Biomedical subjects

James McCluskey

Publications and source records attributed to James McCluskey.

45 records · Page 3Linked to original sources

PubMed2003

Atopy, respiratory function and HLA-DR in Aboriginal Australians.

The Class II genes of the MHC represent a major locus with quantified effects on atopic (allergic) phenotypes in many studies of westernized Caucasians. Although asthma is considered a disease of western societies, typical components of the asthma phenotype, such as elevations of the IgE, are seen with parasitic infestation. We have therefore investigated the effects of the HLA-DRB1 locus on asthma and its intermediate phenotypes in Aboriginal people from the Kimberly region of Australia who were suffering from endemic hookworm infection. Recognizable correlates of allergic asthma were present in the subjects, including skin test positivity to house dust mite (HDM), specific IgE responses to HDM, and the total serum IgE. HLA-DRB1 alleles did not predict the presence of asthma, but multi-allelic tests of association showed the locus accounted for approximately 33% of the variance of the total serum IgE concentration and 17% of the variance of the specific IgE titres to HDM. Genetic admixture was excluded as a cause of the results. These effects of the MHC on IgE levels were an order of magnitude greater than that seen in Caucasians, consistent with the hypothesis that the genetic predisposition to allergic disease may be driven by adaptation to helminth infection. The results further suggest that parasitism per se is not protective against asthma.

Adolescent↗

PubMed2003

A structural basis for the selection of dominant alphabeta T cell receptors in antiviral immunity.

We have examined the basis for immunodominant or "public" TCR usage in an antiviral CTL response. Residues encoded by each of the highly selected genetic elements of an immunodominant clonotype recognizing Epstein-Barr virus were critical to the antigen specificity of the receptor. Upon recognizing antigen, the immunodominant TCR undergoes extensive conformational changes in the complementarity determining regions (CDRs), including the disruption of the canonical structures of the germline-encoded CDR1alpha and CDR2alpha loops to produce an enhanced fit with the HLA-peptide complex. TCR ligation induces conformational changes in the TCRalpha constant domain thought to form part of the docking site for CD3epsilon. These findings indicate that TCR immunodominance is associated with structural properties conferring receptor specificity and suggest a novel structural link between TCR ligation and intracellular signaling.

Amino Acid Sequence↗

PubMed2002

The production, purification and crystallization of a soluble heterodimeric form of a highly selected T-cell receptor in its unliganded and liganded state.

T-cell antigen receptors (TcRs) are heterodimeric cell-surface receptors that play a pivotal role in the cellular immune response. The TcR interacts specifically with a peptide-laden major histocompatability complex (pMHC). A human TcR has been characterized that interacts with an immunodominant epitope, FLRGRAYGL, from the Epstein-Barr virus, a ubiquitous human pathogen, in complex with HLA-B8. Despite the vast TcR repertoire, this TcR is found in up to 10% of the total T-cell population in seropositive HLA-B8+ individuals. In this report, this highly selected TcR is characterized by expressing in Escherichia coli, refolding, purifying and crystallizing the receptor. In addition, the HLA-B8-FLRGRAYGL complex has been expressed in E. coli, refolded and shown to be functionally active. Using native gel electrophoresis, the refolded TcR is shown to be capable of binding specifically to the refolded HLA-B8-FLRGRAYGL and this TcR has been crystallized in complex with the pMHC. The crystals of the unliganded and liganded TcR diffract to 1.5 and 2.5 A, respectively.

Amino Acid Sequence↗

PubMed2002

The structure of HLA-B8 complexed to an immunodominant viral determinant: peptide-induced conformational changes and a mode of MHC class I dimerization.

EBV is a ubiquitous human pathogen that chronically infects up to 90% of the population. Persistent viral infection is characterized by latency and periods of viral replication that are kept in check by a strong antiviral CTL response. Despite the size of the EBV genome, CTL immunity focuses on only a few viral determinants but expands a large primary and memory response toward these epitopes. In unrelated HLA-B8(+) individuals, the response to the immunodominant latent Ag FLRGRAYGL from Epstein Barr nuclear Ag 3A is largely comprised of CTL clones with identical conserved alphabeta TCR structures. To better understand the structural correlates of Ag immunodominance and TCR selection bias, we have solved the crystal structure of the HLA-B8-FLRGRAYGL peptide complex to a resolution of 1.9 A. The structure confirms the importance of P3-Arg, P5-Arg, and P9-Leu as dominant anchor residues involved in peptide binding to HLA-B8. A bulged conformation of the bound peptide provides a structural basis for the critical role of the P7-Tyr residue in T cell recognition. The peptide also induces backbone and side-chain conformational changes in HLA-B8 that are transmitted along the peptide-binding groove in a domino effect. The HLA-B8-FLRGRAYGL complex crystallizes as a dimer in the asymmetric unit and is oriented such that both peptide ligands are projected in the same plane suggesting a higher order arrangement of MHC-peptide complexes that could be involved in formation of the class I Ag-loading complex or in T cell activation.

Antigens, Viral↗

PubMed2002

Identification of a dominant self-ligand bound to three HLA B44 alleles and the preliminary crystallographic analysis of recombinant forms of each complex.

A naturally processed and presented ligand that is shared by human leukocyte antigen (HLA) B*4402, B*4403 and B*4405 molecules has been identified in peptides isolated from immunoaffinity purified HLA B44 complexes. This peptide derived from HLA DPalpha residues 46-54, an endogenous product of HLA DP expressed in the cell line Hmy2.C1R, is a prominent peptide in the mass spectra of species isolated as bound peptides from each allele when the three HLA B44 subtypes were introduced as transfected gene products. Recombinant truncated forms of HLA B*4405(1-276), HLA B*4403(1-276), HLA B*4402(1-276) and beta(2)-microglobulin have been prepared as inclusion bodies in Escherichia coli and refolded in the presence of the DPalpha(46-54) peptide and purified by a combination of size exclusion and anion exchange chromatography. This material was determined to be correctly folded based on detection of a conformational epitope recognized by the W6/32 monoclonal antibody. Large, plate-like crystals of the three complexes were produced using polyethylene glycol as the precipitant. All the crystals belong to the space group P2(1)2(1)2(1) with unit cell dimensions of approximately a=51, b=82, c=110 A. The crystals of three B44/DPalpha complexes diffracted to a resolution of 1.9 A or better. For the first time, using this natural, high abundance ligand of the HLA B44 molecules we have successfully expressed and refolded the three HLA B44 molecules and produced crystals amenable to structural studies.

Alleles↗

PubMed2002

A 320-kilobase artificial chromosome encoding the human HLA DR3-DQ2 MHC haplotype confers HLA restriction in transgenic mice.

MHC class II haplotypes control the specificity of Th immune responses and susceptibility to many autoimmune diseases. Understanding the role of HLA class II haplotypes in immunity is hampered by the lack of animal models expressing these genes as authentic cis-haplotypes. In this study we describe transgenic expression of the autoimmune prone HLA DR3-DQ2 haplotype from a yeast artificial chromosome (YAC) containing an intact similar320-kb region from HLA DRA to DQB2. In YAC-transgenic mice HLA DR and DQ gene products were expressed on B cells, macrophages, and dendritic cells, but not on T cells indicating cell-specific regulation. Positive selection of the CD4 compartment by human class II molecules was 67% efficient in YAC-homozygous mice lacking endogenous class II molecules (Abeta(null/null)) and expressing only murine CD4. A broad range of TCR Vbeta families was used in the peripheral T cell repertoire, which was also purged of Vbeta5-, Vbeta11-, and Vbeta12-bearing T cells by endogenous mouse mammary tumor virus-encoded superantigens. Expression of the HLA DR3-DQ2 haplotype on the Abeta(null/null) background was associated with normal CD8-dependent clearance of virus from influenza-infected mice and development of CD4-dependent protection from otherwise lethal infection with Salmonella typhimurium. HLA DR- and DQ-restricted T cell responses were also elicited following immunization with known T cell determinants presented by these molecules. These findings demonstrate the potential for human MHC class II haplotypes to function efficiently in transgenic mice and should provide valuable tools for developing humanized models of MHC-associated autoimmune diseases.

Amino Acid Sequence↗

PubMed2002

The 1.5 A crystal structure of a highly selected antiviral T cell receptor provides evidence for a structural basis of immunodominance.

Despite a potential repertoire of >10(15) alphabeta T cell receptors (TcR), the HLA B8-restricted cytolytic T cell response to a latent antigen of Epstein-Barr virus (EBV) is strikingly limited in the TcR sequences that are selected. Even in unrelated individuals this response is dominated by a single highly restricted TcR clonotype that selects identical combinations of hypervariable Valpha, Vbeta, D, J, and N region genes. We have determined the 1.5 A crystal structure of this "public" TcR, revealing that five of the six hypervariable loops adopt novel conformations providing a unique combining site that contains a deep pocket predicted to overlay the HLA B8-peptide complex. The findings suggest a structural basis for the immunodominance of this clonotype in the immune response to EBV.

Amino Acid Sequence↗

PubMed2002

Optimization of the MHC class I peptide cargo is dependent on tapasin.

The loading of MHC class I molecules with their peptide cargo is undertaken by a multimolecular peptide loading complex within the endoplasmic reticulum. We show that MHC class I molecules can optimize their peptide repertoire over time and that this process is dependent on tapasin. Optimization of the peptide repertoire is both quantitatively and qualitatively improved by tapasin. The extent of optimization is maximal when MHC class I molecules are allowed to load within the fully assembled peptide loading complex. Finally, we identify a single natural polymorphism (116D>Y) in HLA-B*4402 that permits tapasin-independent loading of HLA-B*4405 (116Y). In the presence of tapasin, the tapasin-independent allele B*4405 (116Y) acquires a repertoire of peptides that is less optimal than the tapasin-dependent allele B*4402 (116D).

ATP Binding Cassette Transporter, Subfamily B, Mem↗

PubMedDate not supplied

Immunological aspects of pregnancy.

Alterations to the immune status of the pregnant woman are necessary to allow mothers to tolerate genetically different fetal tissues during pregnancy. These alterations lead to impaired cell-mediated immunity with increased susceptibility to certain infections such as tuberculosis. During pregnancy, the maternal immune system also shows a relative bias toward T helper type 2 immunity. Several inflammatory dermatoses are either unique to pregnancy or altered by the pregnant state. Immunologists are now beginning to understand the various factors that contribute to the maternal immune tolerance and, in particular, the role of classic (human leukocyte antigen [HLA]-A, HLA-B, HLA-C, and HLA-D) and non-classic (HLA-E, HLA-F, and HLA-G) major histocompatability antigens in this process. Human leukocyte antigen-G, in particular, seems to be important in protecting HLA mismatched tissue from the innate immune system, and investigation of HLA-G expression may help to explain how pregnancy affects inflammatory skin disease. Immunologists are now beginning to understand the alterations to the immune status of the pregnant woman that are necessary to allow mothers to tolerate genetically different fetal tissues during pregnancy. These alterations may help to explain how pregnancy effects inflammatory skin disease.

Female↗