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Biomedical subjects

M B Frank

Publications and source records attributed to M B Frank.

At least 37 records · Page 2Linked to original sources
PubMed1994

Cooperative association of T cell beta receptor and HLA-DQ alleles in the production of anti-Ro in systemic lupus erythematosus.

The immunogenetics of the autoantibody response to Ro (or SS-A) have been explored in patients with systemic lupus erythematous. Data show that alleles of the T cell beta receptor and HLA-DQ loci are cooperatively associated with the presence of anti-Ro autoantibodies in systemic lupus erythematosus. Identification of HLA-DQ by oligonucleotide probe binding to polymerase chain reaction products demonstrates that the combination of DQB1*0201 and one of DQA1*0101, DQA1*0102, or DQA1*0103 is associated with anti-Ro. Patients possessing a particular pair of T cell receptor beta restriction enzyme polymorphisms along with these specific HLA-DQ alleles produce quantitatively more anti-Ro as measured by a sensitive solid-phase immunoassay than patients without these T cell receptor and DQ alleles. Other work has shown that the autoimmune response is directed against the human Ro antigen. These results are consistent with a central role in the disregulation of autoimmunity involving a trimolecular complex composed of the autoantigen bound by a HLA-DQ molecule which, together, are bound in turn by T cells which express a particular subset of T cell receptors.

Alleles↗
PubMed1994

The location of a disease-associated polymorphism and genomic structure of the human 52-kDa Ro/SSA locus (SSA1).

Sera from approximately 30% of patients with systemic lupus erythematosus (SLE) contain high titers of autoantibodies that bind to the 52-kDa Ro/SSA protein. We previously detected polymorphisms in the 52-kDa Ro/SSA gene (SSA1) with restriction enzymes, one of which is strongly associated with the presence of SLE (P < 0.0005) in African Americans. A higher disease frequency and more severe forms of the disease are commonly noted among these female patients. To determine the location and nature of this polymorphism, we obtained two clones that span 8.5 kb of the 52-kDa Ro/SSA locus including its upstream regulatory region. Six exons were identified, and their nucleotide sequences plus adjacent noncoding regions were determined. No differences were found between these exons and the coding region of one of the reported cDNAs. The disease-associated polymorphic site suggested by a restriction enzyme map and confirmed by DNA amplification and nucleotide sequencing was present upstream of exon 1. This polymorphism may be a genetic marker for a disease-related variation in the coding region for the protein or in the upstream regulatory region of this gene. Although this RFLP is present in Japanese, it is not associated with lupus in this race.

Amino Acid Sequence↗
PubMed1994

Autoantibodies of neonatal lupus erythematosus.

The most common manifestations of neonatal lupus erythematosus (NLE) are cutaneous lupus and congenital heart block. Autoantibodies to Ro/SSA occur in almost all cases of NLE. The autoantibody response to Ro/SSA is complex, and antibodies may be detected to 60-kD Ro/SSA, 52-kD Ro/SSA, La/SSB, and U1 ribonuclear protein in anti-Ro/SSA-positive sera. Which of these anti-Ro/SSA-related autoantibody specificities are important in the clinical expression of NLE is not conclusively established. We examined the autoantibody specificities in 20 maternal NLE sera to determine whether autoantibody specificities correlate with the clinical findings and to evaluate the relative importance of autoantibodies to the different Ro/SSA-associated proteins. Autoantibodies were examined using immunodiffusion, immunoblotting, and enzyme-linked immunosorbent assay. Eleven babies had NLE skin disease, 11 had heart block, and two had both skin disease and heart block. All 20 maternal sera had antibodies to 60-kD Ro/SSA. Eighteen of the 20 had antibodies to 52-kD Ro/SSA, nine had antibodies to La/SSB, and one had antibodies to U1 ribonuclear protein. The prevalence of anti-La/SSB was the same in the skin-disease and heart-block subsets of NLE. Titers of anti-60-kD Ro/SSA were significantly (p < 0.02) lower in NLE skin disease maternal sera than in the NLE heart-block maternal sera. These results point out the importance of 60-kD Ro/SSA as a potential target in NLE. We speculate that the lower titers of anti-60-kD Ro/SSA in the sera from mothers of babies with skin disease may be due to substantial deposition of antibodies in the mothers' and babies' skin, leading to lower circulating titers, or may reflect a lower threshold for development of skin disease than for heart block.

Antibodies, Antinuclear↗
PubMed1994

Epitope mapping of the 52-kD Ro/SSA autoantigen.

Autoantibodies to Ro/SSA are commonly found in sera of patients with systemic lupus erythematosus (SLE) and primary Sjögren's syndrome. The presence of these antibodies is related to lymphopenia, photosensitive dermatitis, and pulmonary and renal disease, suggesting that they have an immunopathologic role [1-6]. We previously isolated a cDNA clone which encodes the 52-kD human Ro/SSA protein. In this study we have determined the number and location of epitopes recognized by SLE sera using recombinant proteins encoded by the full-length or overlapping subclones of this cDNA. An immunodominant epitope was detected using Western blots and ELISA on the NH2-terminal side of this protein's putative leucine zipper. The data suggest that 11 amino acids are critical for the recognition of this molecule by these autoantibodies. Although the titres of anti-52-kD Ro/SSA antibodies vary between different patient sera, no heterogeneity in the location of antigenic epitopes to which their autoantibodies bound was detected. This homogeneous pattern of reactivity to a single rather than multiple regions of this protein is unusual for lupus autoantigens which have been identified, and suggests that these antibodies may have arisen as by a cross-reaction to an epitope on another molecule.

Amino Acid Sequence↗
PubMed1994

The autoantibody response to Ro/SSA in cutaneous lupus erythematosus.

BACKGROUND AND DESIGN: Seventeen patients with subacute cutaneous lupus erythematosus (SCLE) were compared with 15 patients with discoid lupus erythematosus (DLE) to evaluate the relationship of 60- and 52-kd Ro/SSA autoantibodies to the clinical diagnosis and to evaluate assays for anti-Ro/SSA. RESULTS: All serum samples from patients with SCLE had precipitating anti-Ro/SSA antibodies in immunodiffusion, and all had high titer anti-60-kd Ro/SSA in enzyme-linked immunosorbent assay. Immunoblotting was inadequately sensitive for detecting anti-60-kd Ro/SSA. Fifteen patients with SCLE had anti-52-kd Ro/SSA (11 high titer, four low titer). Only one of the 15 patients with DLE had precipitating, high-titer anti-Ro/SSA. Nine other patients with DLE had low-titer anti-60-kd Ro/SSA, and four had low-titer anti-52-kd Ro-SSA. Low-titer anti-Ro/SSA did not confer an increased risk for photosensitivity in the DLE group. CONCLUSIONS: High-titer, precipitating antibodies to Ro/SSA are typical of SCLE and unusual in DLE. Low-titer, nonprecipitating antibodies to Ro/SSA are common in DLE and could be an indication of pathogenic factors shared with SCLE. However, low titers of anti-Ro/SSA do not confer a significant risk for SCLE skin lesions. For the purpose of clinical evaluation of skin disease, immunodiffusion assays for anti-Ro/SSA are cost-effective and informative.

Autoantigens↗
PubMed1993

The mapping of the human 52-kD Ro/SSA autoantigen gene to human chromosome 11, and its polymorphisms.

Autoantibodies to Ro/SSA occur in nearly half of the patients with systemic lupus erythematosus and are associated with lymphopenia, photosensitive dermatitis, and pulmonary and renal disease, which suggests that they have an immunopathologic role. The majority of Ro/SSA precipitin-positive patients produce serum antibodies that bind to the 60-kD and 52-kD Ro/SSA proteins. We previously isolated and determined the nucleotide sequence of a cDNA clone that encodes the 52-kD form of the human Ro/SSA protein. In the present study, we have determined the chromosomal location of the gene by in situ hybridization to the end of the short arm of chromosome 11. Hybridization of portions of the cDNA probe to restriction enzyme-digested DNA indicated the gene is composed of at least three exons. The exon encoding the putative zinc fingers of this protein was found to be distinct from that which encodes the leucine zipper. An RFLP of this gene was identified and is associated with the presence of lupus, primarily in black Americans.

Autoantigens↗
PubMed1992

Purification, primary structure, bacterial expression and subcellular distribution of an oocyte-specific protein in Xenopus.

This study defines a novel Xenopus laevis protein (P100) that has recently been shown to be recognized by scleroderma patient sera. Using a combination of differential solubility in detergents, hydroxyapatite chromatography and one-dimensional PAGE, P100 was purified to apparent homogeneity and the amino acid sequence was obtained. An oligonucleotide derived from this sequence was used to clone P100 cDNA through a polymerase-chain-reaction cloning strategy. The entire P100 cDNA sequence was determined, identifying a novel 83,000-Da protein. Two alleles for P100 were transcribed in the oocyte, with only one predicted amino acid change between them. Bacterial expression of a clone containing the entire P100 coding region produced a protein that migrated at a mass 15% greater than that predicted from the amino acid sequence, indicating an aberrant electrophoretic mobility. The mRNA transcript for P100 was only expressed during the previtellogenic stages of oogenesis (stages I and II) and was absent from other Xenopus tissues. Similarly, the P100 protein was found only in Xenopus oocytes and was localized to the cytoplasm of these cells. P100 irreversibly bound single-stranded-DNA--cellulose but not double-stranded-DNA--cellulose. These data demonstrate the presence of a novel oocyte-specific protein in Xenopus.

Amino Acid Sequence↗
PubMed1992

Cloning of a complementary DNA coding for the 100-kD antigenic protein of the PM-Scl autoantigen.

Anti-PM-Scl antibodies are associated with polymyositis-scleroderma overlap or either disease alone. Among sera from 39 patients with anti-PM-Scl, 23 recognized the 100-kD band in immunoblot against HeLa cell extract, 16 of which also stained the 70-kD band. A human thymocyte lambda gt11 cDNA expression library was screened with anti-PM-Scl serum, and two clones were identified whose products reacted with 33 and 37 of 39 anti-PM-Scl sera, respectively, but none of 26 negative control sera. Affinity-purified antibody reacting specifically with plaques of the clone stained the 100-kD band on immunoblot, reacted with nucleoli of HEp-2 cells, and immunoprecipitated the PM-Scl protein complex. Partial sequences of both inserts were identical. One insert was fully sequenced, and additional 5' and 3' sequence was obtained using a gene-specific primer to form a cDNA with HeLa cell RNA as template followed by PCR. The complete nucleotide sequence included 2,739-bp coding for a predicted full-length protein of 98,088 D. There was no homology with the PM-Scl 75-kD protein and no significant homology with other proteins. A mixed-charge cluster was identified, with 22 charged amino acids of 37. In conclusion, the full-length cDNA sequence was determined coding for the PM-Scl 100-kD protein, the most commonly antigenic protein of the PM-Scl complex.

Amino Acid Sequence↗
PubMed1992

Autoantibodies to the Ro/SSA autoantigen are conformation dependent. II: Antibodies to the denatured form of 52 kD Ro/SSA are a cross reacting subset of antibodies to the native 60 kD Ro/SSA molecule.

The immune response to the Ro/SSA particles is conformation dependent. In sera with only anti-Ro/SSA precipitins, the autoantibodies to the 60 kD Ro/SSA are largely to the native 60 kD Ro/SSA while autoantibodies to the 52 kD Ro/SSA particle when present are exclusively to the denatured 52 kD Ro/SSA particle. Antibodies eluted from a recombinant 52 kD Ro/SSA fusion protein reacted in a sandwich ELISA which only measures antibody to native 60 kD Ro/SSA antigens and this reaction is largely inhibited by native homogeneous 60 kD Ro/SSA. In addition, antibody binding to the 52 kD Ro/SSA antigen in Western blot is also strongly inhibited by native 60 kD Ro/SSA. These experiment strongly suggest that reactivity of denatured 52 kD Ro/SSA antigen represents a cross reaction with autoantibodies directed to the native 60 kD Ro/SSA antigen. As a corollary of these experiments, data are presented that suggest the hY-RNAs are not associated with the 52 kD Ro/SSA protein but only with the 60 kD Ro/SSA protein. These data are consistent with the hypothesis that the autoanti-Ro/SSA response is driven by native 60 kD Ro/SSA and the immune response to denatured 52 kD Ro/SSA is largely a cross-reactive subset of the immune response to native 60 kD Ro/SSA.

Antibody Specificity↗
PubMed1991

Protein heterogeneity in the human Ro/SSA ribonucleoproteins. The 52- and 60-kD Ro/SSA autoantigens are encoded by separate genes.

Two cDNA clones encoding the 52-kD form of a protein present in human Ro/SSA ribonucleoprotein complexes were cloned from a lambda gt11 human thymocyte cDNA library. These clones reacted with lupus patient sera which had anti-52-kD Ro/SSA antibodies, and with affinity-purified anti-52-kD Ro/SSA antibodies. Moreover, affinity-purified antibodies isolated from isopropyl-beta-D-thiogalactopyranoside-induced proteins of these clones reacted only with the 52-kD protein of lymphocytes in Western blots and precipitated Ro/SSA hY RNAs, confirming that the clones encode a 52-kD Ro/SSA antigen. The cDNA contains a single open reading frame of 1,425 nucleotides and encodes a predicted 475-amino acid polypeptide with a molecular mass of 54,108 D. This protein appears unique in that both a zinc finger and leucine zipper motif are present on this protein. Surprisingly, no homology was found between the 52-kD Ro/SSA gene or protein and three published 60-kD Ro/SSA sequences. However, significant similarity of the 52-kD Ro/SSA was detected with human rfp and mouse rpt-1. These three proteins each contain similar zinc finger motifs in approximately their first 145 amino acid residues. The cDNA and the protein expressed therefrom are useful in the analysis of the structural and functional properties of this autoantigen.

Amino Acid Sequence↗
PubMed1990

Anti-Ro(SSA) autoantibodies are associated with T cell receptor beta genes in systemic lupus erythematosus patients.

Several of the heterogeneous clinical manifestations of systemic lupus erythematosus have been associated with specific autoantibodies. Associations between HLA class II antigens and autoantibodies to the ribonucleoproteins Ro(SSA) and La(SSB) have been reported in these patients. Because HLA class II molecules present antigen to T cell receptors (TCRs), we have searched for a TCR gene associated with the production of anti-Ro(SSA) antibodies. A pair of restriction fragment length polymorphisms (RFLPs), one of which hybridizes to the TCR constant region C beta 1 and the other to the C beta 2 gene, has been identified, suggesting these may be genotypic markers for an extended region of the TCR beta locus. This RFLP pair occurs in 76% of patients with Ro(SSA) precipitins, 84% of anti-Ro(SSA)-positive patients lacking La(SSB) precipitins, but only 41% of the patients lacking both precipitins (P = 0.0004). This disproportionate occurrence in a subset of lupus patients indicates that these RFLPs are not disease susceptibility markers, but rather are important markers for TCR genes whose products are involved in the production of anti-Ro(SSA) antibodies. The majority of patients who have these RFLPs and HLA class II antigens previously associated with the anti-Ro(SSA) response make this antibody, suggesting that interactions between products of these loci occur in response to Ro(SSA).

Adult↗
PubMed1990

HLA-DQ gene complementation and other histocompatibility relationships in man with the anti-Ro/SSA autoantibody response of systemic lupus erythematosus.

A strong gene interaction between HLA-DQ1 and DQ2 alleles has been associated with anti-Ro/SSA autoantibodies (Harley, J.B., M. Reichlin, F. C. Arnett, E. L. Alexander, W. B. Bias, and T. T. Provost. 1986. Science [Wash. DC]. 232:1145-1147; Harley, J. B., A. S. Sestak, L. G. Willis, S. M. Fu, J. A. Hansen, and M. Reichlin. 1989. Arthritis Rheum. 32:826-836; Hamilton, R. G., J. B. Harley, W. B. Bias, M. Roebber, M. Reichlin, M. C. Hochberg, and F. C. Arnett. 1988. Arthritis Rheum. 31:496-505). To test a gene complementation mechanism for these results, restriction fragment length polymorphisms (RFLP) of the DQ alpha and DQ beta genes have been related to Ro/SSA precipitins in patients with systemic lupus erythematosus. In this study Ro/SSA precipitins are related to the simultaneous presence of a particular pair of RFLPs. A DQ alpha RFLP associated with HLA-DQ1 and a DQ beta RFLP associated with HLA-DQ2 predict that the alpha beta heterodimer in HLA-DQ1/DQ2 heteroxygotes is most closely related to anti-Ro/SSA autoantibodies, thereby supporting a gene complementation mechanism. Beyond this effect, an RFLP associated with HLA-DQ2 and/or DR7 is also related to Ro/SSA precipitins. Multiple molecular histocompatibility mechanisms are implicated, therefore, in the production of anti-Ro/SSA autoantibodies in autoimmune disease. For anti-Ro/SSA autoantibodies in SLE, and perhaps more generally, these data show that the histocompatibility antigens are among the elements that confer autoimmune response specificity and restrict the production of particular autoantibodies among patients with systemic lupus erythematosus.

Autoantibodies↗
PubMed1989

Genomic organization and polymorphisms of the human C3d/Epstein-Barr virus receptor.

The human C3d/Epstein-Barr virus receptor (CR2/CD21) is a 145-kDa protein primarily expressed on mature B lymphocytes. CR2 is a member of the regulators of complement activation (RCA) gene family found on band q32 of chromosome 1. The RCA proteins are characterized by the presence of 60-70 amino acid short consensus repeats (SCR). A full length CR2 cDNA was cloned and used to identify overlapping cosmid genomic clones. Analysis of CR2 exon-intron junctions revealed the presence of three types of exons in the short consensus repeat region of CR2. First, four exons each of which encodes two SCR are present. Five exons encode a single SCR. Six exons encode SCRs which are split in identical positions. The order of these types of exons is in a repeated array of four SCRs, indicating that the contemporary CR2 gene likely evolved from a more primitive gene containing four SCRs. The CR2 full length cDNA clone was used to find restriction fragment length polymorphisms (RFLPs). Restriction enzyme TaqI generated 2.55- and 2.10-kilobase (kb) polymorphic bands. This RFLP was mapped near the exon containing the first two SCRs. HaeIII digestion generated polymorphic bands of 1.45, 1.55, and 1.75 kb. The HaeIII 1.45-kb RFLP band maps near the exon containing the 15th SCR. The TaqI and HaeIII RFLPs will provide tools for the genetic analysis of CR2. The organization of the CR2 gene provides insights into the evolution of human CR2 and the RCA gene family.

Amino Acid Sequence↗
PubMed1988

Two pseudogenes among three rat immunoglobulin lambda chain genes.

In order to examine the number and organization of the immunoglobulin lambda light chain genes of the rat, we have used mouse lambda chain cDNA probes to isolate hybridizing fragments from a partial EcoRI rat liver DNA library. We have determined the partial nucleotide sequence of two such clones. One clone, containing a 5.8 kb EcoRI insert which hybridizes to both mouse C lambda 1 and C lambda 2 probes, includes an apparently expressible C lambda 2-like gene as well as a C lambda 1-like pseudogene (psi C lambda 1.1), arranged similarly to the mouse C lambda gene complexes. Sequence analysis of a second cloned EcoRI fragment (1.15 kb in length) revealed part of a second C lambda 1-like pseudogene (psi C lambda 1.2), the coding regions of both pseudogenes being interrupted by multiple frame-shifting size differences. In the case of psi C lambda 1.2, the degree of sequence identity with mouse C lambda 1 drops abruptly immediately following the termination codon, suggesting that translocation events have played a role in its generation. These two rat pseudogenes, and the mouse C lambda 4 pseudogene, clearly have been rendered unexpressible by separate evolutionary events. Comparisons between C lambda coding and non-coding regions of rats and mice indicate that some of the unusual patterns of divergence we have observed in recently diverged Ck genes may exist in C lambda genes as well.

Animals↗
PubMed1987

Duplication of J kappa genes within genus Rattus.

We have isolated a region containing the immunoglobulin kappa chain joining segments from a liver DNA library of the Australian rat Rattus villosissimus, and determined its nucleotide sequence. While the laboratory rat (Rattus norvegicus) had previously been shown to contain three recently duplicated copies of J kappa 2, R. villosissimus has only two. Furthermore, all three copies of J kappa 2 in R. norvegicus share an 11 bp deletion in their 5' flanking regions which is not evident in either copy of J kappa 2 in R. villosissimus. This suggests that the initial duplication events occurred separately in the two lineages, and were followed by a second duplication in R. norvegicus, all three duplications having occurred within the last 6-12 million years (although more complicated schemes involving gene conversion events cannot be excluded). These results indicate that there is a high degree of plasticity in this region of the genome, and that selective forces must exist which have maintained the number of expressible J kappa segments in humans (5) and rodents (4-6) within their narrow range.

Animals↗
PubMed1987

The structure and evolution of immunoglobulin kappa chain constant region genes in the genus Rattus.

We have cloned and sequenced the C-kappa (Ck) genes from seven species and subspecies of rats which have diverged over the past few million years in Australia. Comparisons of these sequences with each other and the Ck genes of the laboratory rat, Rattus norvegicus, indicate noncoding regions have accumulated fewer mutations than adjacent coding sequences, and amino acid replacing nucleotide substitutions in the coding regions have accumulated at a rate at least as great as silent changes. Exactly the opposite of both of these findings is observed when comparisons are made between Ck or other genes from more distantly related species, indicating that these features may be characteristic of Ck short-term evolutionary gene divergence. Changes in the coding regions of these genes result in a non-random distribution of amino acid substitutions on the three-dimensional alpha-carbon backbone of the Ck domain in the most serologically distinct forms of Ck. While phylogenetic relationships inferred from the Ck nucleotide sequences are in general agreement with those derived from other data, considerable differences are seen in rates of accumulation of Ck gene nucleotide substitutions vs rates of accumulation of enzyme polymorphisms.

Alleles↗
PubMed1984

Kappa-chain constant-region gene sequences in genus Rattus: coding regions are diverging more rapidly than noncoding regions.

We have determined the nucleotide sequence of a 1,200-base pair (bp) genomic fragment that includes the kappa-chain constant-region gene (C kappa) from two species of native Australian rodents, Rattus leucopus cooktownensis and Rattus colletti. Comparison of these sequences with each other and with other rodent C kappa genes shows three surprising features. First, the coding regions are diverging at a rate severalfold higher than that of the nearby noncoding regions. Second, replacement changes within the coding region are accumulating at a rate at least as great as that of silent changes. Third, most of the amino acid replacements are localized in one region of the C kappa domain--namely, the carboxy-terminal "bends" in the alpha-carbon backbone. These three features have previously been described from comparisons of the two allelic forms of C kappa genes in R. norvegicus. These data imply the existence of considerable evolutionary constraints on the noncoding regions (based on as yet undetermined functions) or powerful positive selection to diversify a portion of the constant-region domain (whose physiological significance is not known). These surprising features of C kappa evolution appear to be characteristic only of closely related C kappa genes, since comparison of rodent with human sequences shows the expected greater conservation of coding regions, as well as a predominance of silent nucleotide substitutions within the coding regions.

Animals↗