of MICA and MICB microsatellite and HLA‐B locus in the Guangzhou Han population

The purpose of this study was to investigate the genetic polymorphisms and haplotypes of microsatellite locus in exon 5 of the MICA gene and intron 1 of the MICB gene and human leukocyte antigen-B (HLA-B) gene based on 106 samples of the Guangzhou Han population through means of polymerase chain reaction and the fluorescent technique (6-FAM). The corresponding haplotype frequencies, linkage disequilibrium values and relative linkage disequilibrium values were estimated based on population data. The results show that the genotype distributions of MICA and MICB microsatellite and HLA-B satisfy the Hardy-Weinberg equilibrium. In total, five alleles of MICA microsatellite locus and 14 alleles of MICB microsatellite locus were observed. MICA A5 was the most common allele (0.2877), whereas A4 was the least common (0.1321). MICB CA14 was the most common allele (0.3255), and CA19 and CA28 were the two least common (0.0047). CA27 was not observed at all. Five kinds of MICA-MICB haplotypes, 18 kinds of MICA-HLA-B haplotypes and 12 kinds of MICB-HLA-B haplotypes occurred at frequencies of more than 1%. The common haplotypes of MICA-MICB, MICA-HLA-B and MICB-HLA-B were A5-CA14, A5.1-CA18, A4-CA26, A9-CA15, A5-B*15(62), A5.1-B*1301/1302, A4-B*1301/1302, A6-B*51, A6-B*4403, A9-B*3802, CA14-B*4601, CA18-B*1301/1302 and CA26-B*1301/1302, and these haplotypes showed strong linkage disequilibrium. The polymorphisms and haplotype distributions of MICA and MICB microsatellite and HLA-B locus in the Guangzhou Han population have their own distinct genetic characteristics. The microsatellite locus of exon 5 of the MICA gene and intron 1 of the MICB gene could therefore be used as genetic markers in the studies of anthropology, gene linkage analysis in genetic diseases, individual identification and paternity testing in forensic medicine.

The phylogenetic history of the MHC class I gene families in pig, including a fossil gene predating mammalian radiation.

More than 990 kb of the 1200 kb in the SLA class I region of the pig major histocompatibility complex (MHC) have been sequenced. The present study was designed to establish the evolution of this region which was best understood by distinguishing three periods. The most recent period, which extended from 40 to 15 mya, probably corresponded to five rounds of duplication of a basic unit. This unit consisted of a single class I gene linked to widely dispersed repeats, and one SLA-specific repeat motif. The duplications gave rise to six SLA classical class I genes. The second evolutionary period corresponded to the emergence of the SLA nonclassical class I genes, i.e. after the suidae separated from the other artiodactyl species about 65 mya. The third period appeared to correspond to a much more remote age when the ancestor of the gene SLA-11 existed. Comparative studies of the human and pig sequences of the class I-containing segments indeed revealed the presence within the human HSR1-ZNF segment of relics of a human class I fossil gene which appeared to be orthologous to the 5' moiety of the SLA-11 pseudogene. This was the first evidence that a class I gene existed in this location at least 110-120 mya in the MHC class I region of the precursor of the mammalian species. Human/pig sequence comparison also revealed that the presumably functional pig MIC2 gene was probably orthologous to the human functional MICA or MICB genes.

Association of MHC dimorphic Alu insertions with HLA class I and MIC genes in Japanese HLA-B48 haplotypes.

A large proportion of Japanese with the HLA-B48 allele have a MICA gene deletion associated with a MICB null allele within the class I region of the Major Histocompatibility Complex (MHC). Here, we report for the first time a novel positive association between the presence of a polymorphic Alu insertion, AluyMICB, within the first intron of the MICB gene and the MICAdel/MICBnull/HLA-B48 haplotype for five of six well-characterized Japanese cell-lines. The AluyMICB insertion was found to be present at a frequency of 0.242 in 86 Japanese tissue donors and in four of the five individuals with the HLA-B48 allele. The AluyMICB insertion was also associated with at least three different MICB alleles, *0102, *0107N and *0105, and three different HLA-B alleles, B13, B48 and B57, respectively, in the seven Workshop cell-lines (the 4th Asia-Oceania Histocompatibility Workshop, and the 10th International Histocompatibility Workshop) and the six Japanese cell-lines that were selected for this study. Based on the analysis of associations between different polymorphic markers within the beta block, the MICB*0102 allele was inferred to be the ancestral form of the MICB*0105 and MICB*0107N alleles. The AluyMICB polymorphism can now be used to further investigate its relationship with other MICB alleles and consequently their origins. In addition, we have examined the absence and presence of three other polymorphic Alu markers distributed within the alpha block of the class I region of the HLA-B48/AluyMICB haplotype. We conclude that the extended HLA-B haplotypes are best defined by considering multiple genomic sites including the four polymorphic Alu insertions described in this study.

Alternatively spliced forms of MICA and MICB lacking exon 3 in a human cell line and evidence of presence of similar RNA in human peripheral blood mononuclear cells.

MICA and MICB genes encode MHC class I chain-related proteins, which are polymorphic, do not appear to present peptides or associate with beta(2)-microglobulin, and are expressed predominantly in epithelial cells, endothelial cells, fibroblasts and several cultured cell lines. Alternatively spliced isoforms are known to exist for HLA-A and B, as well as HLA-G and the MHC class I-related gene, MR1. In the course of cloning MICA and MICB cDNA from the colon carcinoma cell line HCT 116, it was observed that two kinds of cDNAs were obtained: a 1161-bp cDNA, representing full-length MICA or MICB, and a shorter variant of 873 bp. The sequences of these short cDNAs were those of the correct MICA or MICB alleles but lacking exon 3. They were found in 7 of 72 clones examined or about 10% and were called MICA2 and MICB2. MICA1 and MICA2 were transfected into Chinese hamster ovary cells and found to be expressed both in the cells and on their surface. PCR with a primer based on a sequence formed by the joining of exons 2 and 4 allowed detection of the isoform RNA in different cells including freshly prepared normal PBMC.

Alu polymorphism within the MICB gene and association with HLA-B alleles

Alu polymorphism within the MICB gene and association with HLA-B alleles

Alu polymorphism within the MICB gene and association with HLA-B alleles.

We describe the finding of an Alu repeat dimorphism within the first intron of the MICB gene. The frequencies of the two AluyMICB alleles, AluyMICB*0(absence of insertion) and AluyMICB*1(presence of insertion), and their associations with the highly polymorphic HLA-B locus were determined for 51 human cell lines and for 109 and 200 Caucasians and northeastern Thais, respectively. Analysis of the AluyMICB and HLA-B allelic relationships revealed that AluyMICB*1 occurred at relatively low gene frequency (0.118-0.157) [corrected] but was strongly associated with HLA-B17 (HLA-B57,HLA-B58) and HLA-B13. The AluyMICB locus provides a useful dimorphic marker for investigations on the level of linkage disequilibrium between MICB, MICA, and HLA-B loci.

Primary sclerosing cholangitis is associated to an extended B8-DR3 haplotype including particular MICA and MICB alleles.

Susceptibility to primary sclerosing cholangitis (PSC) is associated with HLA-B8, -DR3, -DR2, and -DR6. It is not established whether these HLA genes or closely linked genes confer the primary disease susceptibility. MICA and MICB genes are found in the class I region between HLA-B and DRB. MICA is expressed in gastrointestinal epithelium and activates gammadelta T cells in the gut. Because PSC is strongly associated with inflammatory bowel disease, we investigated whether MICA and MICB contribute to the HLA-associated genetic susceptibility to develop PSC. The study included 130 PSC patients and 306 healthy controls, previously typed for HLA class I and II genes, typed for 5 MICA and 15 MICB microsatellite alleles. The phenotype frequencies of MICA5.1 and MICB24 were significantly increased among PSC patients compared with controls (90% vs. 74%; odds ratio [OR] = 3.2; P(c) = 3 x 10(-3) and 58% vs. 29%; OR = 3.3; P(c) < 1 x 10(-7), respectively). When stratified for B8- or DR3-positive and -negative individuals, the association of these markers to PSC was no longer evident. However, we observed that B8 and DR3 were associated to PSC only in the presence of both MICA5.1 and MICB24 markers. The frequency of individuals carrying all 4 alleles was significantly increased among the PSC patients compared with controls (49% vs. 18%; OR = 4.5; P(c) < 1 x 10(-7)). Our data indicate that PSC is associated to the extended B8-MICA5.1-MICB24-DR3 haplotype.

Oxidative stress increases MICA and MICB gene expression in the human colon carcinoma cell line (CaCo-2)

The human major histocompatibility complex class I chain-related A gene (MICA) and the MICB gene are newly identified members of the major histocompatibility complex class I chain-related gene family. We demonstrate here that oxidative stress, induced by H 2O 2, promoted MICA (2.2-fold) and MICB (3.8-fold) gene expression using the human colon carcinoma cell line (CaCo-2) and semi-quantitative RT-PCR.

Wide distribution of the MICA-MICB null haplotype in East Asians.

Stress-inducible MICA (MHC class I chain-related A) is known to bind to NKG2D, which is one of the natural killer (NK) cell receptors, and plays a role in immune surveillance. We have reported that a MICA-MICB null haplotype is in linkage disequilibrium with HLA-B*4801 in the Japanese population. In the haplotype, an approximately 100-kb deletion, including the entire MICA gene, was observed and MICB possessed a premature stop codon. In this study, a multiplex polymerase chain reaction (PCR) method was developed for detecting the MICA deletion. MICB alleles were typed by PCR-single-strand conformation polymorphism (SSCP) method and direct sequencing. The frequency of the MICA-MICB null haplotype was 3.7% on the average, and was strongly associated with HLA-B48 in seven East Asian populations. It was presumed that the stop codon of MICB gene generated after the large-scale deletion. The wide distribution of the null haplotype at polymorphic frequencies suggests that the haplotype has been conservatively maintained because of some selective advantage.

Three novel MICB alleles.

The two members of the MHC class I chain-related (MIC) gene family, MICA and MICB, have been shown by several investigators to be polymorphic. Most of the research effort so far has focussed on MICA, so less is known about the extent of polymorphism in the MICB gene. Here we report three novel MICB alleles, which had been detected in the course of an SSOP typing study on a large cohort of cell lines. Two of these alleles are formed by a non-synonymous nucleotide variation. Our results confirm previous findings that most of the polymorphisms in the MICB gene, as in MICA, are coding and suggest that the extent of polymorphism in the two genes might be comparable.

A new MICA allele with ten alanine residues in the exon 5 microsatellite.

The MICA and MICB genes code for protein products that have structural similarities to major histocompatibility complex (MHC) class I genes. These genes are upregulated by heat stress. They have been shown to interact with a common receptor (NKG2D/DAP10) on gammadelta T cells, CD8+ T cells and natural killer (NK) cells. The MICA gene has an expressed microsatellite, GCT, within the exon 5 which encodes for alanine. So far, four different repetitions of this short tandem have been reported. Also one non-synonymous, one synonymous substitution and a 1-bp insertion within this region have also been described. An association of Behcet's disease with the microsatellite A9 has been reported. Here we report a novel allele with 10 GCT repetitions (A10) which was detected by reference strand mediated conformation analysis and confirmed by DNA sequencing.

Microsatellite polymorphism of the MHC class I chain-related (MIC-A and MIC-B) genes marks the risk for autoimmune Addison's disease.

The major histocompatibility complex class I chain-related MIC-A and MIC-B genes are located on chromosome 6 between the histocompatibility leucocyte antigen (HLA)-B and the B-associated transcript genes. The presence of 21-hydroxylase autoantibodies is a sensitive and specific marker of autoimmune Addison's disease. We studied the polymorphism of exon 5 of the MIC-A gene, of intron 1 of the MIC-B gene, and of HLA-DRB1, -DQA1, and -DQB1 genes in 28 autoimmune (21-hydroxylase autoantibody positive) Addison's disease patients and in 75 healthy subjects from central Italy. The MIC-A5.1 allele was significantly more frequent in Addison's disease patients (79%) than in healthy subjects (36%) [odds ratio (OR) = 6.52, corrected P (Pc) = 0.0015], whereas MIC-A6 was significantly reduced in affected subjects (15% vs. 56%, OR = 0.13, Pc = 0.002). The A5.1/A5.1 genotype had an OR for autoimmune Addison's disease as high as 18.0 and an absolute risk of 1 per 1131. In the presence of MIC-A5.1, MICB-CA-25 was significantly increased in Addison's disease patients (25% vs. 4%, OR = 8.0, P = 0.0039, Pc = 0.047). The MICB-CA-17 allele was absent in Addison's disease patients, but present in more than 25% healthy individuals (OR = 0.10, P = 0.0025, Pc = 0.03). Among HLA-DR and -DQ haplotypes, only DRB1*03-DQA1*0501-DQB1*0201 (DR3/DQ2) was significantly more frequent in Addison's disease patients than in healthy subjects, but only in the presence of MIC-A5.1. The frequency of MIC-A5.1 was significantly increased in Addison's disease patients only in the presence of HLA-DR3-DQ2. Our study demonstrates that susceptibility to autoimmune Addison's disease is linked to the MIC-A microsatellite allele 5.1 and that both MIC-A5.1 and HLA-DR3/DQ2 are necessary to confer increased genetic risk for Addison's disease.

New polymorphic microsatellite markers in the human MHC class I region.

The human major histocompatibility complex (MHC) class I region is believed to contain a large number of genes encoding susceptible factors for diseases such as Behcet's disease, Graves disease and psoriasis vulgaris. To identify the causative genes of those diseases, we have conducted large-scale genomic sequencing and determined the 1.8 Mb entire HLA class I region from the MICB gene to the HLA-F gene. During the course of genomic sequencing, a total of 731 microsatellite sequences with dinucleotide to pentanucleotide repeats were found in this region. Previously, we reported that 26 microsatellites between MICB and S on the most centromeric side of the class I region, and between HSR1 and HLA-92/L in the midst of the class I region were highly polymorphic, and served as excellent genetic markers. In this paper, in order to fill the gaps with no known polymorphic microsatellites available in the HLA class I region, 12 new polymorphic microsatellite markers were recruited from the 1.8 Mb region including the remaining class I segments, namely between S and HSR1, and between HLA-92/L and HLA-F The average number of alleles at these new microsatellite loci was 8.2 with a polymorphism content value (PIC) of 0.63. These 38 markers in total almost uniformly interspersed in the HLA class I region will enable us to search precisely for the location of disease susceptible loci within the HLA class I region by association and for linkage analyses.

MIC-A polymorphism in Japanese and a MIC-A-MIC-B null haplotype.

A polymorphic gene, MIC-A, is one of the MIC family of genes which is composed of a group of homologous genes interspersed in the class III and class I regions of the major histocompatibility complex. MIC-A is located 46 kilobases (kb) centromeric of HLA-B, and is preferentially expressed in the epithelial cells and intestinal mucosa. Recently, MIC-A and the closely related MIC-B were reported as the molecules that conferred specificity in the recognition by the Vdelta1gammadeltaT cells. In the present study, polymorphic exons 2, 3, and 4 of the MIC-A gene were analyzed using the polymerase chain reaction-single-strand conformation polymorphism method. The number of patterns found in exons 2, 3, and 4 were 5, 6, and 4, respectively, in 114 healthy Japanese subjects. Eight MIC-A alleles were observed in Japanese individuals, among which one, tentatively named MIC-AMW, has not previously been reported. There was a strong linkage disequilibrium between MIC-A and HLA-B loci: each MIC-A allele showed strong association with a particular HLA-B group. In contrast, B*3901 showed association with multiple MIC-A alleles. Furthermore, the existence of a MIC-A-MIC-B null haplotype, which is associated with HLA-B*4801, was identified. In this haplotype, a large-scale deletion (of approximately 100 kb) including the entire MIC-A gene was indicated and the MIC-B gene possessed a stop codon.

Genome sequencing analysis of the 1.8 Mb entire human MHC class I region.

The human MHC class I region spans 1.8 Mb from the MICB gene to the HLA-F gene at the telomeric end of the HLA region. There are fewer genes recognized in this region than in the class II or class III region, probably because this region remained uncharacterized for genomic organization. Based on the 1,796,938 bp genomic sequence of the entire class I region determined in our laboratory, the complete gene structure of this region has finally emerged. This region embraces as many as 118 genes (73 known and 45 new genes) with a gene density of one gene every 15.2 kb, which is comparable to that of the gene-rich class III region. The GC content is fairly uniform throughout the class I region, being 45.8% on average, which corresponds to the isochore H1. By investigation of genetic polymorphisms in 26 out of 758 microsatellite repeats identified in the class I region, we could reduce the critical region for Behçet's disease (associated with B51) and psoriasis vulgaris (associated with Cw6) to approximately 50 kb segments, between MICA and HLA-B and between TCF19 and S, respectively. Thus, systematic large-scale genomic sequencing provides an efficient way of identifying genes and of mapping disease-susceptible genes in the genome.

Microsatellite polymorphism within the MICB gene among japanese patients with behçet’s disease

Behçet’s disease (BD) is known to be associated with HLA-B51. In order to investigate the influence of the MICB gene, located about 120 kb centromeric of the HLA-B gene, on the susceptibility to BD, (CA/TG) dinucleotide repeat microsatellite polymorphism in intron 1 of the MICB gene was investigated among 77 Japanese patients with BD, 60 randomly selected controls and 28 HLA-B51-positive unrelated healthy controls. There was no significant difference in the phenotype frequency of the microsatellite polymorphism between the BD patients and controls. This result suggests that the MICB gene itself is not responsible for the development of BD, and that the candidate gene(s) for BD is located between the MICA and HLA-C genes.