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Results for “Immunoglobulin Heavy Chains”

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At least 19 recordsLinked to original sources
PubMed1979

Nuclear transcripts of mouse heavy chain immunoglobulin genes contain only the expressed class of C-region sequences.

In plasmacytoma cells producing IgG, IgA, or IgM immunoglobulin heavy chains, the large precursors of the heavy chain messenger RNA's contain nucleotide sequences that specify only the expressed class of constant region. This indicates that the switch from one class of heavy chain to another during B cell ontogeny does not occur by altered processing of a complex gene transcript.

Animals↗
PubMed1977

Murine plasma cells secreting more than one class of immunoglobulin heavy chain. III. Immunoglobulin production by established cultures and cloned lines of SAMM 368.

SAMM 368, a plasmacytoma which produces IgA-kappa and IgG2b-kappa was established in vitro from primary explants or after animal passage. The 9 lines that were established produced both paraproteins. The production of both immunoglobulins by single cells was demonstrated by immunofluorescent staining and cloning. Continuous culture of 3 parent lines for 18 months and 11 cloned lines for periods from 5 to 7 months demonstrated that double production is a stable characteristic of this plasmacytoma. Two single paraprotein-producing varients (IgG2b or IgA) were derived when cells were cultured in the presence of Fungizone. Chromosomal analysis of SAMM 368 indicates that this double producing tumor has a modal number similar to those observed in myelomas producing a single immunoglobulin class.

Cell Line↗
PubMed1977

Initiation and translation in vitro of mRNA for MOPC 315 immunoglobulin heavy chain and characterization of translation product.

An initiation study of mineral oil-induced plasmacytoma (MOPC) 315 heavy chain immunoglobulin (H315) in vitro has been conducted using formyl-[35S]methionyl-tRNAfMet and a highly purified 18 S message from MOPC 315 solid tumor in a crude rabbit reticulocyte lysate system. The product was specifically precipitated by antibodies directed against MOPC 315 immunoglobulin and H315. The in vitro H315 products terminally labeled with formyl-[35S]methionine or internally labeled with [3H]leucine were electrophoretically identical with in vivo H315 on sodium dodecyl sulfate-polyacrylamide gels. All of the [35S]-methionine was incorporated at the NH2 terminus, not internally, since there is a near complete recovery of [35S]methionine following one cycle of Edman degradation. The NH2-terminal cyanogen bromide peptide, CN2, of in vivo and in vitro H315 co-migrated exactly on gel electrophoresis under conditions which completely resolved two proteins differing in size by only 14 amino acids. These data strongly suggest that there is no NH2-terminal precursor of H315 in this system. Cyanogen bromide peptide profiles of in vivo and in vitro H315 were chromatographically indistinguishable. Three peptides, CN1, CN2, and CN4, which represent approximately 85% of the total amino acids of H315 were isolated and further characterized by electrophoresis and paper chromatography. All were very similar to the corresponding peptides of authentic H315. We conclude that the fidelity of H315 translation is preserved in vitro.

Cell Line↗
PubMed1977

Amino acid sequence of the precursor region of MOPC-315 mouse immunoglobulin heavy chain.

Partially purified mRNA coding for the MOPC-315 heavy immunoglobulin alpha chain was translated in a reticulocyte lysate containing 20 labeled amino acids. Radiolabeled MOPC-315 heavy chain precursor protein, purified by preparative gel electrophoresis and immunoprecipitation, was sequenced by Edman degradation. The labeled phenylthiohydantoin amino acid obtained in each cycle was identified and quantitated by high-pressure liquid chromatography. The precursor sequence of 18 amino acids, Met-Lys-Val-Leu-Ser-Leu-Leu-Tyr-Leu-Leu-Thr-Ala-Ile-Pro-His-Ile-Met-Ser, preceded the sequence corresponding to the NH2 terminus of the mature secreted heavy chain.

Amino Acid Sequence↗
PubMed2025

Comparative Analysis of Mammalian Adaptive Immune Loci Revealed Spectacular Divergence and Common Genetic Patterns.

Adaptive immune responses are mediated by the production of adaptive immune receptors, antibodies, and T-cell receptors, which bind antigens, thus causing their neutralization. Unlike other proteins, adaptive immune receptors are not fully encoded in the germline genome and result from a complex of somatic processes collectively called V(D)J recombination affecting germline immunoglobulin (IG) and T-cell receptor (TR) loci consisting of template genes. While various existing studies report extreme diversity of antibodies and T-cell receptors, little is known about the diversity of germline IG and TR loci. To overcome this gap, the first comparative analysis of full-length sequences of IG/TR loci across 46 mammalian species from 13 taxonomic orders was performed. First, germline gene counts were shown to correlate in immunoglobulin heavy chain immunoglobulin heavy chain (IGH)/immunoglobulin lambda (IGL) loci and T-cell receptor alpha (TRA)/T-cell receptor beta (TRB) and anticorrelate in immunoglobulin kappa (IGK)/IGL, possibly indicating coevolution between corresponding chains. Second, structures of IG/TR loci were analyzed, and it was shown that IG/TR loci formed by long arrays of high multiplicity repeats are more common for species that have experienced population bottlenecks. Finally, haplotypes of IG/TR loci with little or no sequence similarity within a species were found, suggesting that they may have a limited potential for homologous recombination. These results demonstrate that IG/TR loci are rapidly evolving genomic regions whose structural variation is shaped by the population history of the species and open new perspectives for immunogenomics studies.

Animals↗
PubMed1977

Murine plasma cells secreting more than one class of immunoglobulin heavy chain. IV. Sequence differences between kappa-chains of SAMM 368 IgG2b and IgA.

The amino terminal sequences of the kappa light chains from SAMM 368 IgG2b and IgA have been determined. The two chains belong to different kappa subgroups as indicated by 10 amino acid differences among the first 23 residues. An additional three residue differences were demonstrated in the sequence including the first complementarity region. These results indicate that single cells of SAMM 368 produce two unique kappa-chains.

Amino Acid Sequence↗
PubMed1975

Partial amino acid sequence of an IgA2 human immunoglobulin heavy chain.

The amino acid sequence of the heavy chain of an IgA2, AIm(1) polymeric myeloma protein (Avil) was studied. Altogether, sequence data were obtained for some 130 residues. Including the amino acids placed by homology with IgA1, this accounts for some 170 residues, thus representing more than one-third of the alpha2 chain. The sequence includes 26 amino acids from the amino-terminal end (V-H III), and 25 residues at the "hinge" region. Of a total of 17 cysteine residues, 14 were located in regions of the molecule which were identical or homologous in the alpha1 and alpha2 chains. These striking homologies, together with the results obtained by diagonal maps of classes of IgA. Study of the cysteine-containing peptides of the J chain are consistent with the conclusion that the J chains associated with different classes of immunoglobulins are identical.

Amino Acid Sequence↗
PubMed1977

An immunoglobulin heavy chain variable region (VH) marker associated with cross-reactive idiotypes in man.

The VH specificities of two rabbit antisera raised against the H chains of two human IgM cold agglutinins were studied with the aid of proteins on which the VH sequence data are available. They were found to recognize a new VH antigen (VHMar), which may represent a subgroup of VHI. This antigen is found on anti-Ii antibodies which have cross-reactive idiotypes. It is moderately well-expressed in normal gamma globulin and strongly expressed on 8% of unselected myeloma and macroglobulinaemia proteins. There is evidence to suggest that a proportion of the cross-reactive idiotypes among the anti-Ii antibodies involve this VH antigen.

Agglutinins↗
PubMed1977

Primary structure of the variable regions of two canine immunoglobulin heavy chains.

The complete amino acid sequences of the variable regions of two canine immunoglobulin heavy chains have been determined by automated Edman degradation and found to be strongly homologous to the human VHIII subgroup. The canine sequences were identical with each other at 76 of 113 residue positions. Twenty-three of the 37 differences are located within the four hypervariable regions previously defined by the sequences of several human VHIII proteins. Forty-five of 77 framework residue positions are invariant in the seven human and two canine VHIII proteins which have been completely sequences. The canine proteins are 78% homologous to the framework of the human prototype. Phylogenetically associated residues before the first hypervariable region were confirmed and several potential phylogenetically associated residues were identified between the first and third hypervariable regions. This study represents the first complete amino acid sequences of VH regions of spontaneously occurring, nonhuman homogeneous immunoglobulins. The date demonstrate a high degree of preservation of VHIII structure in another species.

Amino Acid Sequence↗
PubMed1978

Location of the genes for human heavy chain immunoglobulin to chromosome 6.

Immunoglobulin synthesis was examined in 31 man-mouse hybrid clones produced by fusing RAG mouse cells with human lymphoid cells. Cells were grown in serum-free medium containing [(14)C]leucine and a (14)C-labeled amino acid mixture. Spent medium was dialyzed, concentrated, and subjected to radioimmunoelectrophoresis. Eighteen clones were found to produce material that gave a radiolabeled precipitin line with anti-human IgG (gamma-chain specific). Production of material which was indistinguishable on radioimmunoelectrophoresis from human Ig gamma heavy chain, was dependent on the presence in hybrid clones of human chromosome 6. The material was found to have the ion-exchange elution characteristics of human IgG. When radiolabeled spent medium from human lymphoid lines and from chromosome 6-positive hybrid clones was exposed to protein A-Sepharose and bound material eluted with 8 M urea was subjected to sodium dodecyl sulfate polyacrylamide gel electrophoresis, three radiolabeled peaks occurred with molecular weights of approximately 55,000 (coinciding with that of Ig gamma heavy chain), 40,000 and 25,000 (coinciding with that of Ig light chains). No similar peaks were detected in experiments where spent medium from RAG cells was treated identically. These studies lead us to conclude that certain RAG-human lymphoid hybrid clones produce human IgG and that the structural genes for gamma heavy chains are located on human chromosome 6. These results also imply that the locus coding for human alpha,-antitrypsin (Pi) is located on chromosome 6.

Cell Line↗
PubMed1979

Chromosomal location of the genes for human immunoglobulin heavy chains.

We have studied somatic cell hybrids between P3x63Ag8 mouse myeloma cells deficient in hypoxanthine phosphoribosyltransferase (EC 2.4.2.8) and either human peripheral lymphocytes or human lymphoblastoid or myeloma cells for the production of human immunoglobulin chains and for the expression of enzyme markers assigned to each of the different human chromosomes. Human chromosome 14 was the only human chromosome present in all independent hybrids producing mu, gamma, and alpha human heavy chains. In two of the independent hybrids that produced human heavy chains, human chromosome 14 was the only human chromosome present in the hybrid cells. Loss of human chromosome 14 from these hybrids resulted in the concomitant loss of their ability to produce human immunoglobulin heavy chains. In view of these results, we conclude that the genes for human immunoglobulin heavy chains are located on human chromosome 14 in immunoglobulin-producing human cells.

Animals↗
PubMed1978

Isolation, purification, and properties of mouse heavy-chain immunoglobulin mRNAs.

A procedure is described for the isolation of highly purified heavy-chain immunoglobulin mRNAs from a variety of mouse plasmacytomas (IgA, IgG, and IgM producers). The use of fresh tissue and the rapid isolation and direct extraction of membrane-bound polyribosomes were found to be essential in obtaining large quantities of undegraded heavy-chain mRNAs. The individual mRNAs were purified by two cycles of oligo(dT)-cellulose chromatography, sodium dodecyl sulfate--sucrose gradient centrifugation, and electrophoresis on 98% formamide containing polyacrylamide gels. When added to a cell-free protein-synthesizing system from wheat germ, the MPC-11 gamma2b and H2020 alpha heavy-chain mRNAs efficiently directed the synthesis of a predominant product of 55 000 molecular weight, while the synthesis of a 70 000 dalton protein in addition to other lower molecular weight polypeptides were observed with MOPC 3741 mu mRNA. All of these proteins were immunoprecipitable with class-specific heavy-chain antisera, and in the case of the gamma2b in vitro products good correspondence in a comparative trypsin--chymotrypsin fingerpring with in vivo labeled gamma2b heavy chain was observed. The gamma2b and a alpha heavy-chain mRNAs possessed a chain length of approximately 1800 nucleotides and the mu mRNA a size of approximately 2150 nucleotides when examined under stringent denaturation conditions. The purities of the alpha, gamma2b, and mu mRNAs were estimated to be 60--80%, 50--70%, and 50--83%, respectively, on the basis of their hybridization rates with cDNA probes in comparison to mRNA standards of known complexity. Heavy-chain mRNAs of the same class isolated from different mouse strains (Balb/C or NZB) display no detectable sequence differences in cross hybridization experiments, even though the cDNA--mRNA hybrids are submitted to stringent S1 nuclease digestion. These results indicate that allotypic determinants represent only a minor fraction of the heavy-chain constant region sequence in the mouse.

Animals↗
PubMed1977

Purification of immunoglobulin heavy chain messenger RNA by immunoprecipitation from the mouse myeloma tumor, MOPC-31C.

Immunoglobulin heavy chain mRNA was purified from immunoprecipitated polysomes derived from the mouse myeloma tumor, MOPC-31C. The purified mRNA migrated predominantly as a single band upon polyacrylamide gel electrophoresis in 98% formamide and the molecular weight of this mRNA was calculated to be 700,000. This mRNA was as active as the purified light chain mRNA when it was employed as a template in a cell-free protein synthesizing system from wheat germ. The translation product had a molecular weight of 55,000 daltons, and migrated slightly faster than mature heavy chain upon polyacrylamide gel electrophoresis in sodium dodecylsulfate. The protein synthesized by the direction of this mRNA was shown to yield tryptic peptides corresponding to those derived from the mature heavy chain protein except that one missing peptide was replaced by another additional peptide. DNA complementary to the mRNA was synthesized by RNA-dependent DNA polymerase from avian myeloblastosis virus. Hybridization kinetic analysis between the heavy chain mRNA and its complementary DNA indicated that the RNA was essentially homogenous with rabbit globin mRNA as a standard.

Animals↗
PubMed1978

Organization of immunoglobulin heavy chain genes and allelic deletion model.

We have assessed the number of times the gene sequence encoding constant regions of mouse immunoglobulin heavy chains gamma1, gamma2a, and gamma3 are represented in the mouse genome by hybridization kinetic analysis. All three genes are present at one copy each per haploid genome in normal tissues and myelomas producing IgM or IgG3. IgG1-producing myelomas, however, contain 1 copy each of the gamma1 and gamma2a genes and 0.5 copy of the gamma3 gene per haploid genome. IgG2b-producing myelomas contain 1 copy of the gamma2a gene and 0.5 copy each of the gamma1 and gamma3 genes per haploid genome. IgG2a-producing myelomas contain 1 copy of the gamma2a gene and 0.5 copy each of the gamma1 and gamma3 genes per haploid genome. In myelomas producing IgA, all three gamma genes are represented 0.5 times per haploid genome. In order to account for the results we propose an allelic deletion model: (i) The specific deletion of heavy chain constant region genes accompanies the recombination of a variable region gene to a constant region gene. (ii) The portion of the chromosome that resides between two joining sequences is excised out of the chromosome. (iii) The recombination occurs on one of the alleles. Based on this model we also propose that heavy chain genes are arranged on one chromosome in the following order; variable region genes, unknown spacer sequence, mu, gamma3, gamma1, gamma2b, gamma2a, and alpha.

Alleles↗
PubMed1975

Immunoglobulin heavy chain mRNA in mitogen-stimulated B cells.

This paper relates the synthesis of DNA, immunoglobulin and heavy chain (H) mRNA in murine spleen cells following activation of B cells with lipopolysaccharide from E. coli (LPS). Spleen cells (CBA/H mice) were cultivated with 10% FCS and 10 mug LPS/ml. 4 h pulses with [3H]thymidine showed that DNA synthesis was stimulated within the first day following LPS activation and exhibited a sharp peak at 24 h. The shape of the DNA synthesis curve suggests that the cells susceptible to LPS stimulation are activated in a synchronous manner. Stimulation of H-chain mRNA (H-mRNA) synthesis proceeded rapidly (within 6 h of LPS addition) and peaked around 24 h, in parallel to DNA synthesis. The H-mRNA was isolated and quantitated by making use of its interaction with IgG[1, 2]. The actual level of H-mRNA in the culture increased threefold during the first 24 h and then doubled within the next 48 h. Estimates of the actual number of H-mRNA were approximately 200 molecules H-m-RNA/cell on day 0 rising to 1800/cell on day 3. In such a mixed cell population these figures will be accurate only within a factor of 2-3 (at least 35% B cells in spleen cell suspensions at the commencement of the culture, with up to 35-60% of plasma blasts by day 3 and 4 of LPS treatment). Translation of the lymphoid cell mRNA in oocytes from Xenopus laevis demonstrated that stimulation of H-mRNA synthesis was restricted to mu-mRNA, although some gamma-mRNA was present in the original spleen cells. High levels of synthesis of immunoglobulin followed after a lag period of about 24 h following LPS addition peaking after 48 and 72 h; the proportional Ig production relative to total protein synthesis reached 26% on days 3 and 4. Stimulation of Ig production was limited to IgM. Rapid stimulation of mitosis and H-mRNA synthesis thus precedes the maximum synthesis of Ig molecules, suggesting a translational block on H-mRNA during cell maturation. There was no apparent block on the transport of H-mRNA from the nucleus during early stages of activation.

Animals↗