Rheumatoid factors from patients with rheumatoid arthritis react with beta 2-microglobulin.
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Amino Acid Sequence↗
Biomedical subjects
R C Williams
Publications and source records attributed to R C Williams.
Serologic cross-reactivities poorly reflect allelic relationships in the HLA-B12 and HLA-B21 groups. Dominant epitopes of the alpha 2 helix.
Previous analysis has emphasized the correlation between primary structures of class I HLA molecules and their patterns of serologic cross-reactivity. Here we describe the structures of two serologic groups of HLA-B alleles for which this is not the case. HLA-B45, an allele associated with black populations, is serologically paired with B44 in the B12 group; its structure, however, is divergent from that of B44 but closely related to B50. The BN21 (B*4005) allele is associated with native Americans and is serologically grouped with B50 in the B21 group; its structure, however, is more closely related to alleles of the B40 group. The B44 and B45 serologically cross-reactive molecules differ at seven functional positions of the Ag recognition site; the B50 and BN21 molecules differ at four such residues. These differences are predicted to alter peptide presentation and be capable of eliciting strong alloreactive T cell responses. For these pairs of B12 and B21 Ag, serology appears dominated by epitopes formed by short sequences of the alpha 2 helix which have been shuffled by recombination between alleles. The implications of these results for HLA matching in transplantation are discussed.
Alleles↗
Differential mapping of Fc gamma-binding and monoclonal antibody-reactive epitopes on gE, the Fc gamma-binding glycoprotein of herpes simplex virus type 1.
The entire 396 residue extracellular sequence of gE the HSV-1 Fc gamma-binding glycoprotein has been studied to determine epitopes binding to two mAb II-481 and 88S previously demonstrated to react with gE at or near the Fc gamma-binding regions. Overlapping 7-mers constructed from the established sequence were tested with mAb II-481 and 88S along with their Fab fragments. Control mAb of the same IgG 2b subclass as well as whole rabbit and human IgG and Fc were also tested for binding to overlapping linear sequences using the ELISA pin assay to map Fc gamma-binding regions. Six sequences PKTSWRRVS, GLYTLSV, QVASVVLVVQP, PAPPRSWP, CLYHPQLP, and ASTWTSRL were found that constituted major regions binding to the two different mAb of the same specificity. Glycine substitution for each residue within these sequences indicated that arginine 29, tryptophane 70, valine 144, valine 157, arginine 208, histidine 283, and arginine 305 constituted important portions of the II 481 mAb-reactive epitope. Many of the same regions along with one other, GPLHPSW, appeared to be involved in Fc gamma binding. Substitution of glycine for each residue indicated that histidine 67, tryptophane 70, valine 71, valine 157, valine 158, valine 160, valine 161, tryptophane 210, serine 279, cysteine 280, leucine 281, tyrosine 282, histidine 283, proline 284, glutamine 285, proline 287, tryptophane 302, and arginine 305 were important for Fc gamma-binding. Inhibition by gE peptides of rosetting of E sensitized with rabbit IgG antibody around HSV-1-infected cells, as well as inhibition of rosetting using F(ab)2 fragments of rabbit antibodies to these peptides was used to assay relative contributions of all seven regions to Fc gamma-binding activity. Our results provide a tentative map of mAb binding and Fc gamma-reactive sites on gE. mAb and Fc gamma binding of a limited number of individual antigenic amino acids widely distributed among the separate reactive regions suggest that many of the same separate residues contribute both to antigenicity as well as to Fc gamma-binding activity.
Amino Acid Sequence↗
Heteroclitic polyclonal and monoclonal anti-Gm(a) and anti-Gm(g) human rheumatoid factors react with epitopes induced in Gm(a-), Gm(g-) IgG by interaction with antigen or by nonspecific aggregation. A possible mechanism for the in vivo generation of rheumatoid factors.
Heteroclitic rheumatoid factors (RF) are specific for allotypic determinants, e.g., Gm(a) or Gm(g) on allogeneic, but not autologous IgG. All polyclonal RF we isolated from nine rheumatoid arthritis patients with circulating Gm(a-), (b+), (g-), (f+) IgG displayed dual heteroclitic activity for the Gm(a) and Gm(g) allotypes, as shown by using appropriate RBC agglutination assays and affinity columns bearing Gm(a+) or Gm(g+) IgG. To investigate possible mechanisms underlying the in vivo generation of heteroclitic RF, we tested the ability of nonspecifically and immune-specifically aggregated Gm(a-), (g-) IgG to function as targets for RF from Gm(a-), (g-) patients with rheumatoid arthritis. Heat aggregation (63 degrees C for 20 min) or binding to Ag (as in tetanus toxoid-antitetanus toxoid complexes) induced a "functional" Gm(a+) and/or (g+) phenotype in Gm(a-), (g-) IgG from five healthy subjects and five rheumatoid patients, as suggested by the ability of these altered IgG to function as efficient targets for six heteroclitic RF in direct binding and competitive inhibition experiments. That heterocliticity and dual Gm(a), Gm(g) specificity can be features of a single antibody molecule was formally demonstrated by analysis of a monoclonal RF (IgM mAb 61) generated from a Gm(a-), (g-) rheumatoid patient. RF mAb 61 displayed a high affinity (Kd, 10(-7) M) for IgG Fc fragment of Gm(a+) and (g+) IgG or aggregated autologous Gm(a-), (g-) IgG but did not bind to native autologous IgG. To investigate the molecular basis of the acquired Gm(a) phenotype, PBMC from five Gm(a-) patients with rheumatoid arthritis and two Gm(a-) normal subjects arthritis and two Gm(a-) normal subjects were cultured in vitro after activation with PWM. In most instances, these PBMC produced IgG that behaved as Gm(a+) in sensitive ELISA. Application of the polymerase chain reaction (PCR), using probes specific for the nucleotide sequence coding for the Gm(a) tetrapeptide, to the amplification of DNA from the in vitro-stimulated Gm(a-) normals or rheumatoid patients' PBMC provided no evidence for Gm(a) nucleotide sequences. The present data suggest that acquisition of the Gm(a) determinant by Gm(a-) IgG may result from subtle changes in the CH2-CH3 RF-binding region. Such changes would occur when Gm(a) IgG are complexed with Ag or nonspecifically altered, thereby providing a possible explanation for the induction of heteroclitic RF in Gm(a-) rheumatoid arthritis patients.
Antibodies, Monoclonal↗
Rheumatoid factors from patients with rheumatoid arthritis react with beta 2-microglobulin.
IgM rheumatoid factors (RF) from 18 sera of rheumatoid arthritis (RA) patients isolated from monomeric IgG affinity columns showed strongly positive ELISA reactions with human beta 2-microglobulin (beta 2m), as well as with recombinant beta 2m. When the same RA sera were adsorbed to beta 2m-Sepharose affinity columns, eluted material showed predominant IgM anti-Fc of IgG and anti-beta 2m reactivity. Inhibition reactions with "RF" obtained from IgG affinity columns showed slightly higher reactivity of RF for Fc over beta 2m; however, when RF from the same RA serum had been adsorbed to and eluted from beta 2m affinity columns, beta 2m showed greater inhibition than Fc for RF reacting with either beta 2m or Fc on ELISA plates. Thus two overlapping populations of RF were identified in RA sera showing reactivity with both beta 2m and Fc of IgG. When RF were isolated from IgG columns, affinity was slightly higher for Fc than beta 2m. Conversely, RF eluted from beta 2m Sepharose reacted slightly more with beta 2 m than Fc. Trypsin digests of a polyclonal RA IgM RF showed no beta 2m reactivity in Fc mu 5 fragments. Fab mu RF retained slight anti-Fc IgG but no residual anti-beta 2m activity. Monoclonal human IgM, IgG, or IgA RF either from mixed cryoglobulins or EBV-stimulated RA lymphoid cell lines showed negative or occasional weakly positive anti-beta 2m activity. Overlapping 7-mer peptide ELISA analysis of the entire 99-amino acid sequence of beta 2m showed a major RF-reactive linear hydrophilic sequence at positions 56-60 which included a 3-amino acid exact homology to positions 401, 403, and 404 of the C gamma 3 domain. A peptide encompassing this sequence produced 90% inhibition of RF binding to whole beta 2m. Substitution of neutral glycines for each amino acid throughout the reactive epitope at positions 56-66 indicated that lysine at position 58 aspartic acid at 59, and tryptophane at 60 represented major portions of the RF-reactive epitope. These findings indicate that human RF derived from patients with RA react with other epitopes besides those present on IgG Fc, including epitopes on human beta 2m. For many years serum RF3 found in patients with RA have been regarded as premier examples of autoantibodies to autologous IgG.(ABSTRACT TRUNCATED AT 400 WORDS)
Amino Acid Sequence↗
Unusual HLA-B alleles in two tribes of Brazilian Indians.
The Kaingang and Guarani are culturally and linguistically distinct tribes of southern Brazil. Like all Amerindian groups they show limited HLA polymorphism, which probably reflects the small founder populations that colonized America by overland migration from Asia 11,000-40,000 years ago. We find the nucleotide sequences of HLA-B alleles from the Kaingang and Guarani to be distinct from those characterized in caucasian, oriental and other populations. By comparison, the HLA-A and C alleles are familiar. These results and those reported in the accompanying paper on the Waorani of Ecuador reveal that a marked evolution of HLA-B has occurred since humans first entered South America. New alleles have been formed through recombination between pre-existing alleles, not by point mutation, giving rise to distinctive diversification of HLA-B in different South American Indian tribes.
Alleles↗
Microtubules grow and shorten at intrinsically variable rates.
Microtubules were assembled from pure tubulin with axonemal pieces serving as nuclei. They were observed by video-enhanced differential-interference-contrast light microscopy. Their lengths were measured from videotaped images at frequent intervals (0.13-5 s). Error analysis indicated that the uncertainty in measuring a single length was quite small; the 95% confidence limit approximated the microscope's limit of resolution. Rates of growth and shortening of the dynamically unstable microtubules, obtained from the length-versus-time data, were found to be highly variable. The variability was far too large to be attributed to known random error of measurement and must be a property of the microtubules. Further experiments were aimed at finding its structural cause. The variability of rates exhibited by a single microtubule was as great as that of the whole population. The locations at which a growing microtubule changed its rate of growth were not related to the locations at which rates changed during its subsequent shortening. The cause of the variability must therefore be both small relative to the size of a microtubule and transient relative to its lifetime. Fluctuations in configuration of the microtubule's cap appear to be the likeliest source.
Animals↗
The conversion of tubulin carboxyl groups to amides has a stabilizing effect on microtubules.
In a recent study, we demonstrated that the conversion of carboxyl residues in the C-termini of tubulin to neutral amides with glycine ethyl ester enhanced the ability of the protein to assemble into microtubules and decreased its interaction with microtubule-associated proteins (MAPs). In this work, we investigated the effects of carboxyl modification on the dynamic behavior of microtubules at polymer mass steady state. After steady state, microtubules assembled from unmodified tubulin were sheared, and the mean polymer lengths decreased to 5 microns and then increased to 29 microns within 130 min. In contrast, lengths of sheared microtubules polymerized from tubulin containing 23 modified carboxyl groups increased by only 2-fold. Stabilization of polymer lengths was also observed directly by video-enhanced light microscopy of microtubules grown off of axonemes. Rapid shortening was seen in microtubules composed of unmodified but not modified tubulin. Further evidence for the less dynamic behavior of microtubules as a result of carboxyl modification was obtained from kinetic studies of the elongation phase during assembly which showed a 3-fold lower off-rate constant, k-, for modified microtubules. Another effect of the modification was a 12-fold reduction in the steady-state rate constant for GTP hydrolysis (165 s-1 for unmodified and 14 s-1 for modified). These results suggest that reduction of the negative charges in the C-termini by modification of the acidic residues stabilizes microtubules against depolymerization. MAPs may stabilize microtubules in an analogous manner.
Amides↗
A CNS specific proteoglycan associated with astrocytes in rat optic nerve.
A novel CNS specific keratan sulfate proteoglycan is identified by a monoclonal antibody (mAb) TED15. The antibody recognizes a carbohydrate epitope that is removed from the core protein following keratanase digestion. The relative molecular weight of the glycoprotein is not altered following digestion with heparinase, heparitinase or chondroitinase ABC. By immunoblot analysis, the TED15 antigen is differentially distributed in diencephalic derivatives, being present in the optic nerve but absent from the retina. Within the optic nerve, the TED15 antigen is associated with astrocytes, while in the retina no labeling of astrocytes of Müller cells is observed.
Animals↗
End-stabilized microtubules observed in vitro: stability, subunit, interchange, and breakage.
We report a reliable method to prepare, in vitro, microtubules that are stabilized at both ends by axonemal structures, and report studies of their properties. Such "end-stabilized" microtubules neither grow nor shorten over times of several hours when tubulin subunits are present in the surrounding solution. When subunits are removed, the microtubules eventually break. Breakage occurs within a sinuous and flexible region, a few microns in length, that begins at a single point on the microtubule and grows. When breakage does occur, the resulting two free ends shorten very rapidly until the flexible part has depolymerized and the region of straight microtubule is reached. The remainder of the microtubule then shortens at rates comparable to those ordinarily observed in dynamic instability. Formation of the flexible region can be reversed if subunits are added to the buffer prior to breakage. End-stabilized microtubules are a useful tool for studying interactions of molecules with the microtubular wall. They may be a good model for interpreting stabilizing events that happen in the cell. A preliminary study of the effects of microtubule poisons on the wall is presented.
Alkaloids↗
Two subtypes of HLA-B51 differing by substitution at position 171 of the alpha 2 helix.
Newly defined antigens of the B5, B35 cross-reacting group have been found in Japanese and North American Indians. Nucleotide sequencing of the alleles encoding the Japanese B5.35 antigen and the variant B5 antigen from the Piman Indians show them to be identical. This new allele, B*5102, differs from B*5101 by a single nucleotide substitution that changes residue 171 from histidine to tyrosine. Residue 171, which is part of the alpha 2 helix, is believed to contribute directly to peptide interaction in the A pocket of the binding groove and is either histidine or tyrosine in all HLA-A, B, C heavy chains. Tyrosine 171 is shared by B*5102, B*3501, B*3502, and B*5301 and must be responsible for the serological cross-reactivities of these molecules not shared with B*5101. Stimulation of lymphocytes from a B*5101 positive donor with B*5102 positive cells failed to generate cytotoxic T cells with specificity for the difference between these molecules. However, one out of five clones of cytotoxic T cells raised against B*5101 failed to lyse targets expressing B*5102. Substitution of histidine for tyrosine at residue 171 affected recognition of HLA-B35-restricted human minor histocompatibility antigen-specific T cell clones.
Amino Acid Sequence↗
Human anti-F(ab')2 antibodies show preferential reactivity for F(ab')2 molecules bearing lambda light chains.
In order to evaluate binding specificities of anti-F(ab')2 antibodies from patients with systemic lupus erythematosus (SLE) and from normal healthy controls, F(ab')2 fragments were prepared from 24 IgG myelomas with defined isoelectric points, DNA-associated idiotypes, and kappa/lambda light chain types. Using ELISA and hemagglutination assays, anti-F(ab')2 antibodies from 12 healthy controls and 29 SLE patients were observed to exhibit preferential binding (lambda > kappa) to myeloma F(ab')2 fragments composed of lambda light chains (P < 0.0001). No correlation of anti-F(ab')2 binding and presence of cationic, neutral, or anionic isoelectric points or for DNA-associated idiotypes on monoclonal F(ab')2 was detected. Anti-F(ab')2 antibodies, often elevated in SLE during remission, show preferential specificity for F(ab')2 fragments bearing lambda light chains.
Adult↗
HLA class II variation in the Gila River Indian Community of Arizona: alleles, haplotypes, and a high frequency epitope at the HLA-DR locus.
A genetic distribution for the HLA class II loci is described for 349 "full-blooded" Pima and Tohono O'odham Indians (Pimans) in the Gila River Indian Community. A high frequency epitope in the *DRw52 family was defined by reactions with 31 alloantisera, which we have designated *DR3X6. It segregates as a codominant allele at HLA-DR with alleles *DR2, *DR4, and *DRw8, and has the highest frequency yet reported for an HLA-DR specificity, 0.735. It forms a common haplotype with *DRw52 and *DQw3 that is a valuable marker for genetic admixture and anthropological studies. Phenotype and allele frequencies, and haplotype frequencies for two and three loci, are presented. Variation at these loci is highly restricted, the mean heterozygosity for HLA-DR and HLA-DQ being 0.361. The Pimans represent a contemporary model for the Paleo-Indians who first entered North America 20,000 to 40,000 years ago.
Alleles↗
HLA class I variation controlled for genetic admixture in the Gila River Indian Community of Arizona: a model for the Paleo-Indians.
The genetic distribution of the HLA class I loci is presented for 619 "full blooded" Pima and Tohono O'odham Native Americans (Pimans) in the Gila River Indian Community. Variation in the Pimans is highly restricted. There are only three polymorphic alleles at the HLA-A locus, *A2, *A24, and *A31, and only 10 alleles with a frequency greater than 0.01 at HLA-B where *Bw48 (0.187), *B35 (0.173), and the new epitope *BN21 (0.143) have the highest frequencies. Two and three locus disequilibria values and haplotype frequencies are presented. Ten three-locus haplotypes account for more than 50% of the class I variation, with *A24 *BN21 *Cw3 (0.085) having the highest frequency. Gm allotypes demonstrate that little admixture from non-Indian populations has entered the Community since the 17th century when Europeans first came to this area. As a consequence many alleles commonly found in Europeans and European Americans are efficient markers for Caucasian admixture, while the "private" Indian alleles, *BN21 and *Bw48, can be used to measure Native American admixture in Caucasian populations. It is suggested that this distribution in "full blooded" Pimans approximates that of the Paleo-Indian migrants who first entered the Americas between 20,000 and 40,000 years ago.
Alleles↗
Radiographic evaluation of crestal bone levels adjacent to nonsubmerged titanium implants.
Several parameters have been described to determine success or failure in long-term evaluations of dental implants. One of these parameters is the observation of changes in peri-implant bone levels. Studies on submerged implants have analyzed the bone level changes in the pre- and post-loading phases. No such data exist for intentionally nonsubmerged implants. The purposes of this study were: (1) to test the applicability and reproducibility of a simple computer-assisted method in the evaluation of changes in peri-implant bone levels; (2) to establish a baseline for the longterm radiographic follow-up; and (3) to evaluate changes in crestal bone levels adjacent to nonsubmerged ITI implants between the 1-year and 2-year examination. Standardized periapical radiographs were obtained from 80 implants at the 1-year and 2-year follow-up examinations after their placement. The implants were located in different jaw areas of 55 patients and supported single crowns or short-span fixed partial dentures. For each implant, the distance from implant shoulder to first crestal bone contact (DIB) was measured at the proximal surfaces with a digitizer/computer assembly. Statistically significant greater mean DIB were found at the 1-year (baseline) evaluation for: (1) maxillary sites overall (4.10 x 1.02 mm (SD)) compared with mandibular sites overall (3.61 +/- 1.03 mm); (2) maxillary anterior sites (4.08 +/- 0.97 mm) compared with mandibular posterior sites (3.60 +/- 1.05 mm); and (3) maxillary posterior sites (4.13 +/- 1.12 mm) compared with mandibular posterior sites. No statistically significant changes in DIB occurred in any of the jaw locations between the 1-year and 2-year evaluations.(ABSTRACT TRUNCATED AT 250 WORDS)
Alveolar Bone Loss↗
Native American mitochondrial DNA analysis indicates that the Amerind and the Nadene populations were founded by two independent migrations.
Mitochondrial DNAs (mtDNAs) from 167 American Indians including 87 Amerind-speakers (Amerinds) and 80 Nadene-speakers (Nadene) were surveyed for sequence variation by detailed restriction analysis. All Native American mtDNAs clustered into one of four distinct lineages, defined by the restriction site variants: HincII site loss at np 13,259, AluI site loss at np 5,176, 9-base pair (9-bp) COII-tRNA(Lys) intergenic deletion and HaeIII site gain at np 663. The HincII np 13,259 and AluI np 5,176 lineages were observed exclusively in Amerinds and were shared by all such tribal groups analyzed, thus demonstrating that North, Central and South American Amerinds originated from a common ancestral genetic stock. The 9-bp deletion and HaeIII np 663 lineages were found in both the Amerinds and Nadene but the Nadene HaeIII np 663 lineage had a unique sublineage defined by an RsaI site loss at np 16,329. The amount of sequence variation accumulated in the Amerind HincII np 13,259 and AluI np 5,176 lineages and that in the Amerind portion of the HaeIII np 663 lineage all gave divergence times in the order of 20,000 years before present. The divergence time for the Nadene portion of the HaeIII np 663 lineage was about 6,000-10,000 years. Hence, the ancestral Nadene migrated from Asia independently and considerably more recently than the progenitors of the Amerinds. The divergence times of both the Amerind and Nadene branches of the COII-tRNA(Lys) deletion lineage were intermediate between the Amerind and Nadene specific lineages, raising the possibility of a third source of mtDNA in American Indians.
Asian People↗
Studies of antibody to herpes simplex virus Fc gamma-binding protein gE in patients with rheumatoid arthritis, juvenile rheumatoid arthritis and normal controls.
IgG antibody to gE, the Fc gamma-binding herpes simplex 1 (HSV-1) viral glycoprotein, was studied in 49 rheumatoid arthritis (RA) patients and 43 normal controls. Antibody to gD, another important HSV-1 antigen, was assayed in parallel. No difference between RA patients and normal controls was found in levels of anti-gE antibody measured by reactivity of IgG F(ab')2 fragments reacting with gE coated to ELISA plates. No difference in anti-gD antibody was recorded between normals and patients with RA. Levels of IgG anti-IgE antibody did not correlate with quantitative elevations of serum rheumatoid factor (RF) in RA patients. When IgG anti-gE and anti-gD were assayed in 20 patients with juvenile rheumatoid arthritis and 22 children controls, no significant differences were noted. However, when individual RFs from patients with RA were tested for reactivity against a panel of affinity-isolated F(ab')2 antibodies to gE, some evidence for individual autospecificity was obtained. Four of 20 monoclonal IgM RFs produced from RA patients' B cells showed marked elevations of reactivity with some RA patients' F(ab')2 antibodies to gE. All four of the monoclonal RFs showing this specificity were derived from RA synovial tissue B cells. These findings may provide support for the concept that some RFs in patients with RA show individual specificity for internal image determinants of IgG antibodies to viral Fc gamma-binding proteins.
Adult↗
The molecular basis for reactivity of anti-Cw1 and anti-Cw3 alloantisera with HLA-B46 haplotypes.
HLA haplotypes containing the HLA-B46 allele react with both anti-Cw1 and anti-Cw3 alloantisera, a pattern of reactivity defined as the Cw11 antigen and postulated to involve either a distinctive Cw11 allele or a duplicated HLA-C locus. From serological characterization of CIR cells transfected with B46 cDNA we now demonstrate that the anti-Cw3 reactivity with these haplotypes is solely due to the B46 molecule and not to an HLA-C molecule. Furthermore, isolation and characterization of HLA-C mRNA from cells expressing B46 strongly suggest that anti-Cw1 reactions are directed against the product of a conventional Cw1 allele. The antigenic cross-reactivities of B46 with B62 and Cw3 correlate with its chimaeric primary structure, which is identical to that of B62, except in the alpha 1 helix where it is identical to both Cw3 and Cw1. The structure, distribution and genetic linkage of B46 indicate it is of recent, Asian origin and is the result of a gene conversion, involving Cw1 as the donor gene and B62 as the recipient. These results demonstrate that the Cw11 antigen neither corresponds to a novel HLA-C allele nor a duplicated HLA-C locus, but to a combination of epitopes contributed by linked Cw1 and B46 alleles. The nucleotide sequence we previously and erroneously attributed to a distinct Cw11 allele is now demonstrated to encode Cw8. Isolation of the cDNA clone with this sequence from a library made from a cell homozygous for the B46 haplotype was probably an artefact of contamination.
Alleles↗