Class II genes of the human major histocompatibility complex. Comparisons of the DQ and DX alpha and beta genes

The human major histocompatibility complex, HLA, contains the genes of several class II molecules. We present here the molecular maps of the DQ and DX subregions and analyze the sequences of the polymorphic DQ alpha and DQ beta genes as well as the DX alpha and DX beta genes. The DQ alpha and DQ beta genes are oriented in opposite directions, approximately 12 kilobases apart. The DX alpha and DX beta genes are similarly oriented about 8 kilobases. The exon-intron organizations of the DQ alpha and DX alpha genes are analogous to those of other class II alpha genes. Comparison of the DQ alpha gene sequence to three DQ alpha cDNA clones shows that amino acid replacements are predominantly located between residues 45 and 80 in the amino-terminal domain. Analysis of the frequency of silent and replacement substitutions indicates that there is little selection against replacements in DQ alpha first domains. The exons encoding the second domains of DQ alpha and DX alpha are virtually identical, suggesting that a gene conversion event has occurred between these genes. The DX beta gene is very similar to the DQ beta gene but differs in the cytoplasmic portion. The DX beta gene contains a separate exon of 24 nucleotides encoding the core of the cytoplasmic tail. This exon is not expressed in the DQ beta genes due to a nonfunctional splice junction. Comparison of the number of nucleotide substitutions in the DQ beta first and second domain exons suggests that little or no phenotypic selection acts on the first domain whereas the second domain is under strong selection.     

Class II genes of the human major histocompatibility complex. The DO beta gene is a divergent member of the class II beta gene family

A novel class II beta chain gene is described. This gene, tentatively called DO beta, displays considerably less polymorphism than beta genes of the DP, DQ, and DR loci. The nucleotide sequence of the DO beta gene is strikingly similar to that of the previously identified murine A beta 2 gene. The DO beta gene displays the same exon/intron organization as other beta genes although the fifth exon and the translated portion of the sixth exon are longer than in other genes. A striking feature of the amino acid sequence deduced from the DO beta gene sequence is the pronounced hydrophobicity of the NH2-terminal region. This feature distinguishes the putative DO beta chain from other class II beta chains and raises the possibility that DO beta chains may interact with an alpha chain that is structurally different from those of the DP, DQ, and DR loci. It further suggests that the putative DO molecule may have a function different from those of other class II antigens.     

Class II genes of the human major histocompatibility complex. Evolution of the DP region as deduced from nucleotide sequences of the four genes

The DP region of the human major histocompatibility complex contains two alpha genes and two beta genes. The DP alpha 1 and beta 1 genes encode the expressed DP histocompatibility antigen molecule, while the DP alpha 2 and beta 2 genes are inactive in the haplotypes examined. Here we present the sequence of the two DP beta genes and of the expressed DP alpha 1 gene. Nucleotide sequence comparisons reveal a considerably greater degree of similarity between the two beta genes than between the two alpha genes. We propose that a duplication giving rise to the DP alpha gene pair evolutionarily preceded the corresponding DP beta gene duplication. We also propose, based on the orientation of other class II gene pairs, that the original DP molecule was encoded by the DP beta 1 and DP alpha 2 genes. At some stage during the evolution of the DP region both of the two pseudogenes appear to have been expressed.     

Linkage map of two HLA-SB beta and two HLA-SB alpha-related genes: an intron in one of the SB beta genes contains a processed pseudogene

Three overlapping cosmid clones contain coding sequences for four HLA Class II genes, provisionally identified as two HLA-SB alpha and two HLA-SB beta genes. The genes are in the order beta, alpha, beta, alpha, inverted with respect to each other. One of the SB beta genes contains a 513 bp sequence that appears to be a processed pseudogene, flanked by direct 17 bp repeat sequences, in the intron upstream of the beta 1 exon. The pseudogene is homologous to a family of sequences of approximately 25-40 members, most of which are not on chromosome 6. A cDNA clone, highly homologous to the pseudogene, except for its 5' end, contains a normal poly(A) addition site and a poly(A) tail. The cDNA clone is homologous to a single-copy gene in both man and mouse, encoded on human chromosome 15. A search of published DNA sequences identified a mouse sequence, with about 77% similarity to the pseudogene sequence, in the negative strand of an intron in a mouse dihydrofolate reductase gene. The second SB beta gene does not contain the pseudogene sequence.     

Molecular map of the human HLA-SB (HLA-DP) region and sequence of an SB alpha (DP alpha) pseudogene.

The human major histocompatibility complex contains the genes for at least three different types of class II antigens, DR, DC and SB (DR, DQ and DP). They are all composed of an alpha and a beta chain. We have cloned a chromosomal region of 70 kb containing the SB (DP) gene family in overlapping cosmid clones. This segment contains two alpha genes and two beta genes, located in the order SB alpha 1, SB beta 1, SB alpha 2 and SB beta 2. The orientation of the alpha genes is reversed compared with that of the beta genes. This organisation suggests that the SB region has arisen by duplication of a chromosomal segment encompassing one alpha and one beta gene. Partial nucleotide sequences of the SB alpha 1 and SB beta 1 exons demonstrate that the genes correspond to SB alpha and beta cDNA clones. Consequently these genes are expressed. In contrast nucleotide sequence determination of the SB alpha 2 gene shows that it is a pseudogene.     

Sequence analysis of the human major histocompatibility gene SX alpha.

The DP subregion of the human major histocompatibility complex contains two closely linked gene pairs, DP alpha, DP beta and SX alpha, SX beta. The exon-intron organization and the complete DNA sequence of the SX alpha gene are reported here. There are several mutations within the SX alpha gene which strongly suggest that it is a pseudogene. These include two frameshift mutations, one in the alpha 1 domain and the other in the cytoplasmic domain. A 5' splice site mutation at the end of the alpha 1 exon also exists. DNA sequence homology between DP alpha and SX alpha suggests that these genes arose through a gene duplication event.     

Structures of the genes encoding the alpha and beta subunits of the Neurospora crassa mitochondrial ATP synthase.

We have isolated and sequenced cDNA and genomic clones encoding the alpha and beta subunits of the Neurospora crassa ATP synthase. The genes are not linked to each other: atp-1(alpha) maps to either linkage group I or V, and atp-2(beta) lies on linkage group II. The two genes resemble each other in having a large number of introns, five in atp-1 and seven in atp-2, mostly positioned near their 5' ends and varying in length from 60-332 bp. The coding regions of both genes have a high G+C content (59%) and use a low number of codons, 46 (atp-1) and 44 (atp-2), a feature associated with highly expressed genes. Northern-blot analysis shows both genes are expressed at high levels during mycelial growth. Comparison of the exon-intron structures of the beta-subunit-encoding gene with those from human and tobacco showed a similar number of introns, several closely positioned, but no exact conservation in position, size or sequence of introns.     

Studies on the organization of the human apolipoprotein B 100 gene

The organization of five exons of the 3' terminal end of the human apolipoprotein B 100 (apo B 100) gene 1906, 184, 115, 7572 and 374 bp long have been determined from two overlapping EMBL3 human genomic clones extending over 18 kb. They encode more than 70% of the apo B 100 amino-acid sequence. The introns between these five exons were sequenced revealing the common intron/exon splice junction sequences. The 7572 bp exon is the longest exon so far reported for mammalian genes with the proposed sequence coding for the LDL receptor binding site. Its possible relationship to apolipoprotein B 48 is discussed.     

Location of the 11 bp exon in the chicken pro alpha 2(I) collagen gene.

During the fine structural analysis of the 5' end of the 38 kb chicken pro alpha 2(I) collagen gene, we failed to locate an exon, only 11 bp in size, which had been predicted from the DNA sequence analysis of a cDNA clone complementary to the 5' end of the pro alpha 2(I) collagen mRNA (1). We know report the location of this 11 bp exon, exon 2, at the 5' end of a 180 bp Pst I fragment, 1900 bp 3' to exon 1 and 600 bp 5' to exon 3. Its sequence, ATGTGAGTGAG, is highly unusual in that it contains two overlapping consensus donor splice sequences. Moreover, it is flanked by two overlapping donor splice sequences but only one of the four splice sequences is actually spliced (1). The first half of intron 1 also has an unusual sequence: it is 68% GC, contains 88 CpG dinucleotides and 11 Hpa II sites. The second half is more like other intron sequences in the collagen gene with a GC content of 41%, 19 CpG, and no Hpa II sites. However it contains two sequences with 7 and 9 bp homology to the 14 bp SV40 enhancer core sequence. It is suggested that some part of intron 1 may be involved in regulation.     

Internal organization of the major adult alpha- and beta-globin genes of X. laevis

We describe the isolation of two recombinant lambda phages, each containing genomic DNA fragments encoding both the major adult alpha- and beta-globin mRNAs of X. laevis. The DNA fragment in the two clones have restriction maps which indicate that they are each derived from a different member of the pair of alleles present in the heterozygote used as the source of DNA for cloning. The characterization of these two clones by restriction mapping, R looping and DNA sequencing shows that the alpha 1- and beta 1-globin genes lie in the orientation separated by 7.7 kb of DNA. There are two introns in the alpha 1-globin gene and two in the beta 1-globin gene, and they interrupt the genes at exactly the same positions as the introns found in all known mammalian alpha- and beta-globin genes. The exon sequences proximal to the introns show a much higher degree of homology with mammalian sequences than the sequences distal to intron/exon junctions, and the introns in the beta 1-globin gene of X. laevis are very similar in length to the corresponding introns in the beta-globin genes of several mammals and the chicken.     

Distribution of introns in frameshift-suppressor proline-tRNA genes of Saccharomyces cerevisiae

Mutations in the suf9, suf10, and suf11 genes of yeast suppress + 1 nucleotide (nt) insertions in proline codons. Nucleotide sequence analysis indicates that the suf9 and suf11 genes are members of the proline tRNA(UGG) gene family, which also includes three other previously identified genes, suf7, suf8, and trn1. All five members of this gene family contain introns. The suf9 and suf11 introns are 31 and 30 nt in length, respectively, and are similar but not identical in sequence to other introns within the family. The suf10 gene is identical in sequence to suf2, which was shown previously to encode proline tRNA(IGG). Both members of this gene family lack introns. Alleles of suf9, suf10, and suf11 that confer frameshift suppression were also analyzed. The SUF9-1 allele results in a G----U substitution at nt position 39 in the anticodon stem. The recessive suf11-1 allele is a double mutant containing the same nt position 39 alteration as in SUF9-1 plus a second U----A substitution at nt position 38 in the anticodon loop. The SUF10-1 suppressor mutation corresponds to a +1G insertion in the anticodon loop. Since the nt substitutions in suf11-1 alter the sequence of the 3' exon/intron boundary, the double mutant pre-tRNA was tested for its ability to be cleaved in vitro by tRNA-splicing endonuclease. It was found that suf11-1 pre-tRNA is cleaved with reduced efficiency at the 3' splice junction.     

Molecular cloning and characterization of the human topoisomerase IIalpha and IIbeta genes: evidence for isoform evolution through gene duplication

Human DNA topoisomerase II is essential for chromosome segregation and is the target for several clinically important anticancer agents. It is expressed as genetically distinct alpha and beta isoforms encoded by the TOP2alpha and TOP2beta genes that map to chromosomes 17q21-22 and 3p24, respectively. The genes display different patterns of cell cycle- and tissue-specific expression, with the alpha isoform markedly upregulated in proliferating cells. In addition to the fundamental role of TOP2alpha and TOP2beta genes in cell growth and development, altered expression and rearrangement of both genes are implicated in anticancer drug resistance. Here, we report the complete structure of the human topoisomerase IIalpha gene, which consists of 35 exons spanning 27.5 kb. Sequence data for the exon-intron boundaries were determined and examined in the context of topoisomerase IIalpha protein structure comprising three functional domains associated with energy transduction, DNA breakage-reunion activity and nuclear localization. The organization of the 3' half of human TOP2beta, including sequence specifying the C-terminal nuclear localization domain, was also elucidated. Of the 15 introns identified in this 20 kb region of TOP2beta, the first nine and the last intron align in identical positions and display the same phases as introns in TOP2alpha. Though their extreme 3' ends differ, the striking conservation suggests the two genes diverged recently in evolutionary terms consistent with a gene duplication event. Access to TOP2alpha and TOP2beta gene structures should aid studies of mutations and gene rearrangements associated with anticancer drug resistance.