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Biomedical subjects

P Parham

Publications and source records attributed to P Parham.

At least 55 records · Page 3Linked to original sources
PubMed1999

Hepatitis C virus envelope glycoprotein E1 originates in the endoplasmic reticulum and requires cytoplasmic processing for presentation by class I MHC molecules.

We investigated whether hepatitis C virus envelope glycoprotein E1 is transported from the endoplasmic reticulum (ER) to the cytoplasm of infected cells for class I MHC processing. Target cells expressing E1 were killed by CTL lines from a hepatitis C virus-infected chimpanzee, and synthetic peptides were used to define an epitope (amino acids 233-GNASRCWVA-241) presented by the Patr-B*1601 class I MHC molecule. An unusually high concentration (>100 nM) of this nonameric peptide was required for target cell lysis, but this could be reduced at least 1000-fold by replacing the asparagine at amino acid position 234 (Asn234) with aspartic acid (Asp), the anticipated anchor residue for NH2-terminal peptide binding to Patr-B*1601. Conspicuously, position 234 is part of an N-glycosylation motif (Asn-Xaa-Ser/Thr), suggesting that the Asn234 to Asp substitution might occur naturally within the cell due to deglycosylation/deamidation of this amino acid by the cytosolic enzyme peptide N-glycanase. In support of this model, we demonstrate that presentation of the epitope depended on 1) cotranslational synthesis of E1 in the ER, 2) glycosylation of the E1 molecule, and 3) a functional TAP transporter to shuttle peptide from the cytosolic to ER compartment. These results indicate for the first time that during infection of the host, viral envelope glycoproteins originating in the ER are processed in the cytoplasm for class I MHC presentation. That a posttranslational change in amino acid sequence from Asn to Asp alters the repertoire of peptides presented to CD8+ CTL has implications for the design of antiviral vaccines.

Animals↗
PubMed1999

Phylogenetic relationships of the Callitrichinae (Platyrrhini, primates) based on beta2-microglobulin DNA sequences.

The phylogenetic relationships of callitrichine primates have been determined by DNA sequence analyses of exons 1, 2, and 3 of the beta2-microglobulin gene. Parsimony, distance, and maximum likelihood analyses of ca. 900 base pairs of 21 taxa, representing all callitrichine genera, indicated that Saguinus was the most basal offshoot. Within Saguinus, S. fuscicollis appeared as the first divergent lineage followed by an unresolved trichotomy formed by S. mystax/S. imperator, S. midas/S. bicolor, and S. oedipus. A second callitrichine lineage was formed by Leontopithecus; each of the three species studied showed identical nucleotide sequences. Callimico appeared as the sister taxon of Callithrix/Cebuella. Genetic distances within this latter group were very small, although a stronger association between Cebuella and species of the Callithrix argentata group was observed. The inclusion of Cebuella in the genus Callithrix is suggested. These studies indicated that tamarins are more plesiomorphic than marmosets in agreement with the phyletic dwarfism hypothesis.

Animals↗
PubMed1999

Molecular phylogeny of new world primates (Platyrrhini) based on beta2-microglobulin DNA sequences.

Neotropical primates, traditionally grouped in the infraorder Platyrrhini, comprise 16 extant genera. Cladistic analyses based on morphological characteristics and molecular data resulted in topologic arrangements depicting disparate phylogenetic relationships, indicating that the evolution of gross morphological characteristics and molecular traits is not necessarily congruent. Here we present a phylogenetic arrangement for all neotropical primate genera obtained from DNA sequence analyses of the beta2-microglobulin gene. Parsimony, distance, and maximum likelihood analyses favored two families, Atelidae and Cebidae, each containing 8 genera. Atelids were resolved into atelines and pitheciines. The well-supported ateline clade branched into alouattine (Alouatta) and ateline (Ateles, Lagothrix, Brachyteles) clades. In turn, within the Ateline clade, Lagothrix and Brachyteles were well-supported sister groups. The pitheciines branched into well-supported callicebine (Callicebus) and pitheciine (Pithecia, Cacajao, Chiropotes) clades. In turn, within the pitheciine clade, Cacajao and Chiropotes were well-supported sister groups. The cebids branched into callitrichine (Saguinus, Leontopithecus, Callimico, Callithrix-Cebuella), cebine (Cebus, Saimiri), and aotine (Aotus) clades. While the callitrichine clade and the groupings of species and genera within this clade were all well supported, the cebine clade received only modest support, and the position of Aotus could not be clearly established. Cladistic analyses favored the proposition of 15 rather than 16 extant genera by including Cebuella pygmaea in the genus Callithrix as the sister group of the Callithrix argentata species group. These analyses also favored the sister grouping of Callimico with Callithrix and then of Leontopithecus with the Callithrix-Callimico clade.

Animals↗
PubMed1999

Residue 3 of beta2-microglobulin affects binding of class I MHC molecules by the W6/32 antibody.

Previous studies of class I MHC molecules have shown that the owl monkey (Aotus) possesses at least two variants of the beta2-microglobulin (beta2m) protein. These two variants have different isoelectric points, and exhibit differential reactivity with the monoclonal antibody W6/32. We report cDNA sequences of the B2m gene, from W6/32-positive and W6/32-negative Aotus cell lines. The two beta2m variants we identified exhibit a single amino acid difference at position three. An arginine residue at position 3 was correlated with W6/32 reactivity, whereas histidine was associated with non-reactivity. W6/32 reactivity was conferred to a W6/32-negative Aotus cell line when it was transfected with the B2m from the W6/32-positive cell line. Residue 3 of beta2m is located at the surface of the class I molecule. It is also close to position 121 of the MHC class I heavy chain, which has previously been shown to influence W6/32 antibody binding. We conclude that W6/32 binds a compact epitope on the class I molecule that includes both residue 3 of beta2m and residue 121 of the heavy chain. We examined the distribution of the two beta2m motifs in a sample Aotus population using an allele-specific polymerase chain reaction assay. The pattern of beta2m segregation we observed matches that which was defined previously by serology. Additionally, we identified laboratory-born hybrid animals who possess both variants of beta2m.

Alleles↗
PubMed1999

A divergent non-classical class I gene conserved in salmonids.

Complementary DNA for two class I genes of the rainbow trout, Oncorhynchus mykiss, were characterized. MhcOnmy-UBA*01 is similar to Onmy-UAC32 and the classical major histocompatibility complex class I genes of other fish species, whereas Onmy-UAA*01 is divergent from all class I genes so far characterized. Onmy-UAA*01 is expressed at lower levels than Onmy-UBA*01. Although Onmy-UAA*01 exhibits restriction fragment length polymorphism on Southern blotting, the encoded protein is highly conserved. Two allotypes, which differ only by substitution at amino acid position 223 of the alpha 3 domain, have been defined. Onmy-UAA*01 has an exon-intron organization like other class I genes and contains a Tc1-like transposon element in intron III. Orthologues of Onmy-UAA*01 have been characterized in four other species of salmonid. Between four species of Oncorhynchus, UAA*01 proteins differ by only 2-6 amino acids, whereas comparison of Oncorhynchus with Salmo trutta (brown trout) reveals 14-16 amino acid differences. The Onmy-UAA*01 gene has properties indicative of a particularly divergent non-classical class I gene.

Amino Acid Sequence↗
PubMed1999

Evidence for an HLA-C-like locus in the orangutan Pongo pygmaeus.

HLA-B and C are related class I genes which are believed to have arisen by duplication of a common ancestor. Previous study showed the presence of orthologues for both HLA-B and C in African apes but only for HLA-B in Asian apes. These observations suggested that the primate C locus evolved subsequent to the divergence of the Pongidae and Hominidae. From an analysis of orangutan Tengku two HLA-C-like alleles (Popy C*0101 and Popy C*0201) were defined as well as three HLA-B-like (Popy-B) alleles. By contrast, no Popy-C alleles were obtained from orangutan Hati, although three Popy-B alleles were defined. Thus an HLA-C-like locus exists in the orangutan (as well as a duplicated B locus), implying that the primate C locus evolved prior to the divergence of the Pongidae and Hominidae and is at least 12-13 million years old. Uncertain is whether all orangutan MHC haplotypes contain a C locus, as the failure to find C alleles in some individuals could be due to a mispairing of HLA-C-specific primers with certain Popy-C alleles. These results raise the possibilities that other primate species have a C locus and that the regulation of natural killer cells by C allotypes evolved earlier in primate evolution than has been thought.

Alleles↗
PubMed1999

Analysis of a successful immune response against hepatitis C virus.

To investigate the type of immunity responsible for resolution of hepatitis C virus (HCV) infection, we monitored antibody and intrahepatic cytotoxic T lymphocyte (CTL) responses during acute (<20 weeks) infection in chimpanzees. Two animals who terminated infection made strong CTL but poor antibody responses. In both resolvers, CTL targeted at least six viral regions. In contrast, animals developing chronic hepatitis generated weaker acute CTL responses. Extensive analysis of the fine specificity of the CTL in one resolver revealed nine peptide epitopes and restriction by all six MHC class I allotypes. Every specificity shown during acute hepatitis persisted in normal liver tissue more than 1 yr after resolution. These results suggest that CD8+CTL are better correlated with protection against HCV infection than antibodies.

Acute Disease↗
PubMed1999

Evolutionary disruptions of human syntenic groups 3, 12, 14, and 15 in Ateles paniscus chamek (Platyrrhini, primates).

Comparative gene assignments of 18 markers, based on analyses of somatic cell hybrids and previous data in the literature, indicated that human (HSA) syntenic groups 3, 12, 14, and 15 are dissociated in the spider monkey species Ateles paniscus chamek (APC). Markers present in HSA 3p were allocated to APC 3 and APC 9. The HSA 12 cluster was split into two syntenic groups, one mainly including HSA 12p markers in APC 16 and the other, including HSA 12q markers, in APC 2p. The HSA 14q cluster split into three syntenic groups, corresponding to APC 2q, APC 6, and APC 12. Finally, the HSA 15 cluster split into two syntenic groups, APC 2q and APC 3. Comparisons with previous gene assignments and human SROs led to the tentative postulation of rearrangements having occurred during the evolutionary divergence of man and A. paniscus chamek. Chromosome painting data in the congeneric species A. geoffroyi, other New World and Old World primates, and several representative non-primate animals were compared in an attempt to delineate the ancestral and derived conditions underlying the evolutionary rearrangement of syntenic groups in mammals.

Animals↗
PubMed1999

Polymorphism and evolution of HLA class I and II genes and molecules.

Genes in the HLA complex, the human major histocompatibility complex (MHC), encode polymorphic HLA class I and II molecules that help T lymphocytes recognise and respond to foreign antigens. Certain HLA class I allotypes also regulate the response of natural killer cells. HLA class I and II molecules with little or no polymorphism contribute a variety of functions to the immune response, as do class I molecules coded by genes outside of the HLA region. Knowledge of the organisation of HLA class I and II genes, of the nucleotide sequences of their alleles, and the three-dimensional structures of their protein products, has facilitated analysis of the evolution and polymorphism of HLA class I and II genes and molecules. In turn, these analyses have provided insight into the mechanisms and selective forces driving change in the HLA complex.

Genes, MHC Class I↗
PubMed1998

Clathrin self-assembly is regulated by three light-chain residues controlling the formation of critical salt bridges.

Clathrin self-assembly into a polyhedral lattice mediates membrane protein sorting during endocytosis and organelle biogenesis. Lattice formation occurs spontaneously in vitro at low pH and, intracellularly, is triggered by adaptors at physiological pH. To begin to understand the cellular regulation of clathrin polymerization, we analyzed molecular interactions during the spontaneous assembly of recombinant hub fragments of the clathrin heavy chain, which bind clathrin light-chain subunits and mimic the self-assembly of intact clathrin. Reconstitution of hubs using deletion and substitution mutants of the light-chain subunits revealed that the pH dependence of clathrin self-assembly is controlled by only three acidic residues in the clathrin light-chain subunits. Salt inhibition of hub assembly identified two classes of salt bridges which are involved and deletion analysis mapped the clathrin heavy-chain regions participating in their formation. These combined observations indicated that the negatively charged regulatory residues, identified in the light-chain subunits, inhibit the formation of high-affinity salt bridges which would otherwise induce clathrin heavy chains to assemble at physiological pH. In the presence of light chains, clathrin self-assembly depends on salt bridges that form only at low pH, but is exquisitely sensitive to regulation. We propose that cellular clathrin assembly is controlled via the simple biochemical mechanism of reversing the inhibitory effect of the light-chain regulatory sequence, thereby promoting high-affinity salt bridge formation.

Amino Acid Sequence↗
PubMed1998

Direct binding and functional transfer of NK cell inhibitory receptors reveal novel patterns of HLA-C allotype recognition.

Cytotoxicity of human NK cells is under negative control of killer cell Ig-like receptors (KIR) specific for HLA class I. To determine the specificity of five KIR containing two Ig domains (KIR2D), direct binding of soluble recombinant KIR2D to a panel of HLA class I transfectants was assayed. One soluble KIR2D, derived from an inhibitory receptor with a long cytoplasmic tail (KIR2DL1), bound to HLA-C allotypes containing asparagine 77 and lysine 80 in the heavy chain, as expected, since these allotypes inhibit lysis by NK cells expressing KIR2DL1. Surprisingly, another KIR2D (KIR2DL2), which inhibits NK lysis of cells expressing HLA-C molecules with serine 77 and asparagine 80, bound to HLA-C allotypes carrying either amino acid motif. Expression of the KIR2DL receptors in NK cells using recombinant vaccinia viruses confirmed these patterns of recognition, and identified KIR2DL3 as another KIR reacting with both groups of HLA-C allotypes. Mutagenesis of amino acid 44 in KIR2DL1 and KIR2DL2 suggested this residue controls the affinity of KIR for the 77/80 motif of HLA-C molecules. Two other soluble KIR2D, derived from noninhibitory receptors with short cytoplasmic tails (KIR2DS), did not bind to any of the HLA class I allotypes tested. One of these receptors (KIR2DS2) is closely related in sequence to KIR2DL2. Substitution of tyrosine 45 with the phenylalanine conserved in other KIR was sufficient to permit specific binding of KIR2DS2 to HLA-C. These results show that KIR2DL receptors are specific for HLA-C, but that recognition of HLA-C allotypes appears more permissive than indicated by previous functional experiments.

Alleles↗
PubMed1998

Characterization of chimpanzee TCRV gene polymorphism: how old are human TCRV alleles?

The functional relevance of the majority of human T-cell receptor A and B variable region gene polymorphisms is controversial. Studies of human and nonhuman primate major histocompatibility complex (MHC) class I and II polymorphisms show that allelic lineages predate human speciation and indicate that selection favors the long-term maintenance of these advantageous mutations. We investigated at the DNA level whether 15 human TCRA and B polymorphisms exist in contemporary chimpanzee populations. Polymorphisms consisted of variable region replacements, a recombination signal sequence base change, and silent mutations. With one exception, none of these human TCR polymorphisms were observed in contemporary chimpanzees. Investigation of the same polymorphisms in a range of other nonhuman primates showed little evidence of the existence of human polymorphism prespeciation. Chimpanzee TCRAV and BV regions were however polymorphic for variation so far not observed in human groups. Levels of mitochondrial and nuclear DNA sequence variation in contemporary chimpanzees suggest that population bottlenecks have not been a feature of chimpanzee evolution and it is therefore probable that most human TCR polymorphisms have evolved in the estimated five million years since the speciation of human and chimpanzees. Thus, over the evolutionary time period studied, ancient TCRA and B polymorphisms have not been maintained by selection to the same degree as postulated for MHC polymorphisms.

Alleles↗
PubMed1998

A major histocompatibility complex class I allele shared by two species of chimpanzee.

Little is known regarding the rates at which natural selection can modify or retain antigen presenting alleles at the major histocompatibility complex (MHC). Discovery of identical [1101 base pairs (bp)] coding regions at the MHC class I C locus in Pan troglodytes and Pan paniscus, chimpanzee species that diverged approximately 2.3 million years ago, now indicates that a class I allotype can survive for at least this period. Remarkable conservation was also reflected in the (1799 bp) introns where a maximum of only six substitutions distinguished five alleles (three from P. troglodytes and two from P. paniscus) that encoded the identical heavy chain allotype. Analysis of a more distantly related human allele, HLA-Cw*0702, corroborated that intron variation was non-uniform along the gene. Thus we provide a clear reference frame for the lifetime of an MHC class I allotype, a direct estimate of allelic substitution rates, and evidence for an unusual evolution of MHC class I introns.

Alleles↗
PubMed1998

The Bw4/Bw6 difference between HLA-B*0802 and HLA-B*0801 changes the peptides endogenously bound and the stimulation of alloreactive T cells.

HLA-B*0801 is unique among HLA-B allotypes in having dominant amino acid anchors at positions 3 and 5 of the peptide-binding motif. HLA-B*0802 is a variant of HLA-B*0801 in which the Bw6 sequence motif is replaced by a Bw4 sequence motif. This change, involving substitutions at positions 77, 80, 81, 82, and 83 of the B*08 heavy chain, is probably the result of a single evolutionary event of interallelic conversion. Moreover, the difference between B*0802 and B*0801 is sufficient to stimulate a cytotoxic T-cell response. To assess further the functional impact of the Bw4 motif on a B8 background, we compared the peptide-binding specificity of the B*0801 and B*0802 allotypes by sequencing the mixture of peptides endogenously bound to B*0802 and 12 individual peptides purified from that mixture. The HLA-B*0802 allotype, while able to bind some peptides bound by B*0801, has a broader repertoire of endogenously bound peptides than B*0801: the peptides bound by B*0802 are more variable in length and exhibit greater diversity in the carboxyl-terminal amino acid which interacts with the F pocket.

Amino Acid Sequence↗