Nucleotide sequences of Caenorhabditis elegans core histone genes. Genes for different histone classes share common flanking sequence elements

We have determined the nucleotide sequence of core histone genes and flanking regions from two of approximately 11 different genomic histone clusters of the nematode Caenorhabditis elegans. Four histone genes from one cluster (H3, H4, H2B, H2A) and two histone genes from another (H4 and H2A) were analyzed. The predicted amino acid sequences of the two H4 and H2A proteins from the two clusters are identical, whereas the nucleotide sequences of the genes have diverged 9% (H2A) and 12% (H4). Flanking sequences, which are mostly not similar, were compared to identify putative regulatory elements. A conserved sequence of 34 base-pairs is present 19 to 42 nucleotides 3' of the termination codon of all the genes. Within the conserved sequence is a 16-base dyad sequence homologous to the one typically found at the 3' end of histone genes from higher eukaryotes. The C. elegans core histone genes are organized as divergently transcribed pairs of H3-H4 and H2A-H2B and contain 5' conserved sequence elements in the shared spacer regions. One of the sequence elements, 5' CTCCNCCTNCCCACCNCANA 3', is located immediately upstream from the canonical TATA homology of each gene. Another sequence element, 5' CTGCGGGGACACATNT 3', is present in the spacer of each heterotypic pair. These two 5' conserved sequences are not present in the promoter region of histone genes from other organisms, where 5' conserved sequences are usually different for each histone class. They are also not found in non-histone genes of C. elegans. These putative regulatory sequences of C. elegans core histone genes are similar to the regulatory elements of both higher and lower eukaryotes. The coding regions of the genes and the 3' regulatory sequences are similar to those of higher eukaryotes, whereas the presence of common 5' sequence elements upstream from genes of different histone classes is similar to histone promoter elements in yeast.     

The uni chromosome of Chlamydomonas: histone genes and nucleosome structure.

The uni linkage group (ULG) of Chlamydomonas reinhardtii contains many genes involved in the basal body-flagellar system. Recent evidence suggests that the corresponding uni chromosome is located in close proximity to the basal body complex. In the course of studies into its molecular organization, we have found a cluster of four histone genes on the ULG. The genes are arranged as divergently-transcribed pairs: H3-H4 and H2B-H2A. Genomic sequencing reveals that these genes lack introns and contain characteristic 3' palindromes similar to those of animals. The predicted amino acid sequences are highly conserved across species, with greatest similarities to the histone genes of Volvox. Southern analysis shows that each histone gene is present in 15-20 copies in Chlamydomonas and suggests a dispersed genomic organization. Northern analysis of mitotically-synchronized cells shows that, like the replication-dependent histones of higher eukaryotes, Chlamydomonas histone genes are expressed during S-phase. Using a gene-specific probe on Northern blots, we provide evidence that the ULG H4 gene is regulated in the same manner as other Chlamydomonas histone genes. Finally, micrococcal nuclease protection experiments show that the uni chromosome itself associates with histone proteins and displays a conventional nucleosomal banding pattern.     

Genes encoding a histone H3.3-like variant in Arabidopsis contain intervening sequences.

Two genes encoding a particular H3 histone variant were isolated from Arabidopsis thaliana. These genes differ from the H3 genes previously cloned from Arabidopsis and other plants by several interesting properties: (1) the two genes are located close to each other; (2) their coding regions are interrupted by two or three small introns, the two closest to the initiation codon being located at the same place in the two genes; (3) another, long intron is located in the 5'-untranslated region just before the initiation codon of gene I as deduced from the sequence of several corresponding cDNAs, and very likely also of gene II; (4) these genes do not show preferential expression in organs containing meristematic tissues contrary to the classical intronless replication-dependent histone genes, thus suggesting that their expression is not replication-dependent; (5) the protein encoded by both genes is the same and corresponds to a minor H3 variant highly conserved among all the plant species studied up to now. All these characteristics are common with the animal replication-independent H3.3 histone genes and it is assumed that the genes described here are the first example of the equivalent H3.3 gene family in plants. Interestingly, the promoter regions of the two genes have the same general structure as the Arabidopsis intronless genes. Possible implications on the regulation of H3 genes expression are discussed.     

The human H2A and H2B histone gene complement

Sequences and expression patterns of newly isolated human histone H2A and H2B genes and the respective proteins were compared with previously isolated human H2A and H2B genes and proteins. Altogether, 15 human H2A genes and 17 human H2B genes have been identified. 14 of these are organized as H2A/H2B gene pairs, while one H2A gene and three H2B genes are solitary genes. Two H2A genes and two H2B genes turned outto be pseudogenes. The 13 H2A genes code for at least 6 different amino acid sequences, and the 15 H2B genes encode 11 different H2B isoforms. Each H2A/H2B gene pair is controlled by a divergent promoter spanning 300 to 330 nucleotides between the coding regions of the two genes. The highly conserved divergent H2A/H2B promoters can be classified in two groups based on the patterns of consensus sequence elements. Group I promoters contain a TATA box for each gene, two Oct-1 factor binding sites, and three CCAAT boxes. Group II promoters contain the same elements as group I promoters and an additional CCAAT box, a binding motif for E2F and adjacent a highly conserved octanucleotide (CACAGCTT) that has not been described so far. Five of the 6 gene pairs and 4 solitary genes with group I promoters are localized in the large histone gene cluster at 6p21.3-6p22, and one gene pair is located at 1q21. All group II promoter associated genes are contained within the histone gene subcluster at D6S105, which is located at a distance of about 2 Mb from the major subcluster at 6p21.3-6p22 containing histone genes with group I promoters. Almost all group II H2A genes encode identical amino acid sequences, whereas group I H2A gene products vary at several positions. Using human cell lines, we have analyzed the expression patterns of functional human H2A/H2B gene pairs organized within the two histone gene clusters on the short arm of chromosome 6. The genes show varying expression patterns in different tumor cell lines.     

Structure of the three beta-tubulin-encoding genes of the unicellular alga, Polytomella agilis

The quadriflagellate, unicellular, colorless alga, Polytomella agilis, contains several distinct microtubule arrays. To study the genetic basis of microtubule heterogeneity in P. agilis, we characterized its tubulin(Tub)-encoding genes (tub). The three beta tub genes detected in blots of P. agilis DNA were isolated from a genomic library. The structure and organization of the genes were examined by restriction mapping and nucleotide (nt) sequencing. S1 nuclease protection studies showed that all three genes are expressed. The predicted amino acid (aa) sequences are more than 98% conserved with the Chlamydomonas reinhardtii and Volvox carteri beta-Tubs, underscoring the close phylogenetic relationship of these species. Evolutionary divergence among the P. agilis genes is demonstrated by differences in intron number, nt sequences in noncoding regions, and silent nt substitutions in the coding regions. However, the proteins encoded by the beta 1 and beta 3 tub genes are identical; the beta 2 gene product differs by one conservative aa substitution. These results are in striking contrast to the C-terminal aa diversity reported within beta tub gene families in animal, higher plant and fungal systems. The data support the hypothesis that those tub genes whose products assemble into axonemal microtubules are subject     

Sequence comparisons of non-allelic late histone genes and their early stage counterparts. Evidence for gene conversion within the sea urchin late stage gene family

We have determined the nucleotide sequence of sea urchin (Lytechinus pictus) late stage H3 and H4 histone genes contained on the clone pLpH3H4 -21 and of the early stage H3 gene contained on the plasmid pLpA . Comparison of these differentially regulated histone genes with each other and with other L. pictus late and early stage histone H3 and H4 genes previously sequenced confirms that members of each histone gene family (early and late) are more homologous to each other than they are to members of other histone gene families. The spacer regions between two late H3-H4 gene pairs on the clones pLpH3H4 -19 and pLpH3H4 -21 have diverged to the point where they are no longer homologous. However, comparative analysis of the 5' flanking DNA has identified a sequence 5'C-T-C-A-T-G-T-A-T-T3' upstream of both late H4 genes and another, 5'A-G-A-T-T-C-A3', upstream of both H3 genes. Except for a short conserved sequence near the initiation codon, the transcribed 5' leaders of the late mRNAs differ in length and sequence in the two non-allelic late histone gene pairs. This divergence contrasts with the 95 to 96% conservation found between late histone gene coding sequences. The results suggest that there is intergenic exchange in the germline among members of the late histone gene family and that the unit of exchange is the individual gene rather than the heterotypic dimer which includes the common spacer DNA.     

Nonallelic histone gene clusters of individual sea urchins (Lytechinus pictus): mapping of homologies in coding and spacer DNA

The linear arrangement and lengths of the spacers and coding regions in the two nonallelic histone gene variant clusters of L. pictus are remarkably homologous by R loop analysis and are similar in general topography to the histone gene repeat units of other sea urchins examined to date. No interventing sequences were detected. The coding regions of these two histone gene variants share considerable sequence homology; however, there are areas of nonhomology in every spacer region and the lengths of the nonhomologous spacers between the H2A and H1 genes are not the same for the two repeat unit classes (inter-gene heterogeneity). Combining length measurements obtained with both R loops and heteroduplexes suggests that the DNA sequences of the analogous leader regions for the two H1 mRNAs are nonhomologous. Similar observations were made for the H4 leader sequences, as well as the trailer region on H2B. S. purpuratus spacer DNA segments share little sequence homology with L. pictus; however, the analgous coding (and possibly flanking) regions have conserved their sequences. The various coding and spacer regions within a repeat unit do not share DNA sequences. Thus certain areas in the sea urchin histone gene repeat units have been highly conserved during evolution, while other areas have been allowed to undergo considerable sequence change not only between species but within a species.     

Distribution of introns in fungal histone genes

Saccharomycotina and Taphrinomycotina lack intron in their histone genes, except for an intron in one of histone H4 genes of Yarrowia lipolytica. On the other hand, Basidiomycota and Perizomycotina have introns in their histone genes. We compared the distributions of 81, 47, 79, and 98 introns in the fungal histone H2A, H2B, H3, and H4 genes, respectively. Based on the multiple alignments of the amino acid sequences of histones, we identified 19, 13, 31, and 22 intron insertion sites in the histone H2A, H2B, H3, and H4 genes, respectively. Surprisingly only one hot spot of introns in the histone H2A gene is shared between Basidiomycota and Perizomycotina, suggesting that most of introns of Basidiomycota and Perizomycotina were acquired independently. Our findings suggest that the common ancestor of Ascomycota and Basidiomycota maybe had a few introns in the histone genes. In the course of fungal evolution, Saccharomycotina and Taphrinomycotina lost the histone introns; Basidiomycota and Perizomycotina acquired other introns independently. In addition, most of the introns have sequence similarity among introns of phylogenetically close species, strongly suggesting that horizontal intron transfer events between phylogenetically distant species have not occurred recently in the fungal histone genes.     

Genomic organization and nucleotide sequences of two corn histone H4 genes

The sea urchin histone H4 gene has been used as a probe to clone two corn histone H4 genes from a lambda gtWES X lambda B corn genomic library. The nucleotide (nt) sequences of both genes showed that the encoded amino acid sequences were identical to that of the H4 of pea and one variant of wheat. The nt sequences of the coding regions showed 92% homology. 5'- and 3'-flanking regions do not show extensive nt sequence analogies. Southern blotting of the EcoRI digested genomic DNA suggests the existence of multiple H4 genes dispersed throughout the genome.     

Histone genes inPhysarum polycephalum: Transcription and analysis of the flanking regions of the two H4 genes

The histone H4 multigene family of Physarum polycephalum consists of two genes, H41 and H42. Both genes have an unusual structure in that they are interrupted by a small intron. The structure of the P. polycephalum H4 genes is discussed and compared to the structure of histone genes of other organisms. S1 nuclease analysis was used to map the 5' and 3' ends of the histone H4 messengers. We show that the histone H4 genes have a hybrid structure; they are interrupted by an intervening sequence, as in replacement variant histone genes of higher eukaryotes, but their 5' and 3' noncoding regions have the properties of replication-dependent histone genes: the 5' and 3' leader and trailer sequences are short, possess a 3'-hyphenated dyad symmetry element, and a CAGA sequence is found 3' to the hyphenated hairpin structure. This report also provides evidence that both genes are expressed in late G2 phase as well as in S phase and that their expression is temporally coordinated and quantitatively similar during the cell cycle.