Asako Ando

HLA class II molecules and autoimmune hepatitis susceptibility in Japanese patients

To investigate the association between autoimmune hepatitis and HLA alleles in Japanese patients, serological typing and class II genotyping were performed using the polymerase chain reaction-restriction fragment length polymorphisms (PCR-RFLP) method. Serological typing showed that HLA-B54, -DR4, -DR53, and -DQ4 were significantly more frequent in patients with autoimmune hepatitis than in controls. HLA-DR4 was most frequently associated with autoimmune hepatitis (88.7%). In PCR-RFLP typing, the frequency of DRB1*0405 was significantly higher in autoimmune hepatitis than in controls. However, there was no significant difference in the frequency of Dw between the patients and the controls who were DR4-positive. The significant increase observed in DQA1*0301 and DQB1*0401 was explained by a linkage disequilibrium with DR4. Six DR4-negative patients had DR2, but there was no significant difference in the frequency of the DR2-associated Dw-alleles compared with the DR2-positive controls. No DPB1 allele was significantly associated with autoimmune hepatitis. These findings suggest that the basic amino acid at position 13, which is present only on the DR2 and DR4 B1 molecules (Arg on DR2 and His on DR4), contributes to the susceptibility to autoimmune hepatitis among the Japanese.

Southern hybridization analysis of DNA polymorphism in the HLA-D region

Restriction fragment-length polymorphisms (RFLP) were systematically analyzed by Southern hybridization with restriction endonuclease-digested genomic DNA from 28 HLA-homozygous B cell lines with Dw1-Dw19 specificity using the DR beta and DQ beta chain cDNAs as probes. These probes detected polymorphic fragments unique to each HLA-DR specificity. Furthermore, the DQ beta chain probes permitted us to distinguish between different Dw specificities with an identical DR type much more efficiently than with the DR beta chain probe. Distribution analysis of restriction fragments hybridizing to DR beta in relation to the DR and DQ specificities showed several sets of them forming ten clusters, some of which correlate with DRw53, DQw1, and DR alleles. This DNA typing technique allows the direct definition of HLA types at the gene level and provides a powerful tool for isolating genes controlling HLA-associated diseases.

Precise switching of DNA replication timing in the GC content transition area in the human major histocompatibility complex.

The human genome is composed of long-range G+C% (GC%) mosaic structures thought to be related to chromosome bands. We previously reported a boundary of megabase-sized GC% mosaic domains at the junction area between major histocompatibility complex (MHC) classes II and III, proposing it as a possible chromosome band boundary. DNA replication timing during the S phase is known to be correlated cytogenetically with chromosome band zones, and thus the band boundaries have been predicted to contain a switch point for DNA replication timing. In this study, to identify to the nucleotide sequence level the replication switch point during the S phase, we determined the precise DNA replication timing for MHC classes II and III, focusing on the junction area. To do this, we used PCR-based quantitation of nascent DNA obtained from synchronized human myeloid leukemia HL60 cells. The replication timing changed precisely in the boundary region with a 2-h difference between the two sides, supporting the prediction that this region may be a chromosome band boundary. We supposed that replication fork movement terminates (pauses) or significantly slows in the switch region, which contains dense Alu clusters; polypurine/polypyrimidine tracts; di-, tri-, or tetranucleotide repeats; and medium-reiteration-frequency sequences. Because the nascent DNA in the switch region was recovered at low efficiency, we investigated whether this region is associated with the nuclear scaffold and found three scaffold-associated regions in and around the switch region.

A boundary of long-range G+C% mosaic domains in the human MHC locus: Pseudoautosomal boundary-like sequence exists near the boundary

The human genome is composed of long-range G+C% (GC%) mosaic structures related to chromosome bands. We found the human MHC locus to be an example of megabase-level GC% mosaic structures and predicted a possible boundary of the megabase-level domains within an undercharacterized 450-kb region harboring the junction of MHC classes II and III. Chromosome walking of the 450-kb region and base-compositional analysis precisely located the boundary of the mosaic domains, disclosing a sharp GC% transition. Near the transition point there was a 20-kb dense Alu cluster, a 30-kb dense LINE-1 cluster, and a sequence highly homologous with the pseudoautosomal boundary of the short arms of human sex chromosomes (PAB1X and PAB1Y); PAB1X and PAB1Y are the interface between sex-specific and pseudoautosomal regions. Many PAB1XY-like sequences (PABLs) were detected by hybridization against genomic DNA, and the new sequences defined the complete form of PABLs to be about 650 nt.

Nomenclature for factors of the SLA system, update 2008

This report summarizes the new swine leukocyte antigen (SLA) allele sequences and haplotypes designated by the SLA Nomenclature Committee of the International Society for Animal Genetics. There have been 74 new SLA alleles, comprising 18 SLA-1 alleles, 11 SLA-2 alleles, six SLA-3 alleles, two SLA-6 alleles, one SLA-DRA allele, 20 SLA-DRB1 alleles, three SLA-DQA alleles and 13 SLA-DQB1 alleles. Twelve new SLA class I and four new class II haplotypes have also been designated. This is the first official update since the 2005 reports on the nomenclature for factors of the SLA class I and II systems. This report also summarizes recent updates to the Immunopolymorphism Database-Major Histocompatibility Complex (IPD-MHC) website (http://www.ebi.ac.uk/ipd/mhc/sla/). All information has now been integrated to the SLA section of the IPD-MHC database, which serves as the repository for maintaining a list of all recognized SLA genes and their allelic sequences.

Extracellular matrix protein tenascin-like gene found in human MHC class III region

The major histocompatibility complex (MHC) plays a central role in immune response, and susceptibility to a number of diseases, including autoimmune diseases, is suspected to be controlled by genes in this complex (Tiwari and Terasaki 1985; Klein 1986; Todd et al. 1988; Kostyu 1991). It is not clear whether this susceptibility is due to already known gene products or those of not yet identified neighboring genes. Many additional genes have been discovered in the MHC (Spies et ai. 1989, 1990; Kendall et al. 1990; Ragoussis et al. 1991). We reported previously a cluster of fibronectin type III repeats (MHC-F3) located approximately 20 kilobases (kb) centromeric of CYP21 in class III, which showed sequence similarity with human tenascin (Matsumoto et al. 1992a). Tenascin is a multidomain and multifunctional extracellular matrix glycoprotein, sharing several features with fibronectin and laminin but expressed in a more restricted fashion in tissues such as the central nervous system, cartilage, and soft tissue stroma (ChiquetEhrismann et al. 1986; Spring et al. 1989; ChiquetEhrismann 1990; Wehrle-Haller et al. 1991). Tenascin, whose gene is on chromosome 9, is made up of heptad, epidermal growth factor (EGF), fibronectin type III repeats, and a fibrinogen domain, arranged in this order from the Nto C-terminus (Gulcher et al. 1990; Siri et al. 1991; Gulcher et al. 199l). If the gene in MHC is actually a tenascin-like gene, heptad and EGF repeats may exist upstream from the clustered type III repeats. Taking into account the human tenascin gene structure (Gulcher et al. 1991), 12 kb Eco RI fragment approximately 60 kb centromeric of CYP21 (Fig. 1) was sequenced. Both the heptad and the EGF repeats, showing clear similarity with corresponding regions of tenascin, were found in the 2.6 kb sequence (Fig. 2). There were four heptads (MHC-

Rapid assignment of the swine major histocompatibility complex (SLA) class I and II genotypes in Clawn miniature swine using PCR-SSP and PCR-RFLP methods

Abstract:  Background:  Inbred miniature swine with defined novel SLA haplotypes will be useful in allo‐ and xeno‐transplantation studies, which can be carried out representing variable combinations of SLA haplotypes.

Gene organization of human NOTCH4 and (CTG)n polymorphism in this human counterpart gene of mouse proto-oncogene Int3

The cDNA and genomic clones for the human counterpart of the mouse mammary tumor gene Int3 were isolated and sequenced. We designated this human major histocompatibility complex (MHC) class III gene as NOTCH4, since very recently, by sequencing cDNA clones, the complete form of the mouse proto-oncogene Int3 has been clarified and named Notch4. The present human NOTCH4 sequence is the first example of the genomic sequence for the extracellular portion of the mammalian Notch4, and by comparing it with the mouse Notch4 cDNA sequence, the exon/intron organization was clarified. The comparison of the predicted amino acid sequence of human NOTCH4 with those of other Notch homologues of a wide range of species revealed four subfamilies for mammalian Notch. In the protein coding region of human NOTCH4, we found (CTG)n repeats showing a variable number tandem repeat (VNTR) polymorphism for different human leukocyte antigen (HLA) haplotypes. Ten genes mapped on 6p21.3, including NOTCH4, were found to have counterparts structurally and functionally similar to those mostly mapped on 9q33-q34, indicating segmental chromosome duplication during the course of evolution. Similarity of genes on chromosomes 1, 6, 9 and 19 was also discussed.

Cloning of a new kinesin-related gene located at the centromeric end of the human MHC region

We previously reported the presence of a new gene (HSET) with an unknown function, in the centromeric side of the class II gene region of the human major histocompatibility complex (MHC). cDNA clones corresponding to the HSET gene were isolated from a human testis cDNA library. A 2.4 kilobase transcript from the HSET gene was abundantly expressed in testis, B-cell, T-cell, and ovary cell lines but was not detected in lung or stomach. Analysis of the nucleotide sequence of the HSET cDNA clones revealed significant similarity to kinesin-related proteins in yeast, Drosophila, and human. Its predicted amino acid sequence contains a domain with strong sequence similarity to the ATP-binding and motor domains of a plus end-directed microtuble motor protein, kinesin, which might be involved in mitotic chromosome segregation, suggesting that the HSET gene encodes a novel kinesin-related protein.

HLA-DP typing by analysis of DNA restriction fragment length polymorphisms in the HLA-DPβ subregion

HLA class II antigens are encoded in the HLA-D region and are highly polymorphic. Southern hybridization technique was used to analyze restriction fragment length polymorphisms (RFLPs) in the DP beta gene and an attempt was made to correlate these with DP haplotypes derived from primed lymphocyte typing (PLT) analysis. Digestion of DNA from 32 Epstein-Barr virus (EBV)-transformed cell lines (of haplotypes DPw2, DPw3, DPw4, DPw5, and Cp63) with three different restriction endonucleases. Southern transfer, and hybridization to the DP beta cDNA probe revealed multiple fragments in all cell lines tested. The polymorphic patterns of these fragments were found to correlate with DP haplotypes, suggesting the possibility that the analysis of DNA RFLPs (DNA typing) in the HLA-DP beta subregion can distinguish and identify HLA-DP haplotypes.