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Biomedical subjects

H A Erlich

Publications and source records attributed to H A Erlich.

At least 163 records · Page 9Linked to original sources
PubMed1990

Prenatal diagnosis of alpha-thalassemia by polymerase chain reaction and dual restriction enzyme analysis.

Asian couples at risk for a fetus with homozygous alpha-thalassemia (hydrops fetalis) are often identified by their low erythrocyte mean corpuscular volume (MCV) and normal hemoglobin electrophoresis when little time remains to test their genotypes by restriction enzyme analysis. DNA analysis is performed directly on chorionic villi or amniocytes remaining after an aliquot is used to establish a backup cell culture. The polymerase chain reaction (PCR) protocol quickly determines whether the fetus has hydrops fetalis without waiting for cultured cells to grow. Previously, growing cultured fetal cells to obtain more fetal material to establish unambiguously the fetal genotype with two independent restriction enzyme digests absorbed a significant portion of the time remaining to complete prenatal diagnosis. A dual restriction enzyme digestion protocol was development using a 3' zeta-globin probe to clearly distinguish the most common alpha-thalassemia deletions that represent nearly all the alpha-thalassemia haplotypes in Southeast Asia.

Base Sequence↗
PubMed1990

Unique sequences for two HLA-DRB1 genes expressed on distinct DRw6 haplotypes.

We have used restriction fragment length polymorphism (RFLP) analysis and DNA sequencing to characterize two distinct DRB1 alleles expressed on DRw52 and DQw7-associated haplotypes but not readily defined by conventional DR serology. These two haplotypes, designated HLA-D "HAG" and "PEV", react variably with DRw13(w6), DRw14(w6), and the more broad DR"3 + 6" antisera. Analysis of RFLP revealed that HLA-D "HAG" and "PEV" are associated with different DRw52 variants, and that "HAG" is indistinguishable from DRw18(3) haplotypes. Sequencing of the "HAG" and "PEV" DRB1 genes showed each to represent novel alleles. Nevertheless, these sequences show similarities with the other alleles of the DR5,w6, and w8 family. "HAG" (DRB1*1303) appears to have arisen either from two recombinational events involving at least three DRB1 sequences (DRB1*1101, DRB1*0803, DRB1*0401) or from a single recombinational event together with multiple point mutational events. "PEV" appears to represent a DRB1*1301-1302/DRB1*1101 recombinant allele, with recombination having occurred in the region of bases 175-198. The results of this study suggest that the DRw52 family of haplotypes is derived from a relatively restricted number of ancestral sequences, with diversity among DRB1 alleles within this family arising through gene conversion or recombination events.

Amino Acid Sequence↗
PubMed1990

HLA-DP beta and susceptibility to multiple sclerosis: an analysis of caucasoid and Japanese patient populations.

Nonradioactive sequence-specific oligonucleotide (SSO) probes specific for the HLA-DP beta locus have been used in a simple dot-blot assay to DP beta-type samples amplified by the polymerase chain reaction (pcr) from Caucasoid (n = 24) and Japanese (n = 23) patients with multiple sclerosis (ms) as well as ethnically matched controls. In contrast to previous reports, no DP beta allele was found to be increased in either patient population. However, the results do show a dramatic difference in the allele frequencies between the two control populations, further emphasizing the need for ethnically matched controls in studies of HLA and disease.

Alleles↗
PubMed1990

Shared molecular markers of genetic predisposition to seropositive rheumatoid arthritis.

Rheumatoid arthritis is associated with the human class II major histocompatibility complex antigens known as HLA-DR4. HLA-DR4 can be subdivided by cellular typing into five subtypes: Dw4, Dw10, Dw13, Dw14, and Dw15. By traditional serologic methods, 60-80% of rheumatoid arthritis patients type HLA-DR4 compared to approximately 20% of the general population. It has been demonstrated, using a panel of four alloreactive T-cell clones, each of which recognized HLA-DR4, Dw14 homozygous typing cells, that cells from all of a group of 23 rheumatoid arthritis patients could be recognized by one or more of these clones regardless of the patients' serologic typing. As the expressed polymorphism of the DR molecule is accounted for by the beta 1 gene, this gene was amplified, using the polymerase chain reaction, and sequenced. Seven patients whose cells were recognized by one of the DR4, DW14-specific T-cell clones, T431, were analyzed. All of these patients shared a common sequence in the third hypervariable region of the DR beta 1 chain gene. The sequence identified is the one normally associated with DR4, Dw14 and DR1. Patients and DR4-positive controls whose cells did not stimulate this clone did not share this sequence. These results suggest that this hypervariable region might be an important contribution to a restriction site for the putative causative agent(s) in rheumatoid arthritis.

Amino Acid Sequence↗
PubMed1990

Allelic diversification at the class II DQB locus of the mammalian major histocompatibility complex.

The allelic diversity at HLA class II loci either arose after the divergence of hominoid lineages or, alternatively, the polymorphism was present before speciation and has been maintained by selection. Here, we report the use of oligonucleotide primers to amplify, by the polymerase chain reaction, and sequence the polymorphic second exon of the DQB locus from 11 species, spanning more than 40 million years of mammalian evolution. Phylogenetic analysis reveals that of the four human DQB allelic types (DQB1-B4), three (DQB1-3) were found in chimpanzee and gorilla and two (DQB3 and -4) were identified in the rhesus monkey, suggesting that some of these types are 5-20 million years old. The ratio of replacement to silent substitutions was calculated between members of the same allelic type from different species. These results suggest that the evolution of the DQB3 allelic type is more constrained than that of the DQB1 allelic type; both evolve more slowly than the DXB locus, a linked but presumably nonexpressed locus. Further, the clustering of allelic subtypes by species in the phylogenetic tree indicates that allelic diversification has occurred subsequent to the divergence of hominoids. Finally, some haplotype combinations of DQA and DQB alleles are common to several hominoid species and may have been maintained for at least 5 million years.

Alleles↗
PubMed1990

HLA-DQ beta sequence polymorphism and genetic susceptibility to IDDM.

The analysis of HLA-DQ beta nucleotide sequence polymorphism in insulin-dependent diabetes mellitus (IDDM) patients and control subjects suggests a role for the DQ beta-chain in genetic susceptibility. Sequence determination and oligonucleotide hybridization was carried out on enzymatically amplified DNA from various HLA-DR-typed individuals, including the rare class of DR2+ patients. In the analysis of DQ beta variation in DR4, DRw6, and DR2 haplotypes, a correlation was observed between the presence of the negatively charged residue Asp at position 57 and low susceptibility and the presence of an Ala (DR4), Val (DRw6), or Ser (DR2) and higher susceptibility. However, important exceptions to this pattern have been identified in the analysis of heterozygous DR1/4 IDDM patients. In these individuals, susceptibility appears to correlate with specific DR beta l alleles (Dw4) on the DR4 haplotype, rather than with the DQ beta allele (DQB3.2) that contains Ala at position 57. The DQ beta alleles found in some Chinese IDDM patients also proved discordant with the position-57 correlations. Thus, although there is a general correlation between the residue at position 57 of the DQ beta-chain and IDDM susceptibility, these data do not support the notion that Asp 57 confers complete resistance or protection to IDDM. In general, these results suggest that IDDM susceptibility is conferred by specific combinations of DQ beta and DR beta sequences.

Alleles↗
PubMed1989

A combination of a particular HLA-DP beta allele and an HLA-DQ heterodimer confers susceptibility to coeliac disease.

Coeliac disease is an autoimmune disease of the intestinal mucosa, elicited by ingestion of wheat gluten in genetically susceptible individuals. Susceptibility to coeliac disease has been associated with the serologically defined variants DR3 and DR7 of the class II antigens encoded by the HLA-D region. Three related class II antigens, each consisting of an alpha and a beta glycoprotein chain, have been identified and are designated HLA-DR, HLA-DQ, and HLA-DP. These highly polymorphic transmembrane proteins bind peptides derived from the processing of foreign antigens and present them to T lymphocytes; they also influence the specificity of the mature T-cell repertoire. The role of HLA-DP polymorphism in susceptibility has not been as fully explored as that of the other class II antigens because of the complexity of the primed lymphocyte typing (PLT) method for determining DPw specificities. Here we use a new DNA-based method of HLA-DP typing to analyse the distribution of DP beta alleles in a group of coeliac disease patients and healthy controls. Two specific DP beta alleles (DPB4.2 and DPB3) are increased in the patient population. Comparison of the DP beta sequences suggests that the polymorphic residues at position 69 and at 56 and 57 may be critical in conferring susceptibility. Further, the contribution of the susceptible DP beta alleles appears to be independent of linkage to the previously reported DR3 and DR7 markers for coeliac disease. The distribution of DQ alpha and beta alleles in patients suggests that a specific DQ heterodimer may be responsible for the observed DR associations. Individuals with both this DQ antigen and a specific DP beta allele are at increased risk for coeliac disease.

Alleles↗
PubMed1989

Molecular genetic analysis of 178 I-Abm12-reactive T cells.

We have studied the genetic diversity of the TCR repertoire to the murine alloantigen I-Abm12 by generating a panel of 178 C57BL/10-derived I-Abm12-reactive T cell hybridomas. The expression of V alpha and V beta gene families was examined in this panel and the frequency of expression of V beta, but not ofV alpha, gene families differed significantly from that observed in a companion panel of random C57BL/10-derived hybridomas. The V beta 5 gene family was expressed significantly less frequently while the V beta 14, V beta 15, and V beta 16 genes were expressed significantly more frequently in the panel of I-Abm12-reactive than in the panel of random hybridomas. The junctional regions (VJ alpha and VDJ beta) of TCR V alpha and V beta genes from selected I-Abm12-specific hybridomas were amplified using the polymerase chain reaction, and directly sequenced. Surprisingly, no conserved J alpha, D beta, J beta, or N region-encoded sequences among these selected I-Abm12-reactive TCRs were identified. Thus, the T cell response to an I-A alloantigen that differs by only three amino acid residues from the I-A molecule of the responding strain is genetically complex but nonrandom. We have estimated that the repertoire to this alloantigen is comprised of at least 37 different TCRs.

Amino Acid Sequence↗
PubMed1989

Polymerase chain reaction.

The PCR, like recombinant DNA technology, has had an enormous impact in both basic and diagnostic aspects of molecular biology because it can produce large amounts of a specific DNA fragment from small amounts of a complex template. Recombinant DNA techniques create molecular clones by conferring on a specific sequence the ability to replicate by inserting it into a vector and introducing the vector into a host cell. PCR represents a form of "in vitro cloning" that can generate, as well as modify, DNA fragments of defined length and sequence in a simple automated reaction. In addition to its many applications in basic molecular biological research, PCR promises to play a critical role in the identification of medically important sequences as well as an important diagnostic one in their detection.

Animals↗