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J Thibodeau

Publications and source records attributed to J Thibodeau.

36 records · Page 2Linked to original sources
PubMed1994

Subsets of HLA-DR1 molecules defined by SEB and TSST-1 binding.

Superantigens bind to major histocompatibility complex class II molecules on antigen-presenting cells and stimulate T cells. Staphylococcus aureus enterotoxin B (SEB) and toxic shock syndrome toxin-1 (TSST-1) bind to the same region of human lymphocyte antigen (HLA)-DR1 but do not compete with each other, which indicates that they bind to different subsets of DR1 molecules. Here, a mutation in the peptide-binding groove disrupted the SEB and TSST-1 binding sites, which suggests that peptides can influence the interaction with bacterial toxins. In support of this, the expression of the DR1 molecule in various cell types differentially affected the binding of these toxins.

Animals↗
PubMed1994

T cell receptor-major histocompatibility complex class II interaction is required for the T cell response to bacterial superantigens.

Bacterial and retroviral superantigens (SAGs) stimulate a high proportion of T cells expressing specific variable regions of the T cell receptor (TCR) beta chain. Although most alleles and isotypes bind SAGs, polymorphisms of major histocompatibility complex (MHC) class II molecules affect their presentation to T cells. This observation has raised the possibility that a TCR-MHC class II interaction can occur during this recognition process. To address the importance of such interactions during SAG presentation, we have used a panel of murine T cell hybridomas that respond to the bacterial SAG Staphylococcal enterotoxin B (SEB) and to the retroviral SAG Mtv-7 when presented by antigen-presenting cells (APCs) expressing HLA-DR1. Amino acid substitutions of the putative TCR contact residues 59, 64, 66, 77, and 81 on the DR1 beta chain showed that these amino acids are critical for recognition of the SAG SEB by T cells. TCR-MHC class II interactions are thus required for T cell recognition of SAG. Moreover, Mtv-7 SAG recognition by the same T cell hybridomas was not affected by these mutations, suggesting that the topology of the TCR-MHC class II-SAG trimolecular complex could be different from one TCR to another and from one SAG to another.

Animals↗
PubMed1994

Binding sites for bacterial and endogenous retroviral superantigens can be dissociated on major histocompatibility complex class II molecules.

Bacterial and retroviral superantigens (SAGs) interact with major histocompatibility complex (MHC) class II molecules and stimulate T cells upon binding to the V beta portion of the T cell receptor. Whereas both types of molecules exert similar effects on T cells, they have very different primary structures. Amino acids critical for the binding of bacterial toxins to class II molecules have been identified but little is known of the molecular interactions between class II and retroviral SAGs. To determine whether both types of superantigens interact with the same regions of MHC class II molecules, we have generated mutant HLA-DR molecules which have lost the capacity to bind three bacterial toxins (Staphylococcus aureus enterotoxin A [SEA], S. aureus enterotoxin B [SEB], and toxic shock syndrome toxin 1 [TSST-1]). Cells expressing these mutated class II molecules efficiently presented two retroviral SAGs (Mtv-9 and Mtv-7) to T cells while they were unable to present the bacterial SAGs. These results demonstrate that the binding sites for both types of SAGs can be dissociated.

Animals↗
PubMed1993

Molecular and biochemical features of poly (ADP-ribose) metabolism.

In the past five years, poly(ADP-ribosyl)ation has developed greatly with the help of molecular biology and the improvement of biochemical techniques. In this article, we describe the physico-chemical properties of the enzymes responsible for the synthesis and degradation of poly(ADP-ribose), respectively poly(ADP-ribose) polymerase and poly(ADP-ribose) glycohydrolase. We then discuss the possible roles of this polymer in DNA repair and replication as well as in cellular differentiation and transformation. Finally, we put forward various hypotheses in order to better define the function of this polymer found only in eucaryotes.

Amino Acid Sequence↗
PubMed1993

Expression in Escherichia coli of the 36 kDa domain of poly(ADP-ribose) polymerase and investigation of its DNA binding properties.

We have expressed in Escherichia coli the 36 kDa domain of the human poly(ADP-ribose) polymerase. This polypeptide comprises the C-terminal part of the DNA binding domain, as well as the automodification region of the enzyme, but lacks the zinc-finger motifs of the N-terminal region and the C-terminal catalytic domain. By probing the crude E. coli protein extracts with radioactive DNA probes (South-Western blots), we have shown that the 36 kDa domain binds a DNA probe of 222 bp but does not bind a shorter probe of 66 bp. This interaction is stronger when the polypeptide is fused to the 55 kDa catalytic domain of the enzyme.

Amino Acid Sequence↗
PubMed1993

Superantigens of microbial origin.

Food poisoning associated staphylococcal enterotoxins and other bacterial products of diverse origin are now the focus of immunological research. These substances have special properties which determine their designation as superantigens. They influence T cell functions by controlling their repertoire, their cytokines production and their modulation of the immune response. As a consequence, superantigens might be at the origin of bacterial and autoimmune diseases. In this review we describe mainly the staphylococcal enterotoxins as representative members of the superantigen family.

Animals↗
PubMed1993

Interactions between staphylococcal superantigens and MHC class II molecules.

Superantigen binding to MHC class II molecules is a prerequisite for T cell stimulation. While the presentation of superantigens is not MHC restricted, allelic and isotypic variations in the ability of class II molecules to interact and present these superantigens exist. SEs bind to MHC class II molecules outside of the peptide binding groove, differences in binding affinities of the toxins for class II alleles and isotypes have been shown. In addition, a direct interaction between T cell receptor and MHC class II molecules could be responsible for these differences. In this review we provide a molecular analysis of the interactions of SEs with class II molecules.

Animals↗
PubMed1992

Selection of mouse cells with amplified metallothionein genes retaining their glucocorticoid inducibility.

Two new mouse cell mutants, resistant to either 80 or 100 mM CdCl2, were isolated to study the regulation of transcription by the glucocorticoid hormones. Their metallothionein mt-1% and mt-2+ genes were amplified coordinately to a maximum of 30 copies per cell. By Southern blot analysis, no gross rearrangement was detectable near the mt+ loci. Contrary to other mutants previously isolated, the metallothionein-specific mRNAs of these mutants are inducible by dexamethasone.

Animals↗
PubMed1992

Structural analysis of the putative regulatory region of the rat gene encoding poly(ADP-ribose) polymerase.

A lambda EMBL3 clone containing the first three exons along with part of the 4th exon of the rat poly(ADP-ribose) polymerase gene was isolated from a genomic DNA library. This clone also contains 6.6 kbp of upstream sequences. Nucleotide sequence analysis of the proximal 5' 670 nucleotides flanking the major RNA start site of the rat gene does not reveal significant global homology with the same region of the human gene, but a series of short sequences are identical. Among these sequences are found two putative Sp1 binding sites along with a decanucleotide sequence responsible for the attachment of the transcription factor AP-2.

Amino Acid Sequence↗
PubMed1991

Identification of the staphylococcal enterotoxin A superantigen binding site in the beta 1 domain of the human histocompatibility antigen HLA-DR.

The staphylococcal enterotoxin A (SEA) is a superantigen that must bind to class II molecules of the major histocompatibility complex to be recognized by T cells. In humans, most HLA-DR class II allelic and isotypic forms, such as DR1, bind SEA well. DRw53 is an exception, binding SEA very poorly. We have localized this difference to a single residue (amino acid 81) in the beta 1 domain. A highly conserved histidine at residue 81 allows SEA binding, but a tyrosine does not. Residue 81 is predicted to lie in an alpha-helix on the surface of the molecule, with its side chain pointing up out of the pocket associated with binding of conventional peptide antigens. This finding supports the hypothesis that superantigens and conventional antigens bind to different sites on the class II molecule.

Amino Acid Sequence↗
PubMed1990

Expression in E. coli of the catalytic domain of rat poly(ADP-ribose)polymerase.

A 2 kilobase pair cDNA coding for the entire C-terminal catalytic domain of rat poly(ADP-ribose)polymerase has been expressed in E. coli. The overproduced 55 kDa polypeptide is active in synthesizing poly(ADP-ribose) and the 4 kDa N-terminal region of this domain is recognized by the monoclonal antibody C I,2 directed against the calf enzyme. Also, the minor alpha-chymotrypsin cleavage site found in the human catalytic domain is not present in the rat enzyme as revealed by the absence of the 40 kDa specific degradation product in the E. coli cells expressing the rat domain. The expression of this partial rat cDNA should thus permit the rapid purification and subsequent crystallization of the catalytic domain of the enzyme.

Amino Acid Sequence↗
PubMed1990

New mouse somatic cell mutants resistant to cadmium affected in the expression of their metallothionein genes.

Fluctuation tests à la Luria and Delbruck were performed with mouse LMTK cells, and the results indicate that the appearance of variants resistant to cadmium is due to random spontaneous mutations and not to epigenetic events. The rate of spontaneous mutations leading to cadmium resistance was calculated to be 0.92 x 10(-6) per cell per generation. This rate increased 14-fold on treatment with ethyl methane sulfonate. Several stable mutant cell lines resistant to cadmium were selected and characterized with respect to metallothionein (MT) induction. Based on the copy number of mt+ genes and the levels of MT proteins and mRNA, the mutants could be divided into two classes, A and B. Although group A mutants have the same number of mt1+ and mt2+ genes as wild-type cells, upon induction with cadmium, the amount of MT proteins and mRNA in the mutants are greatly increased over wild-type levels. This observation strongly suggests a mutation that regulates MT gene transcription in these cells. In group B mutants, the mt+ genes are amplified about three- to fourfold, and their MT protein and mRNA basal levels are, as expected, much higher than in the wild-type cells, under uninduced and induced conditions.

Animals↗
PubMed1989

Cloning of rodent cDNA coding the poly(ADP-ribose) polymerase catalytic domain and analysis of mRNA levels during the cell cycle.

We have isolated a partial 2.0 kb cDNA (pRATC) encoding the entire 489 amino acids of the NAD binding domain located at the C terminus of the rat poly(ADP-ribose) polymerase. pRATC sequences were analysed and compared with the human mRNA. Our analysis reveals a remarkable homology between the rat and human nucleotide and amino acid sequences. Although a few minor amino acid changes were detected, we have found that the total number of possible phosphorylation sites remained constant in the NAD binding domain of both enzymes. We have also found that a 102 amino acid sequence, containing the putative nucleotide binding site Gly-Lys-Gly (position 378), is perfectly conserved between the rat and human sequences. Strong homology was also detected between pRATC and genomic DNA isolated from various vertebrates. In addition, we have analysed the levels of poly(ADP-ribose) polymerase mRNA throughout the cell cycle. Our results show that the levels of mRNA culminate in the G1 phase. We have also found that the increase in enzymatic activity observed in rats following treatment with phenobarbital did not correspond to an increase in the mRNA levels.

Amino Acid Sequence↗
PubMed1987

Cloning and characterization of the metallothionein-I gene from mouse LMTK cells.

A clone of about 14 kb containing the metallothionein MT-I gene and three repetitive sequences, was isolated from a genomic library of mouse LMTK DNA. The MT-I gene was functional. Transfected cells became cadmium resistant. Two of the three repetitive sequences were moderately repetitive while the other was closely related to the R family.

Animals↗