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Biomedical subjects

D Givol

Publications and source records attributed to D Givol.

At least 127 records · Page 7Linked to original sources
PubMed1981

Skeletal muscle actin mRNA. Characterization of the 3' untranslated region.

Plasmids p749, p106, and p150 contain cDNA inserts complementary to rat skeletal muscle actin mRNA. Nucleotide sequence analysis indicates the following sequence relationships: p749 specifies codons 171 to 360; p150 specifies codons 357 to 374 together with 120 nucleotides of the 3'-non-translated region; p106 specifies the last actin amino acid codon, the termination codon and the entire 3' non-translated region. Plasmid p749 hybridized with RNA extracted from rat skeletal muscle, cardiac muscle, smooth (stomach) muscle, and from brain. It also hybridizes well with RNA extracted from skeletal muscle and brain of dog and chick. Plasmid p106 hybridized specifically with rat striated muscles (skeletal and cardiac muscle) mRNA but not with mRNA from rat stomach and from rat brain. It also hybridized to RNA extracted from skeletal muscle of rabbit and dog but not from chick. Thermal stability of the hybrids and sensitivity to S1 digestion also indicated substantial divergence between the 3' untranslated end of rat and dog skeletal muscle actins. The investigation shows that the coding regions of actin genes are highly conserved, whereas the 3' non-coding regions diverged considerably during evolution. Probes constructed from the 3' non-coding regions of actin mRNAs can be used to identify the various actin mRNA and actin genes.

Actins↗
PubMed1981

Polymorphism of germ-line immunoglobulin VH genes correlates with allotype and idiotype markers.

The polymorphic nature of the immunoglobulin VH genes was investigated by Southern blot analysis of liver DNA of sixteen different mouse strains and hybridization with VH probes. Differences in restriction enzyme pattern (REP) were observed and six different patterns of restriction fragments were found for the sixteen strains analyzed. No equivalent polymorphism was observed in another multigene family, the actins. The six patterns correlate with immunoglobin constant region allotypes (Igh-1). Experiments with Igh-1-congenic strains suggest that the VH REP is linked to immunoglobulin constant region haplotype. Mouse strains which share inherited idiotypes also share identical VH restriction pattern. This provides a structural basis for the genetic linkage between idiotypes and allotypes. It also indicates that different strains carry different VH gene repertoires, which may be the basis for the expression of different inherited idiotypes in various strains. We propose that a VH group in a set of linked genes that are coinherited as a cluster with the constant region genes and that VH and Ch can be regarded as an extended haplotype.

Animals↗
PubMed1980

Cloning and sequence of the cDNA corresponding to the variable region of immunoglobulin heavy chain MPC11.

Poly(A)-containing mRNA from mouse myeloma MPC11 was transcribed into cDNA which was cloned in the PstI site of the plasmid pBR322. The transformants were screened by hybridization with a cDNA fragment, derived from plasmid p gamma(11)7, corresponding to the 5' portion of the constant region of MPC11 heavy chain. Several positive transformants were found to contain various lengths of the variable region of the heavy chain. We describe the structure and sequence of one of these clones, pV(11)2, which contains cDNA corresponding to the entire variable region of MPC11 heavy chain and extends to codon 248 in the constant region. The protein sequence deduced from the DNA sequence indicates that the variable region of MPC11 heavy chain contains 121 amino acids and belongs to subgroup II of mouse heavy chains. Comparison of this sequence with other heavy chain sequences suggests a J (joining) segment of 16 residues which overlaps five residues of the third hypervariable region. The cDNA sequence shows that there is no discontinuity between the end of the variable region and the beginning of the constant region.

Animals↗
PubMed1980

T-cell hybridoma bearing heavy chain variable region determinants producing (T,G)-A--L-specific helper factor.

Thymus-derived lymphocytes (T cells) exert their regulatory effect (help or suppression) on the antibody production by B cells either by direct cell to cell interaction or by soluble mediators or factors. The low frequency of specific T cells, the heterogeneity of their responses and their relatively short life span have hampered the molecular characterization of the antigen recognition unit of T cells, and its structure is largely unknown. The lymphocyte hybridization technique, which has been found very useful for the production of B-cell hybridomas secreting specific monoclonal antibodies, has also been used for the generation of homogeneous and stable T-cell hybridomas with unlimited growth potential. So far the only specific effector function demonstrated in the established T hybridomas is the property to generate a factor(s) which suppresses antibody responses. We now describe the establishment of hybrid lines which exhibit characteristic T-cell markers. One of the hybridomas (denoted R-9) releases into the culture supernatant factor(s) with helper activity specific to the synthetic polypeptide (T,G)-A--L and bears surface determinants of the immunoglobulin heavy chain variable region (VH). Such hybrid cell lines are of great value for studies on the nature of the T-cell receptor.

Animals↗
PubMed1980

Structure of immunoglobulin gamma 2b heavy chain gene cloned from mouse embryo gene library.

A mouse DNA clone containing the constant part of the immunoglobulin gamma 2b heavy chain was isolated from a mouse gene library. The library was constructed in Charon 4A from a partial EcoRI digest of mouse embryo DNA and was screened with a plasmid (p gamma (11)7) containing a cDNA insert of the heavy chain constant region of the plasmacytoma MPC-11 (1). The Charon 4A clone contains a 14 kb insert which is cleaved by EcoRI into a 6.8 kb and 7.2 kb fragments, of which only the 6.8 kb contains the sequence for gamma 2b heavy chain. Restriction analysis and partial sequence of the insert in p gamma (11) 7 enabled us to obtain three fragments corresponding to the 5' (amino acid 161-302) middle (amino acid 302-443) and 3' (mostly non coding 107 bp) regions of the constant region. Restriction analysis of the Charon 4A clone and hybridisation to these nick translated fragments revealed that the gamma 2b constant region gene contains about 1.5 kb and has three intervening sequences.

Animals↗
PubMed1980

Correlated expression of VH framework and VH idiotypic determinants on T helper cells and on functionally undefined T cells binding group A streptococcal carbohydrate.

Antibodies to framework determinants of the VH and V lambda fragments of MOPC 315 and antisera to the VH idiotype determinants of the A 5 A antibody were used to analyze the antigen receptors of mouse T (and B) cells. This was done by using the antibodies as inhibitors in (a) an assay in which the binding of radiolabeled streptococcal carbohydrate (A-CHO) antigen by primed and unprimed T and B cells is determined and (b) an assay in which the helper activity of group A streptococcal vaccine-primed T cells is determined. The results suggest that the major proportion of primed and unprimed T cells binding A-CHO (70-90%) exhibit VH framework and VH idiotypic determinants. This population appears to include the helper T cells. A minor proportion of T cells (10-30%) express V lambda-related framework determinants and lack VH framework and VH idiotypic determinants. This population does not include T helper cells. Taken together, the data suggest that a subpopulation of T cells, including the helper cells, uses entire Ig VH regions as part of their antigen receptor system.

Animals↗
PubMed1980

Antibodies to immunoglobulin heavy chain variable regions protect helper cells from specific suicide by radiolabeled antigen.

The antigen-binding receptor of helper T cells was studied by radioactive antigen-caused suicide in vitro. Purified antibodies to immunoglobulin variable regions, obtained from sera of rabbits immunized with isolated VH and VL fragments of mouse myeloma proteins (MOPC 315, XRPC 25), were used to inhibit the binding of radiotoxic antigen. Anti-VH, but not anti-V lambda or anti-V chi inhibited suicide of carrier-primed cells.

Antibodies, Anti-Idiotypic↗
PubMed1980

Antigen-binding receptors on T cells from long-term MLR. evidence of binding sites for allogeneic and self-MHC products.

Antibody inhibition of radiolabelled stimulator membrane vesicle binding by T blasts activated in the mixed lymphocyte reaction (MLR) was used to identify responder-cell determinants involved in the binding phenomenon. Antisera or monoclonal antibodies against Thy-1, Lyt-1, Lyt-2 and Ly-6 antigens were not inhibitory. However, antibodies against heavy-chain V region (VH) determinants strongly inhibited vesicle binding by both primary and long-term MLR blasts. Anti-Ia (both alloantisera and monoclonal reagents) caused inhibition of antigen binding by primary MLR blasts only. T blasts from long-term MLR lines were neither Ia-positive, nor susceptible to blocking of antigen binding with anti-Ia. However, these cells were capable of specifically absorbing soluble syngeneic Ia material, with the concomitant appearance of vesicle-binding inhibition with anti-Ia sera. Acquisition of syngeneic Ia by T blasts was effectivelly blocked with the anti-VH reagent. Passively bound self-Ia did not interfere with vesicle binding in the absence of anti-Ia. These results strongly suggest the existance of specific self-Ia acceptor sites closely linked to the receptors for stimulator alloantigens on T cells proliferating in MLR. A receptor model based on these findings is briefly discussed.

Animals↗
PubMed1979

Anti-V region framework antibodies affect the ligand binding of VL dimer.

The effect of antibodies to the light chain variable region (VL) of protein MOPC-315 (alpha, lambda 2), on the binding of hapten by VL315 dimer or Fv315 (VL + VH) was studied by equilibrium dialysis. Anti-VL did not change the binding properties of Fv but affected the binding properties of VL dimer. At pH 5, the binding properties of VL in the presence or absence of anti-VL were the same, whereas at pH 8, anti-VL reduced the number of ligands bound to VL from two to one. It has previously been shown that VL dimer binds one ligand at pH 5 and two ligands at pH 8, and that VL conformation at pH 5 is tighter. Hence, our results suggest that anti-VL tightens the conformation of VL dimer at pH 8.0 such that it can bind only one ligend. Since Fv is not affected by anti-VL, the results indicate that a combining site made of two identical chains (VL dimer) can undergo a conformational change upon interaction with its antibody. Such conformational change can indirectly affect the binding properties.

Animals↗
PubMed1978

The role of nitro groups in the binding of nitroaromatics to protein MOPC 315.

Two series of dinitrophenyl haptens, in which chlorine replaces one or both nitro groups, were used to investigate, by a combination of high-resolution 1H n.m.r. and fluorescence quenching, the presence of groups in the combining site of protein MOPC 315, which form hydrogen bonds to the aromatic-ring substituents of the hapten. The large differences in binding constants on successive replacement of nitro groups were shown to be due to specific hapten-substituent-protein interactions by (a) showing that there was little difference in the interaction between these haptens and 3-methylindole (a model for the residue tryptophan-93L with which the hapten stacks in protein MOPC 315), (b) proving by 1H n.m.r. that the mode of hapten binding is constant and (c) showing that the differences in Kd were consistent with the relative hydrogen-bonding capacities of chlorine and the nitro moiety. In this way it was established that each nitro group forms a hydrogen bond. Furthermore, from consideration of the 1H n.m.r. chemical shifts of several dinitrophenyl haptens and their trinitrophenyl analogues, it was shown that there is no distortion of the o-nitro group on binding to the variable fragment of protein MOPC 315.

Antigen-Antibody Reactions↗
PubMed1978

Sequence of chicken ovalbumin mRNA.

The complete sequence of chicken ovalbumin mRNA is presented; it is 1,859 residues long, excluding its terminal 'cap' and poly(A). The region coding for ovalbumin lies close to the 'cap' but is separated from the poly(A) by an extensive 3' noncoding region of 637 nucleotides which may have no function that is precisely dependent on its sequence.

Amino Acid Sequence↗