Identification of a novel MICB allele,MICB*030, by cloning and sequencing

A novel MICB allele, MICB*030, has been identified in a healthy Chinese individual of Mongol ethnicity residing in northern China by polymerase chain reaction sequence‐based typing (PCR‐SBT) and confirmed by cloning and sequencing. MICB*030 was linked to HLA‐B*35. Aligned with MICB*005:02, MICB*030 has a nonsynonymous adenine substitution at nucleotide position 50 in exon 3, leading to amino acid change from serine to arginine at codon 102 of the mature MICB molecule.     

A new MICA allele,MICA*007:07, characterized by cloning and sequencing

A new MICA allelic variant, MICA*007:07, was identified in an individual of Mongol ethnicity in the Inner Mongolia Autonomous Region, northern China. Following polymerase chain reaction‐sequence‐based typing (PCR‐SBT), this new allele was further confirmed by cloning and sequencing. MICA*007:07 differs from MICA*007:01 by a synonymous mutation from G to A at the 2nd nucleotide position in exon 2. MICA*007:07 was linked to HLA‐B*27:05.     

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 the major histocompatibility complex class I chain-related gene B (MICB) polymorphism in a northern Chinese Han population: the identification of a new MICB allele, MICB*023

Major histocompatibility complex class I chain-related gene B (MICB) has only been characterized for allelic variation in very few human populations. The MICB polymorphism remains largely unknown in Chinese populations. In this study, 104 healthy unrelated Han subjects recruited from central Inner Mongolia Autonomous Region, northern China, were investigated by sequence-based typing for MICB allelic variation, the association of MICB alleles with AluyMICB insertion/deletion dimorphism located in MICB intron 1, linkage disequilibrium of MICB with human leukocyte antigen (HLA)-B and MICA, and HLA-A-C-B-MICA-MICB haplotypic diversity. Ten kinds of MICB alleles were observed, among which MICB*005:02/010, MICB*002:01, and MICB*004:01 were the most frequent alleles with frequencies of 51.44, 16.35, and 11.54%, respectively. Significant linkage disequilibrium (LD) was observed for 9 of the 21 HLA-B-MICB haplotypes and 6 of the 17 MICA-MICB haplotypes with a frequency >1.5%. In particular, HLA-B*13:01 and HLA-B*13:02, both of which were frequently represented in this population, exhibited a distinct LD pattern with the MICB allele. A new MICB allele, MICB*023, was identified, which differed from MICB*005:02/010 by a single mutation of G to A at position 86 in exon 2, resulting in an amino acid change from arginine to histidine at codon 6. HLA-A*30-C*06-B*13:02-MICA*008:01-MICB*005:02/010 was the most common haplotype, with a frequency of 8.64% in this population. HLA-A*02-C*08-B*48-MICA*Del-MICB*009N demonstrated a frequency of 2.4% in this population. Our results provide for the first time data regarding the MICB genetic polymorphism in northern Chinese Han populations and will form the basis for future studies of the potential role of MICB in allogeneic organ transplantation and disease association in related ethnic groups.     

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.     

MICB polymorphisms and haplotypes with MICA and HLA alleles in Koreans

Major histocompatibility complex (MHC) class I chain-related gene B (MICB) is located within the human MHC class I region. The location of MICB in the MHC region may imply the presence of linkage disequilibrium with polymorphic MICA and human leukocyte antigen (HLA) loci. MICB is also polymorphic; however, MICB polymorphisms have not been investigated in Koreans. Using sequence-based typing (SBT), we estimated the allelic frequencies of MICB and haplotypes with MICA, HLA-B, and HLA-DRB1 at high resolution in a population of 139 unrelated Korean individuals. Eight MICB alleles were identified. The most frequent allele was MICB*005:02/*010 (57.2%), followed by *002 (11.5%), *004 (8.3%), *005:03 (8.3%), and *008 (6.8%). The most common two-locus haplotypes were MICB*005:02/*010-MICA*010 (19.4%), MICB*005:02/*010-DRB1*15:01 (6.5%), and MICB*005:02/*010-B*15:01 (10.4%); the most common three-locus haplotypes were B*15:01-MICA*010-MICB*005:02/*010 (5.8%) and MICA*010-MICB*005:02/*010-DRB1*04:06 (10.4%); and the most common four-locus haplotype was B*15:01-MICA*010-MICB*005:02/*010-DRB1*04:06 (5.8%). This is the first study to provide information about MICB allele frequencies and haplotypes with HLA in Koreans. These study results should help understand mechanisms of disease association between the MICB locus and neighboring loci in Koreans.     

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.     

MICB polymorphism in a southern Chinese Han population: the identification of two new MICB alleles, MICB*005:06 and MICB*026

In this study, we investigated the major histocompatibility complex (MHC) class I chain-related gene B (MICB) allelic variation by sequence-based typing (SBT) in 201 healthy, unrelated Han subjects from Hunan province, southern China. Eleven MICB alleles were observed, among which MICB^∗005:02 predominated with a frequency of 64.93%. Significant linkage disequilibrium (LD) was observed for 5 HLA-B-MICB and 6 MICA-MICB haplotypes. Compared with a northern Chinese Han population, several MICB-containing haplotypes appeared to be highly specific to this southern Chinese Han population. Two new MICB alleles, MICB^∗005:06 and MICB^∗026, were identified. Aligned with MICB^∗005:02, MICB^∗005:06 has a synonymous T replacement at nucleotide 762 in exon 4; MICB^∗026 has probably arisen from MICB^∗004:01 through a single nucleotide substitution from G to A at position 826 in exon 4, leading to an amino acid change from glutamic acid to lysine at codon 253. HLA-A^∗02-C^∗01-B^∗46-MICA^∗010-MICB^∗005:02-DRB1^∗09 was the most prevalent six-locus haplotype with a frequency of 8.49%. HLA-A^∗30-C^∗06-B^∗13:02-MICA^∗008:01-MICB^∗005:02-DRB1^∗07 appeared to be a conserved extended haplotype. Our results provide new information about MICB genetic polymorphism in Chinese Han populations, and will inform future studies of the potential role of MICB in allogeneic organ transplantation and disease susceptibility in related ethnic groups.     

HLA class I chain-related MICA and MICB genes polymorphism in healthy individuals from the Bulgarian population

Although human leukocyte antigen (HLA) gene polymorphism has been investigated in many populations around the world, the data on MHC class I chain-related (MIC) genes are still limited. The present study is aimed to analyze the allelic polymorphism of MICA and MICB genes and haplotype associations with HLA-B locus in 132 healthy, unrelated individuals from the Bulgarian population by next generation sequencing (NGS). A total of 36 MICA and 16 MICB alleles were observed with the highest frequency detected for MICA*008:01 (17.1%) and MICB*005:02 (32.4%). Further, two and three-loci haplotype frequencies and pairwise linkage disequilibrium were estimated. Highly significant global linkage disequilibrium was found between either HLA-B and MICA and MICB genes. This is the first study on MICA and MICB allelic polymorphism, linkage disequilibrium, and haplotype polymorphism in the Bulgarian population. These results will allow for better characterization of the genetic heterogeneity of the Bulgarian population and could contribute to further analyses on MICA and MICB clinical significance.     

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.     

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.     

Identification of a new HLA‐G allele,HLA‐G*01:19, by cloning and phasing

A new HLA‐G allelic variant, HLA‐G*01:19, was identified in a southern Chinese Han population by polymerase chain reaction–sequence‐based typing (PCR‐SBT), cloning and phasing. HLA‐G*01:19 differs from HLA‐G*01:04:01 by a nonsynonymous cytosine at position 99 in exon 2, resulting in amino acid change from valine to leucine at codon 34 of the mature HLA‐G molecule.     

PDX1 ‐MODY and dorsal pancreatic agenesis: New phenotype of a rare disease

Maturity‐Onset Diabetes of the Young (MODY) type 4 or PDX1 ‐MODY is a rare form of monogenic diabetes caused by heterozygous variants in PDX1 . Pancreatic developmental anomalies related to PDX1 are reported only in neonatal diabetes cases. Here, we describe dorsal pancreatic agenesis in 2 patients with PDX1 ‐MODY. The proband presented with diabetes since 14 years of age and maintained regular glycemic control with low doses of basal insulin and detectable C‐peptide levels after 38 years with diabetes. A diagnosis of MODY was suspected. Targeted next‐generation sequencing identified a heterozygous variant in PDX1 : c.188delC/p.Pro63Argfs*60. Computed tomography revealed caudal pancreatic agenesis. Low fecal elastase indicated exocrine insufficiency. His son had impaired glucose tolerance, presented similar pancreatic agenesis, and harbored the same allelic variant. The unusual presentation in this Brazilian family enabled expansion upon a rare disease phenotype, demonstrating the possibility of detecting pancreatic malformation even in cases of PDX1 ‐related diabetes diagnosed after the first year of life. This finding can improve the management of MODY4 patients, leading to precocious investigation of pancreatic dysgenesis and exocrine dysfunction.     

Association of MICA and MICB alleles with symptomatic dengue infection

Dengue viruses (DV) are one of the most important arthropod-borne viral diseases in the developing world. DV can cause syndromes that are either self-limiting or severe. Allelic variants of human leukocyte antigen (HLA) genes have been demonstrated to be associated with disease susceptibility. Here we report the association of nonclassical HLA class I MICA–MICB genes with disease outcome during DV infection. A sequencing-based typing method and genotyping of MICA and MICB in a well-characterized group of Cuban individuals with dengue hemorrhagic fever (DHF), dengue fever (DF), or asymptomatic dengue infection (ADI) was performed. Statistical analysis revealed a tendency for MICA*008 and MICB*008 to associate with susceptibility to illness when symptomatic versus asymptomatic cases (odds ratio [OR] = 2.1, p_v = 0.03, and OR = 10.4, p = 0.0096, respectively) were compared. Surprisingly, a stronger association of both allelic forms was observed for the DF patients compared with the ADI group (MICA*008, OR = 5.2, p = 0.0001; and MICB*008, OR = 13.2, p = 0.0025) rather than the severe cases. Major histocompatibility class I-related gene-related natural killer cells and/or γδ and αβ T-cell activation might regulate the development of symptomatic DF and DHF.     

Characterization of a novel HLA‐B*40 allele,HLA‐B*40:186:02, by cloning and sequencing

A novel HLA‐B*40 variant, HLA‐B*40:186:02, has been identified by cloning and sequencing in a southern Chinese Han population. Aligned with HLA‐B*40:01:01, HLA‐B*40:186:02 has a nonsynonymous cytosine mutation at nucleotide position 165 in exon 2, leading to amino acid change from glycine to arginine at codon 56. It differs from HLA‐B*40:186:01 by a synonymous change (adenine to cytosine) at position 165 in exon 2.     

Identification of the first homozygous 1‐bp deletion in GDF9 gene leading to primary ovarian insufficiency by using targeted massively parallel sequencing

Targeted massively parallel sequencing (TMPS) has been used in genetic diagnosis for Mendelian disorders. In the past few years, the TMPS has identified new and already described genes associated with primary ovarian insufficiency (POI) phenotype. Here, we performed a targeted gene sequencing to find a genetic diagnosis in idiopathic cases of Brazilian POI cohort. A custom SureSelect^XT DNA target enrichment panel was designed and the sequencing was performed on Illumina NextSeq sequencer. We identified 1 homozygous 1‐bp deletion variant (c.783delC) in the GDF9 gene in 1 patient with POI. The variant was confirmed and segregated using Sanger sequencing. The c.783delC GDF9 variant changed an amino acid creating a premature termination codon (p.Ser262Hisfs*2). This variant was not present in all public databases (ExAC/gnomAD, NHLBI/EVS and 1000Genomes). Moreover, it was absent in 400 alleles from fertile Brazilian women screened by Sanger sequencing. The patient's mother and her unaffected sister carried the c.783delC variant in a heterozygous state, as expected for an autosomal recessive inheritance. Here, the TMPS identified the first homozygous 1‐bp deletion variant in GDF9. This finding reveals a novel inheritance pattern of pathogenic variant in GDF9 associated with POI, thus improving the genetic diagnosis of this disorder.     

Homozygous variants in MAPRE2 and CDON in individual with skin folds,growth delay,retinal coloboma,and pyloric stenosis

Cases with multiple molecular diagnoses are challenging to diagnose clinically, yet may be resolved by unbiased exome sequencing analysis. We report an infant with developmental delay, severe growth delay, dysmorphic features, and multiple congenital anomalies including retinal coloboma, congenital pyloric stenosis, and circumferential skin creases. Exome sequencing identified a homozygous missense variant in MAPRE2 and a homozygous stopgain (nonsense) variant in CDON. Variants in MAPRE2, encoding a regulator of microtubule dynamics, lead to congenital symmetric circumferential skin creases type 2, with associated dysmorphism, small growth parameters, and congenital cardiac and genital anomalies. Monoallelic variants in CDON, encoding a coreceptor for sonic hedgehog, have been associated with autosomal dominant pituitary stalk interruption syndrome and holoprosencephaly. Cdon?/? mice have multiple eye defects including coloboma, consistent with the observed human phenotype. Thus, the complex phenotypic presentation of the infant may potentially be attributed to a dual molecular diagnosis. Furthermore, we present CDON as a candidate gene for coloboma formation in addition to the known holoprosencephaly phenotype, and propose to expand the allelic spectrum of CDON to variants associated with autosomal recessive inheritance in addition to dominant inheritance.     

The new neuromuscular disease related with defects in the ASC‐1 complex: report of a second case confirms ASCC1 involvement

Next‐generation sequencing technology aided the identification of the underlying genetic cause in a female newborn with a severe neuromuscular disorder. The patient presented generalized hypotonia, congenital bone fractures, lack of spontaneous movements and poor respiratory effort. She died within the first days of life. Karyotyping and screening for several genes related with neuromuscular diseases all tested negative. A male sibling was subsequently born with the same clinical presentation. Whole‐exome sequencing was performed with variant filtering assuming a recessive disease model. Analysis focused on genes known to be related firstly with congenital myopathies, extended to muscle diseases and finally to other neuromuscular disorders. No disease‐causing variants were identified. A similar disorder was described in patients with recessive variants in two genes: TRIP4 (three families) and ASCC1 (one family), both encoding subunits of the nuclear activating signal cointegrator 1 (ASC‐1) complex. Our patient was also found to have a homozygous frameshift variant (c.157dupG, p.Glu53Glyfs*19) in ASCC1 , thereby representing the second known case. This confirms ASCC1 involvement in a severe neuromuscular disease lying within the spinal muscular atrophy or primary muscle disease spectra.     

海南汉族人群MICB等位基因多态性与类风湿性关节炎相关性研究

目的 研究海南汉族人群MHC I类链相关基因B(MHC class I chain-related gene B,MICB)等位基因多态性与类风湿性关节炎(rheumatoid arthritis,RA)的相关性.方法 用世界卫生组织推荐的标准盐析法提取外周静脉血DNA,采用PCR-序列特异性引物(PCR sequence specific primer,PCR-SSP)和PCR产物直接测序分型(PCR-sequence based genotyping,PCR-SBT)方法对样本MICB基因的多态性进行检测分析.结果 RA患者及对照组中共有10种MICB等位基因被检测出,两组中MICB*005:02等位基因频率分布最高,其频率为52.6％比42.0％,但MICB* 002等位基因频率在RA组明显低于对照组(13.1％比25.5％),差异具有统计学意义(Pc＜0.05),MICB* 014等位基因频率在RA组亦明显低于对照组(8.1％比15.3％),差异具有统计学意义(Pc＜0.05).结论 MICB* 002和MICB*014等位基因与RA的易感性之间存在负相关,可能是RA的保护性基因.