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J M Ahearn

Publications and source records attributed to J M Ahearn.

33 records · Page 2Linked to original sources
PubMed1991

Mapping of the C3b-binding site of CR1 and construction of a (CR1)2-F(ab')2 chimeric complement inhibitor.

CR1/CR2 chimeric receptors in which various short consensus repeats (SCRs) of CR1 were attached to CR2 were transiently expressed on COS cells, and assessed for the binding of polymerized C3b (pC3b) and anti-CR2 by immunofluorescence. Of COS cells expressing chimeras containing SCR 1-4, 1-3, 2-4, 1-2, and 2-3 of the long homologous repeats (LHRs) -B or -C, 96%, 66%, 23%, 0%, and 0%, respectively, bound pC3b. K562 cells were stably transfected with wild-type CR1, deletion mutants of CR1, and the CR1/CR2 chimeras, respectively, and assayed for binding of 125I-pC3b. The dissociation constants (Kd) for pC3b of wild-type CR1 and the LHR-BD and -CD constructs were in the range of 1.0-2.7 nM, and of the CR1/CR2 chimeras containing SCRs 1-4, 1-3, and 2-4 of LHR-B or -C were 1.8-2.4, 6-9, and 22-36 nM, respectively. The factor I-cofactor function of the CR1/CR2 chimeras paralleled the C3b-binding function of the constructs. A CR1/immunoglobulin (Ig) chimeric protein was prepared by fusing SCRs 1-4 of LHR-B to the heavy chains of a murine F(ab')2 anti-nitrophenacetyl (NP) monoclonal antibody. The (CR1)2-F(ab')2 chimera, which retained its specificity for NP, was as effective as soluble, full-length CR1 in binding pC3b, serving as a cofactor for factor I-mediated cleavage of C3b, and inhibiting activation of the alternative pathway, indicating that the bivalent expression of these SCRs reconstitutes the alternative pathway inhibitory function of CR1. The feasibility of creating CR1/Ig chimeras makes possible a new strategy of targeting complement inhibition by the use of Ig fusion partners having particular antigenic specificities.

Animals↗
PubMed1991

Suppression of the immune response by a soluble complement receptor of B lymphocytes.

The CD19-CR2 complex of B lymphocytes contains proteins that participate in two host-defense systems, the immune and complement systems. The ligand for the subunit of the immune system, CD19, is not known, but the complement receptor subunit, CR2 (CD21), binds activation fragments of the C3 component of the complement system and may mediate immunopotentiating effects of complement. A recombinant, soluble CR2 was prepared by fusing the C3-binding region of the receptor to immunoglobulin G1 (IgG1). The (CR2)2-IgG1 chimera competed with cellular CR2 for C3 binding and suppressed the antibody response to a T cell-dependent antigen when administered to mice at the time of immunization. This inhibitory effect of (CR2)2-IgG1 demonstrates the B cell-activating function of the CD19-CR2 complex and suggests a new method for humoral immunosuppression.

Amino Acid Sequence↗
PubMed1991

Interaction of iC3b with recombinant isotypic and chimeric forms of CR2.

CR2 is a component of a signal transduction complex on B lymphocytes that augments B cell responses to Ag. We have quantitatively assessed binding by the two isotypic forms of CR2 for two of its ligands, the polymerized iC3b (p(iC3b)) fragment of C3, and gp350/220, the EBV membrane protein. The recombinant 15-SCR or 16-SCR forms of CR2 bound p(iC3b) with identical affinities. Full binding activity of CR2 for p(iC3b) was observed with a chimera comprised of SCR-1 and -2 of CR2 fused to SCR-17 through -30 of CR1. Therefore, the alternatively spliced SCR-10a has no role in binding p(iC3b), and the binding activity of wild type receptor for iC3b can be reconstituted with SCR-1 and -2 of CR2. The binding affinities of the two isoforms of CR2 for soluble gp350/220 were also similar. Additional sites in the C3c region of C3 have been postulated also to interact with CR2. However, monomeric iC3b and C3d were equally effective in inhibiting the binding of p(iC3b) to CR2, indicating that the C3c region of iC3b does not contribute to the interaction of iC3b with CR2. Finally, the relative abilities of C3b and iC3b to bind to CR1 and CR2 were compared. The conversion of C3b to iC3b generated a ligand with an approximate 100-fold decrease in affinity for CR1 and a 10-fold increased affinity for CR2, resulting in a 1000-fold greater likelihood for binding to the latter receptor that may then promote B cell activation.

Animals↗
PubMed1991

Molecular interactions of complement receptors on B lymphocytes: a CR1/CR2 complex distinct from the CR2/CD19 complex.

The complement system augments the humoral immune response to low concentrations of antigen. This effect may be partly mediated by complement receptors on the surface of B lymphocytes that bind immunogenic complexes bearing fragments of C3 and C4. We have shown by immunoprecipitation analysis that the two complement receptors expressed by B lymphocytes, complement receptor 1 (CR1) and CR2, form a detergent-sensitive complex on the surface of tonsillar B lymphocytes and on K562 erythroleukemia cells that were co-transfected with cDNAs encoding CR1 and CR2. The CR1/CR2 complex is distinct from the CR2/CD19 complex and may assist B cell activation by efficiently capturing C3b-containing immunogens and maintaining such immunogens on the B cell after CR1 and factor I-mediated cleavage to iC3b and C3dg. The complement activating immunogen may then trigger signal transduction by the CR1/CR2 complex, the CR2/CD19 complex, or membrane immunoglobulin.

Antigens, CD19↗
PubMed1989

Mapping of the Epstein-Barr virus and C3dg binding sites to a common domain on complement receptor type 2.

Complement receptor type 2 (CR2;CD21), a member of the superfamily of proteins containing short consensus repeats (SCRs), is the B cell receptor for both the gp350/220 envelope protein of Epstein-Barr virus (EBV), and for the C3dg protein of complement. By analysis of CR2 deletion mutants and chimeras formed with CR1 (CD35) we determined that of the 15 SCRs in CR2, the NH2-terminal two SCRs are necessary and sufficient to bind both gp350/220 and C3dg with affinities equivalent to those of the wild-type receptor. The epitope for OKB-7, a mAb that blocks binding of both EBV and C3dg and shares with these ligands B cell-activating capabilities, also requires both SCR-1 and SCR-2, whereas mAbs lacking these functions bind to other SCRs. Thus, EBV, a polyclonal activator of B cells, has selected a site that is proximate or identical to the natural ligand binding site in CR2, perhaps reflecting the relative immutability of that site as well as its signal transducing function.

Animals↗
PubMed1989

Characterization of the class I HLA 9.2-kb PVU II restriction fragment length polymorphism. Linkage to HLA-A and lack of disease association.

The strongest reported association between a class I HLA allele and disease is that of HLA-B27 with ankylosing spondylitis (AS). However, it has not been shown whether B27 is the gene that predisposes to the development of AS or if it is merely linked with the disease-susceptibility locus. Furthermore, if B27 itself is the disease-susceptibility gene, there may be epistatic loci that also contribute to the development of AS or modify its clinical manifestation. A class I HLA 9.2-kb Pvu II restriction fragment was recently identified, which, when present in a B27-positive individual, further increased the relative risk for developing AS (from 119 to 297). This study was therefore designed to confirm the association between AS and this restriction fragment length polymorphism (RFLP) and to map the location of this fragment in the genome. The data presented here suggest that the class I HLA 9.2-kb Pvu II RFLP represents a Pvu II polymorphism at the 5' end of the HLA-A locus that is tightly linked with both HLA-A3 and A9 alleles. However, there is no association between this RFLP and AS in a population of patients living in Baltimore.

Arthritis, Reactive↗
PubMed1988

Epstein-Barr virus (EBV) infection of murine L cells expressing recombinant human EBV/C3d receptor.

The normal host range of Epstein-Barr virus (EBV) is limited to primate B lymphocytes and certain epithelial cells that express the C3d/EBV receptor [complement receptor 2 (CR2, CD21)]. In the present study, expansion of the tissue tropism of EBV has been accomplished by stably transfecting the murine fibroblast L cell line with pMT.CR2. neo.1, a eukaryotic expression vector promoting the transcription of a complementary DNA insert encoding human CR2. High CR2-expressing transfected L cells were selected by fluorescence-activated cell sorting. The recombinant CR2 was shown to have the same molecular weight as wild-type CR2 from Raji cells and to mediate the binding by the transfectants of particles bearing the iC3b and C3d fragments of the third component of complement. All CR2-expressing L cells, but not nontransfected controls, also bound EBV, as assessed by indirect immunofluorescence. After a 60-hr culture, approximately 0.5% of the CR2-expressing cells preincubated with EBV demonstrated immunofluorescent staining of EBV nuclear antigen with serum from a patient with nasopharyngeal carcinoma. No fluorescent staining of cells was seen with monoclonal antibodies to the early antigen complex or to gp350/220, indicating that the infection was predominantly latent. Infected cells cultured for up to 4 weeks remained EBV nuclear antigen-positive. The capacity of recombinant human CR2 to confer on murine L cells susceptibility to stable latent infection by EBV indicates that this receptor is a primary determinant of the tissue tropism of EBV and may facilitate studies of cell-specific factors that regulate the viral growth cycle.

Animals↗
PubMed1988

Epidemiology and genetics of ankylosing spondylitis.

The spondyloarthropathies represent a group of disorders characterized by the presence of inflammatory peripheral and axial arthritis with sacroiliac involvement and a tendency to familial aggregation; prototype clinical syndromes are ankylosing spondylitis (AS) and Reiter's syndrome. The descriptive epidemiology of AS, including course and prognosis in patients cohorts, is reviewed. Particular attention is focused on the genetic epidemiology of AS, especially the evidence supporting causal association of disease with the class I histocompatibility antigen HLA-B27. In addition, analytical studies that support associations of infectious agents are reviewed. The spondyloarthropathies, especially AS and Reiter's syndrome, clearly illustrate the epidemiologic concept of host-environment interaction in the development and expression of disease.

Disease Susceptibility↗
PubMed1987

Cloning and sequence analysis of the mouse genomic locus encoding the largest subunit of RNA polymerase II.

The genomic locus (RPII215) encoding the largest subunit of mouse RNA polymerase II has been cloned by low stringency hybridization to a Drosophila RPII215 probe. The mouse gene consists of 28 exons which span 30 kilobases. Analysis of the nucleotide and predicted protein sequences indicates that the protein is comprised of two domains. There is a 1500 residue amino-terminal domain which contains seven regions strikingly similar to those in the beta' subunit of Escherichia coli RNA polymerase, and a carboxyl-terminal domain comprised of 52 repeats of a 7-amino-acid consensus sequence Tyr-Ser-Pro-Thr-Ser-Pro-Ser. Among the seven highly conserved regions are a strongly basic domain consistent with a DNA-binding site and a consensus sequence characteristic of a potential zinc-binding domain. The 5' upstream region contains three tandem sequences similar to binding sites for the transcription factor SP1. Two of the introns in this gene splice at donor GC dinucleotides as opposed to previously described invariant GT sites. The identification of regions which are highly conserved as compared with bacterial and yeast RNA polymerase and other regions which are unique to the mouse protein suggests which domains of RNA polymerase large subunits are involved in aspects of transcription common to both procaryotes and eucaryotes and which are characteristic of transcription in higher organisms.

Amino Acid Sequence↗
PubMed1985

A unique structure at the carboxyl terminus of the largest subunit of eukaryotic RNA polymerase II.

Purified eukaryotic nuclear RNA polymerase II consists of three subspecies that differ in the apparent molecular masses of their largest subunit, designated IIo, IIa, and IIb for polymerase species IIO, IIA, and IIB, respectively. Subunits IIo, IIa, and IIb are the products of a single gene. We present here the amino acid composition of calf thymus subunits IIa and IIb and the C-terminal amino acid sequence of subunit IIa (IIo) inferred from the nucleotide sequence of part of the mouse gene encoding this RNA polymerase subunit. The calculated amino acid composition of the peptide unique to subunit IIa indicates that subunit IIa contains a domain rich in serine, proline, threonine, and tyrosine. The sequence at the 3' end of the mouse RNA polymerase II largest subunit gene reveals that the C-terminal domain consists of 52 repeats of a seven amino acid block with the consensus sequence Tyr-Ser-Pro-Thr-Ser-Pro-Ser. This sequence is also unusual in that it contains a high percentage of potential phosphorylation sites.

Amino Acid Sequence↗
PubMed1985

Systemic lupus erythematosus: a review of clinico-laboratory features and immunogenetic markers in 150 patients with emphasis on demographic subsets.

Clinical and laboratory features as well as immunogenetic markers were analyzed in 150 patients with SLE to determine if demographic factors--age at diagnosis, sex and race--influenced the expression of disease. The overall series included 103 white females, 35 black females, 10 white males and 2 black males; the mean age at diagnosis was 32.5 years. Males had a significantly older mean age at diagnosis than females (40.4 versus 31.8 years) and a significantly higher frequency of peripheral neuropathy (50% versus 18.8%). No other differences in clinical or laboratory features or HLA-DR or DQ phenotype frequencies were noted. Blacks had a significant younger mean age at diagnosis than whites (26.9 versus 33.4 years) as well as significantly higher frequencies of nephritis, hypertension, acute lupus pneumonitis, discoid rash, hyperglobulinemia and hypocomplementemia. There were no differences in autoantibody frequencies between race-specific subgroups. HLA-DR2, DRw52 and DQ1 were significantly associated with SLE in whites compared to controls; no HLA-DR or DQ associations were found with SLE in blacks. In whites, HLA-DR2 was associated with the presence of anti-Ro(SS-A) antibody while HLA-DR3 was associated with the presence of both anti-Ro(SS-A) and anti-La(SS-B) antibody. In blacks, HLA-DR2 was associated with the presence of anti-nDNA antibody. In whites, patients with late-onset SLE (age at diagnosis greater than or equal to 50 years) had significantly lower frequencies of nephritis and mesenteric vasculitis but, on the other hand, a higher frequency of secondary Sjögren syndrome than patients with age at diagnosis less than or equal to 22 years. Similar findings were noted when blacks aged 35 and above were compared to those aged 17 and below at diagnosis. In whites, the frequency of both anti-Ro(SS-A) and La(SS-B) antibodies increased with increasing age as did that of HLA-DR3; HLA-DR2, however, was more frequent in those with younger age at diagnosis. These data suggest the existence of two serologic-genetic subsets of SLE with different age at diagnosis.

Adolescent↗