Inefficient assembly limits transport and cell surface expression of HLA-Cw4 molecules in C1R.

HLA-C antigens are expressed to the cell surface at roughly 10% the level of HLA-B or -A, and their serological definition remains persistently difficult. To characterize the factors limiting surface expression, the processes of assembly and intracellular transport of HLA-Cw4 molecules were investigated in the C1R cell line. When appropriate peptides were added to cultured cells or in cell lysates significant amounts of conformed HLA-C molecules that associate with beta 2-microglobulin (beta 2 m) are detected, but are indeed not sufficient to restore expression to the level observed for HLA-A or -B molecules. Furthermore, a precursor/product relationship exists between the free class I heavy chain and the mature conformation of HLA-Cw4 molecules. Thus, HLA-C assembly promotes the conversion of HC-10-reactive molecules (weakly-beta 2m-associated non-ligand associated free HC form) into the beta 2m-associated class I molecules recognized by W6/32. To further investigate the factors that regulate cell surface expression, intracellular transport of HLA-Cw4 was studied in pulse chase analysis. In contrast to some HLA-A and B, maturation of HLA-Cw4 heavy chains and their export to the medial and trans-Golgi compartments are quite inefficient. After 4 h of chase period, roughly half of the pulse-labeled HLA-Cw4 molecules have transited to the medial-Golgi and acquired complex oligosaccharides characteristic of mature form. In addition, treatment with gamma-interferon does not appear to improve maturation of HLA-Cw4 heavy chains, suggesting that increased supply of peptides does not influence intracellular transport. Moreover, only a small fraction in the pool of HLA-Cw4 molecules was subsequently transported through the trans-Golgi network, as indicated by their acquisition of sialic acids. Taken together these studies show that HLA-Cw4 molecules are inefficiently transported through the Golgi apparatus and presumably retained in the endoplasmic reticulum or cis-Golgi compartment.

Structural diversity in the HLA-A10 family of alleles: correlations with serology.

The HLA-A10 crossreacting group consists of the A25, A26, A34, A43 and A66 antigens. Here, we report allelic sequences for A43 and for 2 subtypes of both A26 and A34. Combining these results with previously determined sequences for A25, A26 and A66 enables molecular comparison of all the serologically defined A10 antigens. They form a closely related and well-defined group of alleles which may have originated with A*2601. Patterns of serological crossreactivity are correlated with sequence and a public epitope shared by A33 and members of the A10 family is localized to residues R62 and N63. The A*2501, A*4301 and A*6601 alleles appear to have derived from A*2601 by single gene conversion events with other HLA-A alleles. In the case of A*4301, the donor allele was probably an A29 allele as A*4301 has a small element of sequence in the alpha 1 helix (residues L62 and Q63) uniquely shared with A29. The chimaeric structure of A43 explains the reactivity of A43 molecules with both A10 and A29 alloantisera. The rare Oriental variant of A26 (A26v*) is encoded by an allele (A*2602) that differs from A*2601 by a unique nucleotide substitution which changes aspartate to asparagine at position 116 in the floor of the peptide binding groove. Thus A*2602 is a functionally distinct allele that originated by a point mutation. Alleles encoding A34 and A66 antigens are found to have very similar structures, explaining the difficulty in their serological definition.(ABSTRACT TRUNCATED AT 250 WORDS)

HLA class I nucleotide sequences, 1992.

The HLA class I sequences included in this compilation are taken from publications listed in the papers: Nomenclature for factors of the HLA system, 1991 (Bodmer et al. 1992); Nomenclature for factors of the HLA system, 1990 (Bodmer et al. 1991); and Nomenclature for factors of the HLA system, 1989 (Bodmer et al. 1990). Due to the increased number of sequences we have only included sequences for exons 2, 3, and 4 in this compilation. Where discrepancies have arisen between reported sequences, the original authors have been contacted where possible, and necessary amendments to published sequences have been incorporated into this alignment. Future sequencing may identify errors in this list and we would welcome any evidence that helps to maintain the accuracy of this compilation. In the sequence alignments, identity between nucleotides is indicated by a hyphen (-). An unavailable sequence is indicated by a period (.). Gaps in the sequence are inserted to maintain the alignment between different alleles showing variation in amino acid number.

HLA Class I nucleotide sequences, 1992.

The HLA Class I sequences included in this compilation are taken from publications listed in the papers: Nomenclature for factors of the HLA system, 1991 (1), Nomenclature for factors of the HLA system, 1990 (2) and factors of the HLA system, 1989 (3). Due to the increased number of sequences we have only included sequences for exons 2, 3 and 4 in this compilation. Where discrepancies have arisen between reported sequences, the original authors have been contacted where possible, and necessary amendments to published sequences have been incorporated into this alignment. Future sequencing may identify errors in this list and we would welcome any evidence that helps to maintain the accuracy of this compilation. In the sequence alignments, identity between nucleotides is indicated by a hyphen (-). An unavailable sequence is indicated by a period (.). Gaps in the sequence are inserted to maintain the alignment between different alleles showing variation in amino acid number. A number of sequences that do not appear in the last Nomenclature report are given in the sequence alignment. The HLA allele and the submitting authors are the following; -BeWo C.1 (S. Ellis); Cl.10, Cl.9 (L. Cianetti), Cw6W (E. Weiss); A*68012, B*5104, Cw*0803 (P. Parham); B*4802, B*52012, B*3903, C*X (D. Watkins); B*39013 (M. Takiguchi). Full information regarding these sequences will be given in the next Nomenclature report. In addition, the B*2705W allele shown in this sequence alignment and reported by the group of E. Weiss was found to differ by 3 silent substitutions from the B*2705 (that appeared in the last published alignment). Differences are the following: B*2705: position 245 (Exon 3) 'T'; position 71 (Exon 4) 'A'; position 161 (Exon 4) 'G'. B*2705W: position 245 (Exon 3) 'G'; position 71 (Exon 4) 'G'; position 161 (Exon 4) 'A'.

Serologic cross-reactivities poorly reflect allelic relationships in the HLA-B12 and HLA-B21 groups. Dominant epitopes of the alpha 2 helix.

Previous analysis has emphasized the correlation between primary structures of class I HLA molecules and their patterns of serologic cross-reactivity. Here we describe the structures of two serologic groups of HLA-B alleles for which this is not the case. HLA-B45, an allele associated with black populations, is serologically paired with B44 in the B12 group; its structure, however, is divergent from that of B44 but closely related to B50. The BN21 (B*4005) allele is associated with native Americans and is serologically grouped with B50 in the B21 group; its structure, however, is more closely related to alleles of the B40 group. The B44 and B45 serologically cross-reactive molecules differ at seven functional positions of the Ag recognition site; the B50 and BN21 molecules differ at four such residues. These differences are predicted to alter peptide presentation and be capable of eliciting strong alloreactive T cell responses. For these pairs of B12 and B21 Ag, serology appears dominated by epitopes formed by short sequences of the alpha 2 helix which have been shuffled by recombination between alleles. The implications of these results for HLA matching in transplantation are discussed.

HLA class I nucleotide sequences, 1992.

The HLA Class I sequences included in this compilation are taken from publications listed in the papers: Nomenclature for factors of the HLA system, 1991 (1), Nomenclature for factors of the HLA system, 1990 (2), and Nomenclature for factors of the HLA system, 1989 (3). Due to the increased number of sequences, we have only included sequences for exons 2, 3 and 4 in this compilation. Where discrepancies have arisen between reported sequences, the original authors have been contacted where possible, and necessary amendments to published sequences have been incorporated into this alignment. Future sequencing may identify errors in this list and we would welcome any evidence that helps to maintain the accuracy of this compilation. In the sequence alignments, identity between nucleotides is indicated by a hyphen (-). An unavailable sequence is indicated by a period (.). Gaps in the sequence are inserted to maintain the alignment between different alleles showing variation in amino acid number.

Distinctive HLA-A,B antigens of black populations formed by interallelic conversion.

Alleles encoding five HLA-A and B Ag characteristic of black populations have been isolated and their nucleotide sequences determined. In each case, the "black" allele is similar to a "related" allele found in caucasoid populations. The primary differences between these pairs of alleles are localized clusters of nucleotide substitutions that change two to five residues of the Ag recognition site. The pattern of differences indicates that the pairs of black and caucasoid alleles diverged primarily as a result of interallelic conversion events.

Distinctive polymorphism at the HLA-C locus: implications for the expression of HLA-C.

The HLA-C locus remains an enigma. The serological polymorphism is poorly defined, HLA-C molecules are expressed at the cell surface at about 10% the levels of HLA-A and -B, and their importance for antigen presentation to either CD8-bearing T cells or natural killer cells is unclear. Our understanding of HLA-C polymorphism has also lagged behind that of HLA-A and -B. We have applied the polymerase chain reaction to the characterization of cDNA encoding HLA-C antigens. Combining the recent results with previously characterized HLA-C alleles gives a data base of 26 sequences, which was used to analyze the nature of HLA-C polymorphism and compare it to the variation seen in HLA-A and -B. The sequences form 10 families of alleles that correlate well with the patterns of serological crossreactivity, including the C blanks, and all major HLA-C allelic families appear to have been sampled. The families further divide into two groups of HLA-C alleles defined on the basis of linked substitutions in the 3' exons. In comparison with HLA-A and -B, HLA-C alleles are more closely related to each other, there being less variation in residues of the antigen recognition site and more variation at other positions. In particular, the helix of the alpha 1 domain of HLA-C molecules is unusually conserved. Despite the reduced diversity in the antigen recognition site, it is evident that HLA-C genes have been the target of past selection for polymorphism. Within the antigen recognition site, it is the alpha 1 domain that is most diagnostic of HLA-C, whereas the alpha 2 domain is similar to that of HLA-B, the locus to which HLA-C is most closely related. In particular, conserved motifs in the alpha 1 helix and the conserved glycine at the base of the B pocket (position 45) provide a combination of features that is uniquely found in HLA-C molecules. We hypothesize that these features restrict the peptides bound by HLA-C molecules and in this manner reduce the efficiency of HLA-C assembly and expression at the cell surface. The overall picture HLA-C polymorphism obtained from this sampling of HLA-C alleles is unlikely to change as further alleles are characterized.

HLA class I nucleotide sequences, 1992.

The HLA class I sequences included in this compilation are taken from articles listed in the literature: "Nomenclature for Factors of the HLA System, 1991" [1], "Nomenclature for Factors of the HLA System, 1990" [2], and "Nomenclature for Factors of the HLA System, 1989" [3]. Because of the increased number of sequences, we have only included sequences for exons 2-4 in this compilation. Where discrepancies have arisen between reported sequences, the original authors have been contacted where possible, and necessary amendments to published sequences have been incorporated into this alignment. Future sequencing may identify errors in this list, and we would welcome any evidence that helps to maintain the accuracy of this compilation. In the sequence alignments, identity between nucleotides is indicated by a hyphen (-). An unavailable sequence is indicated by a period (.). Gaps in the sequence are inserted to maintain the alignment between different alleles showing variation in amino acid number.

HLA-J, a second inactivated class I HLA gene related to HLA-G and HLA-A. Implications for the evolution of the HLA-A-related genes.

Ragoussis and co-workers (Genomics 4:301) previously described a class I HLA gene (now designated HLA-J) that maps to within 50 kb of HLA-A. The nucleotide sequences of three HLA-J alleles are reported here. Comparison of the nucleotide sequences of HLA-J alleles shows this gene is more related to HLA-G, A, and H than to HLA-B, C, E, and F. All four alleles of HLA-J are pseudogenes because of deleterious mutations that produce translation termination either in exon 2 or exon 4. Apart from these mutations, the predicted proteins have structures similar to those of HLA-A, B, and C molecules. There is, however, little polymorphism at HLA-J and none at functional positions of the Ag-recognition site. The polymorphism is less than found for HLA-H another HLA-A-related pseudogene. HLA-J appears, like HLA-H, to be an inactivated gene that result from duplication of an Ag-presenting locus related to HLA-A. Nucleotide sequence comparisons show that the HLA-A, H, J, and G genes form a well defined group of "HLA-A-related" loci. Evolutionary relationships as assessed by construction of trees suggest the four modern loci: HLA-A, G, H, and J were formed by successive duplications from a common ancestral gene. In this scheme one intermediate locus gave rise to HLA-A and H, the other to HLA-G and J.

Unusual HLA-B alleles in two tribes of Brazilian Indians.

The Kaingang and Guarani are culturally and linguistically distinct tribes of southern Brazil. Like all Amerindian groups they show limited HLA polymorphism, which probably reflects the small founder populations that colonized America by overland migration from Asia 11,000-40,000 years ago. We find the nucleotide sequences of HLA-B alleles from the Kaingang and Guarani to be distinct from those characterized in caucasian, oriental and other populations. By comparison, the HLA-A and C alleles are familiar. These results and those reported in the accompanying paper on the Waorani of Ecuador reveal that a marked evolution of HLA-B has occurred since humans first entered South America. New alleles have been formed through recombination between pre-existing alleles, not by point mutation, giving rise to distinctive diversification of HLA-B in different South American Indian tribes.

The molecular basis for reactivity of anti-Cw1 and anti-Cw3 alloantisera with HLA-B46 haplotypes.

HLA haplotypes containing the HLA-B46 allele react with both anti-Cw1 and anti-Cw3 alloantisera, a pattern of reactivity defined as the Cw11 antigen and postulated to involve either a distinctive Cw11 allele or a duplicated HLA-C locus. From serological characterization of CIR cells transfected with B46 cDNA we now demonstrate that the anti-Cw3 reactivity with these haplotypes is solely due to the B46 molecule and not to an HLA-C molecule. Furthermore, isolation and characterization of HLA-C mRNA from cells expressing B46 strongly suggest that anti-Cw1 reactions are directed against the product of a conventional Cw1 allele. The antigenic cross-reactivities of B46 with B62 and Cw3 correlate with its chimaeric primary structure, which is identical to that of B62, except in the alpha 1 helix where it is identical to both Cw3 and Cw1. The structure, distribution and genetic linkage of B46 indicate it is of recent, Asian origin and is the result of a gene conversion, involving Cw1 as the donor gene and B62 as the recipient. These results demonstrate that the Cw11 antigen neither corresponds to a novel HLA-C allele nor a duplicated HLA-C locus, but to a combination of epitopes contributed by linked Cw1 and B46 alleles. The nucleotide sequence we previously and erroneously attributed to a distinct Cw11 allele is now demonstrated to encode Cw8. Isolation of the cDNA clone with this sequence from a library made from a cell homozygous for the B46 haplotype was probably an artefact of contamination.

The HLA-A,B "negative" mutant cell line C1R expresses a novel HLA-B35 allele, which also has a point mutation in the translation initiation codon.

The HLA-A,B negative mutant cell line C1R is widely used as a transfection recipient in functional studies of class I MHC genes. It was derived from a normal B cell line, Hmy2, by three rounds of mutagenesis and immunoselection with anti-HLA mAb. Serology characterizes C1R to be negative for the HLA-A2, A3, B35, Bw62, and Cw3 Ag of the parental cell line while retaining expression of HLA-Cw4. We find, however, that CTL specific for HLA-B35 lyse C1R cells, suggesting that expression of HLA-B35 is also retained. To resolve this paradox we examined the expression of HLA-A,B,C genes and proteins in C1R cells. The results are consistent with deletion of the HLA-A3, Bw62, Cw3 haplotype and retention of the HLA-A2, B35, Cw4 haplotype in C1R. Although present, the HLA-A2 gene appears not to be transcribed. As expected, the HLA-Cw4 gene is transcribed and the protein expressed at normal levels. Transcription of the HLA-B35 gene is also normal and comparable to that of HLA-Cw4. However, expression of the HLA-B35 protein is reduced to a few percent of the parental level. Comparison of the nucleotide sequence of B35 alleles from C1R and Hmy2 revealed that reduced translation in C1R is caused by a point mutation (ATG to TTG) in the translation initiation codon. The HLA-B35 allele from C1R and Hmy2 represents a novel subtype, B*3503, differing from B*3501 by replacement of serine by phenylalanine at the peptide binding position 116. This study shows cell surface levels of a class I molecule which are insensitive to lysis by antibody and complement can be readily recognized by alloreactive T cells, further illustrating the relative sensitivity of Ag recognition by T cells.

HLA class I nucleotide sequences, 1991.

The HLA class I sequences included in this compilation are taken from publications listed in the accompanying paper, Nomenclature for factors of the HLA system, 1990 (Bodmer et al., 1991), and also in Nomenclature for factors of the HLA system, 1989 (Bodmer et al., 1990). Where discrepancies have arisen between reported sequences the original authors have been contacted where possible, and necessary amendments to published sequences have been incorporated into this alignment. Future sequencing may identify errors in this list and we would welcome any evidence that helps to maintain the accuracy of this compilation. In the sequence alignments identity between residues is indicated by a hyphen (-). An unavailable sequence is indicated by a full point (.). Gaps in the sequence are inserted to maintain the alignment between different alleles showing variation in amino acid number.

HLA class I nucleotide sequences, 1991.

The HLA Class I sequences included in this compilation are taken from publications listed in the accompanying paper, Nomenclature for factors of the HLA system, 1990 and Nomenclature for factors of the HLA system, 1989. Where discrepancies have arisen between reported sequences the original authors have been contacted where possible, and necessary amendments to published sequences have been incorporated into this alignment. Future sequencing may identify errors in this list and we would welcome any evidence that helps to maintain the accuracy of this compilation. In the sequence alignments identity between residues is indicated by a hyphen (-). Unavailable sequence is indicated by a period (.). Gaps in the sequence are inserted to maintain the alignment between different alleles showing variation in amino acid number.