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H Erlich

Publications and source records attributed to H Erlich.

At least 37 records · Page 2Linked to original sources
PubMed1995

Molecular mapping of a recombination hotspot located in the second intron of the human TAP2 locus.

Recombination across the HLA class II region is not randomly distributed, as indicated by both strong linkage disequilibrium within the 100 kb encompassing the DRB1-DQA1-DQB1 loci and complete equilibrium between TAP1 and TAP2, the closest variant sites of which are < 15 kb. In an attempt to explain these observations, 39 novel polymorphic markers in a region encompassing the TAP, LMP, and DOB genes were used to delineate the site of crossover in 11 class II recombinant chromosomes. SSCP demonstrated that two recombination events occurred within an 850-bp interval in the second intron of TAP2, which separates the variant sites of TAP1 and TAP2. These data indicate the presence of a recombination hotspot, the first to be identified from the analysis of familial transmission in the human major histocompatibility complex. The region of crossover was cloned and sequenced from one of the recombinants, further defining the crossover site to a 138-bp segment nested within the 850-bp region. This represents the most precisely defined region of recombination in the human genome.

ATP Binding Cassette Transporter, Subfamily B, Mem↗
PubMed1994

Higher autoantibody levels and recognition of a linear NH2-terminal epitope in the autoantigen GAD65, distinguish stiff-man syndrome from insulin-dependent diabetes mellitus.

The smaller form of the GABA-synthesizing enzyme glutamic acid decarboxylase (GAD65) is a major autoantigen in two human diseases that affect its principal sites of expression. Thus, destruction of pancreatic beta cells, which results in insulin-dependent diabetes mellitus (IDDM), and impairment of GABA-ergic synaptic transmission in Stiff-Man syndrome (SMS) are both characterized by circulating autoantibodies to GAD65. Anti-GAD65 autoantibodies in IDDM are predominantly directed to conformational epitopes. Here we report the characterization of humoral autoimmune responses to GAD65 in 35 SMS patients, of whom 13 (37%) also had IDDM. All SMS patients immunoprecipitated native GAD65 and the main titers were orders of magnitude higher than in IDDM patients. Furthermore, in contrast to the situation in IDDM, autoantibodies in 35 of 35 (100%) of SMS patients recognized denatured GAD65 on Western blots. Two major patterns of epitope specificity were identified on Western blots. The first pattern, detected in 25 of 35 SMS patients (71%), of whom 11 had IDDM (44%), was predominantly reactive with a linear NH2-terminal epitope residing in the first eight amino acids of GAD65. Nine of nine individuals who were HLA-haplotyped in this group carried an IDDM susceptibility haplotype and HLA-DR3, DQw2 was particularly abundant. The second pattern, detected in 10 of 35 patients (29%) of whom two had IDDM (20%), included reactivity with the NH2-terminal epitope plus strong reactivity with one or more additional epitope(s) residing COOH-terminal to amino acid 101. The second epitope pattern may represent epitope spreading in the GAD65 molecule, but may also include some cases of epitope recognition associated with IDDM resistant HLA-haplotypes. The principal NH2-terminal linear epitope in GAD65 distinguishes the reactivity of SMS and IDDM autoantibodies and may be a determinant of pathogenicity for GABA-ergic neurons. The greater magnitude and distinct specificity of the humoral response to GAD65 in SMS may reflect a biased involvement of the T helper cell type 2 (Th2) subset of CD4+ T cells and antibody responses, whereas IDDM is likely mediated by the Th1 subset of CD4+ T cells and cytotoxic T cell responses.

Adult↗
PubMed1993

Genetic analysis using polymerase chain reaction-amplified DNA and immobilized oligonucleotide probes: reverse dot-blot typing.

The reverse dot-blot method is a simple and rapid diagnostic procedure that allows screening of sample for a variety of mutations/polymorphisms in a single hybridization reaction. Several methods of immobilizing the oligonucleotide probes are discussed. The reverse dot-blot method has several unique properties that are valuable in a diagnostic setting: (1) the typing results from a single sample can be located on a single strip. This facilitates scanning and interpretation of the probe reactivity patterns and minimizes the potential for user error. (2) The test can utilize premade typing strips. This minimizes user labor as well as error potential and allows the use of standardized reagents. (3) Unlike dot-blot/oligonucleotide typing, only the PCR product is labeled, eliminating the potential problem of probes labeled to different specific activities. This method has already been used in the areas of forensic genetic typing (the HLA-DQ alpha Amplitype test), tissue typing for transplantation (the HLA-DR beta) test, cystic fibrosis screening, as well as in a variety of research applications.

Amino Acid Sequence↗
PubMed1993

Analysis of HLA class II polymorphism using polymerase chain reaction.

A simple, rapid, and precise method of typing HLA class II polymorphism would be valuable in the areas of disease susceptibility, tissue transplantation, individual identification, and anthropological genetics. Herein, we describe a method of analyzing class II sequence polymorphism based on polymerase chain reaction (PCR) amplification and hybridization with oligonucleotide probes. Many more DNA-defined alleles at the class II loci have been identified than can be distinguished by conventional serologic typing. Consequently, matching transplant donors and recipients by PCR/oligonucleotide typing should reduce graft rejection and graft-vs-host disease. Also, the ability to identify alleles conferring genetic predisposition to specific diseases (eg, insulin-dependent diabetes mellitus) is significantly enhanced by distinguishing the many alleles or "subtypes" within a serologic type (eg, DR4). One valuable property of sequence-based HLA typing strategies, like oligonucleotide probe hybridization, is that they reveal how and where two alleles differ, not simply that they can be operationally distinguished. The nature and location of HLA polymorphisms appears to be critical in disease association studies and are important in tissue typing for transplantation. New alleles at the DRB1, DPB1, and DQB1 loci are likely to be identified as this technology is applied to more and more samples, particularly in nonwhite ethnic groups. A new allele is uncovered as an unusual pattern of probe binding and then confirmed by sequencing. This pattern is observed because class II polymorphism is localized to specific regions and virtually all "new" alleles represent "shuffled" combinations of polymorphic sequences found in previously known alleles. Since these polymorphisms are in the region of probe binding, these new alleles can be detected without increasing the probe panel. Obviously, any new allele with a new polymorphic sequence in a region for which typing probes are not available would not be revealed by oligonucleotide typing. With the PCR primers and probes described here, 7 DQ alpha 1 alleles, 15 DQ beta 1 alleles, 18 DPB1 alleles, and 32 DRB1 alleles are distinguished. Additional primers and/or probes can, of course, increase the allelic discrimination of PCR/oligonucleotide probe typing. These horseradish peroxidase-labeled oligonucleotide probes are stable (> 2 years when stored at 4 degrees C) and the typing system is simple and robust. Although this dot blot/oligonucleotide hybridization procedure is a powerful and precise method of HLA class II typing, the complexity of the procedure increases as the number of probes required for analysis increases.(ABSTRACT TRUNCATED AT 400 WORDS)

Alleles↗
PubMed1992

HLA alleles in the population of Cartagena (Colombia).

It is known from historical records that the population of Cartagena (Colombia) is a mixture of caucasoids (Spaniards) and negroids (Africans) that is named mulattos. The present study was carried out to determine the distribution of HLA antigens in this population. A sample of 370 individuals was HLA serologically typed; 91 of them were typed for Complement alleles and 47 were DNA typed using PCR amplification and oligonucleotide probes for the HLA-DQA1 locus. The analysis of the allele and haplotype frequencies obtained, together with the genetic distances calculated with other populations, showed that most of the HLA genes of mulattos are from negroids and caucasoids. The Asian and Amerindian contribution is small, consistent with the historical and socioanthropological documentation. For example, A23, Aw34, Aw36 and Bw45, as well as DQA1*0101, DQA1*0301 and BfF have frequencies similar to that observed in negroids. In contrast, many significant differences were observed when comparing the antigen and allele frequencies of mulattos with that of Asians and Amerindians. Our results add evidence to the notion that the arrival of Spaniards in America 500 years ago established several racial mixtures. The HLA characterization of one of these, mulattos, will allow a better application of the HLA technology in the fields of basic research, organ transplantation and forensic medicine among this population.

Alleles↗
PubMed1992

Prognostically significant heterogeneity of cytoplasmic islet cell antibodies in relatives of patients with type I diabetes.

A significant proportion of relatives of patients with insulin-dependent (type I) diabetes with high titers of cytoplasmic islet cell autoantibodies (ICAs) do not progress to overt diabetes with up to 8 yr of follow-up. This may reflect that follow-up of such relatives has not been long enough to observe diabetes, that despite expression of identical ICAs, some relatives will not progress to diabetes; or that there is heterogeneity in what is identified as ICA. We identified a subset of ICA that was restricted in its species (not reacting with mouse islets) and cell-type reactivity within islets (beta-cell specific). Only one of eight relatives whose sera had the restricted pattern of reactivity progressed to overt diabetes, and on sequential evaluation, all but the one relative who progressed to diabetes have maintained normal first-phase insulin secretion to intravenous glucose. In contrast, by life-table analysis, 70% of relatives expressing nonrestricted ICA became diabetic within 5 yr of follow-up (1 of 8 vs. 16 of 25 diabetic at last follow-up, P less than 0.02). Moreover, preliminary data suggest a significant association of the human leukocyte antigen DQB1*0602 allele of DR2 haplotypes with the restricted ICA pattern (4 of 5 DQB1*0602 restricted vs. 0 nonrestricted ICA, P = 0.006). We propose that expression of a genetically determined restricted ICA pattern confers a markedly lower risk for progression to diabetes.(ABSTRACT TRUNCATED AT 250 WORDS)

Adolescent↗
PubMed1992

Polymerase chain reaction (PCR) amplification and human leukocyte antigen (HLA)-DQ alpha oligonucleotide typing on biological evidence samples: casework experience.

The polymerase chain reaction (PCR) method of specific gene amplification was used in casework to synthesize millions of copies of the polymorphic second exon of the human leukocyte antigen (HLA)-DQ alpha (or DQA1) locus from a variety of evidence samples. The HLA-DQ alpha allelic variants in the amplified deoxyribonucleic acid (DNA) were determined in a rapid non-radioactive test by hybridization to sequence-specific oligonucleotide probes in both the dot-blot and reverse dot-blot formats. This genetic typing system has been subjected to blind proficiency testing; the performance of this test in the analysis of experimentally mixed samples was also evaluated. As of August 1990, over 250 cases have been tested and more than 2000 individual evidence (bloodstains, semen stains, individual hairs, bone fragments, and tissue sections) and reference samples have been analyzed. The first 198 of these cases are summarized in this paper; in 65% of the cases with conclusive results a suspect was included, and in 35%, all suspects were excluded. Individual cases as well as some of the general issues relating to forensic science analysis and this genetic typing system are discussed. The high rate of exclusion reported here combined with the ability of PCR to type old evidence samples suggests the relevance of this genetic test for postconviction review; two cases in which the convicted suspect was excluded are discussed.

Adolescent↗
PubMed1991

Specific HLA-DQA and HLA-DRB1 alleles confer susceptibility to Sjögren's syndrome and autoantibody production.

Primary Sjögren's syndrome (1. SS) is a systemic autoimmune disease characterized by lymphocytic infiltration of the salivary glands and autoantibody production. In order to identify genetic factors that play a role in pathogenesis and predict extent of disease, we used Southern blot and polymerase chain reaction (PCR) methods to detect polymorphisms of the HLA-DRB1 (DR), HLA-DRB3 (DRw52), and HLA-DQA1 genes among 75 Caucasoid 1. SS patients and 150 Caucasoid controls living in the same geographic region of Southern California. We found significantly increased frequency of HLA-DR3 (P less than .001), HLA-DW52a (P less than .001), and HLA-DQA4 (P less than .05), in comparison to normal controls. Also, an increased frequency of heterozygosity for HLA-DQA1/DQA4 (P less than .05) was present among 1. SS patients. Autoantibodies to SS-A and to SS-B were significantly associated with DR3 (P less than .001), HLA-DQA4, (P less than .05), and DQA4/DQA1 heterozygotes (P less than .01). Among the 1. SS patients, clinical and laboratory features such as hypergammaglobulinemia, symmetric peripheral neuropathy, and hypothyroidism were significantly associated with HLA-DR3 (P less than .01) but not with HLA-DR2 (P greater than .10). In comparison, 1. SS patients with leukocytoclastic vasculitis were more frequently HLA-DR2 (P less than .05). These results using PCR methods confirm and extend prior studies that have used serologic methods.

Adolescent↗
PubMed1991

MHC class-II molecules and autoimmunity.

Molecular and genetic studies of HLA class-II genes provide new insights into the basis for MHC associations with autoimmunity. Polymorphisms among class-II genes identify specific haplotypes associated with autoimmune diseases such as type-I diabetes, rheumatoid arthritis, celiac disease, and pemphigus vulgaris. In some cases, single genes within those haplotypes are themselves implicated in disease susceptibility. Interactions, both cis and trans, between candidate susceptibility genes suggest a number of possible mechanisms critical for autoimmune triggering events involving class-II molecules. Amino acid sequence comparisons between products of candidate susceptibility genes and other class-II genes pinpoint a limited number of critical sites within HLA molecules which appear to be responsible for pathogenic events.

Amino Acid Sequence↗
PubMed1991

[Relationship between HLA D region gene and primary Sjögren's syndrome].

Primary Sjögren's syndrome is characterized by lymphocytic infiltration of the salivary and lacrimal glands and by autoantibody production. In order to identify the genetic factors that play a role in the pathogenesis and to predict the progress of the disease we used the restriction fragment length polymorphism and polymerase chain reaction method to detect the polymorphism of HLA-DRB1, DRB3, DQA and DQB genes among 75 Caucasoid primary SS patients, and compared the results with normal controls living in the same area. We found (1) significant increased frequency of HLA-DR3 (54%) and DRw8 (13%) (P less than 0.001); (2) increased frequency of the HLA-DRB3 allele DRw52a and of the HLA-DQA allele DQA 4 (P less than 005); (3) increased frequency of heterozygosity for DQA 1/DQA 4 (P less than 005); (4) autoantibody SSA and SSB associated with DR3 (P less than 002 and P less than 0001 respectively).

Adolescent↗
PubMed1991

A dimorphic 4-bp repeat in the cystic fibrosis gene is in absolute linkage disequilibrium with the delta F508 mutation: implications for prenatal diagnosis and mutation origin.

The gene causing cystic fibrosis (CF) has been recently cloned, and the major mutation (delta F508) accounting for approximately 70% of CF chromosomes has been uncovered. We have identified at the 3' end of intron 6 in the CF gene a 4-bp tandem repeat (GATT) that exhibits interesting features. First, PCR screening of 103 normal individuals revealed that the repeat exists only in two polymorphic allelic forms, either as a hexamer or a heptamer. These two alleles are in Hardy-Weinberg equilibrium and predict a heterozygote frequency of 41% (p[seven repeats] = .71; q [six repeats] = .29). Second, the allele with six repeats was found linked to delta F508 on all 76 CF chromosomes investigated, demonstrating strong linkage disequilibrium and suggesting that delta F508 had originated on the gene bearing six repeats. Third, when the repeat alleles are linked to the DNA markers XV2c and KM19, extended haplotypes are generated. These new haplotypes become informative in situations in which prenatal diagnosis cannot be performed solely with XV2c and KM19. Since this repeat marker is located in the CF gene and would be very less likely to recombine with the gene, it can serve as a valuable DNA marker for haplotype analysis. A possible crossover, however, was identified between XV2c and KM19, transferring delta F508 to a different haplotype.

Alleles↗
PubMed1989

Rapid detection and sequencing of alleles in the 3' flanking region of the interleukin-6 gene.

The 3' flanking region of the interleukin 6 gene is polymorphic due to insertions of different size. Within this region lies a sequence of approximately 500 base pairs that is AT rich. Based on flanking sequence information we have constructed oligonucleotides which prime the polymerase chain reaction (PCR) and amplify this AT rich region. The amplification products visualized by agarose gel electrophoresis gave fragment sizes for both homozygous and heterozygous individuals that were concordant with those observed by conventional genomic blotting techniques. Alleles that could not be typed by Southern analysis were resolved with this approach. These results illustrate the value of PCR for the rapid detection of length polymorphisms such as those due to variable numbers of tandem repeats. In contrast to RFLP analysis this procedure takes less than a day to perform, is cheaper, avoids the use of radioactivity and requires far less substrate DNA. Three different human alleles were sequenced, and differences were detected that were due to both large duplications and loss of one or two bases, suggesting that AT rich regions identify highly polymorphic loci. The same primers also amplified non-human primate DNA, allowing a comparison of the human sequence with that of the common chimpanzee and baboon.

Alleles↗
PubMed1989

A beta-thalassemia gene caused by a 290-base pair deletion: analysis by direct sequencing of enzymatically amplified DNA.

The base composition around a recently detected deletion in the human beta-globin gene was determined by direct DNA sequencing of an enzymatically amplified DNA segment. The deletion removes 290 base pairs (bp), including the entire exon 1 and the mRNA cap site. In the vicinity of the deletion endpoints, the normal beta-globin gene contains direct and inverted repeats which may have taken part in generation of this deletion.

Base Sequence↗