Histone H4 and H2B genes in rainbow trout (Salmo gairdnerii)

Summary The complete nucleotide sequence of the 3.0-kb BamH I-Sst I restriction fragment contained within the rainbow trout genomic clone TH2 has been determined. This region contains the rainbow trout H4 and H2B histone genes and 5 and 3 flanking and spacer sequences, and represents the 5 half of the histone-gene cluster; the remaining half has been characterized previously. The genes are uninterrupted, and are transcribed from the same strand. The protein sequence of H4, as determined from the nucleic acid sequence, is the same as that derived for other vertebrate H4 proteins, although comparison of nucleotide sequences shows a great deal of sequence divergence, especially in the third base position. The amino acid sequence of H2B, though largely homologous to those of other vertebrate H2B proteins, displays some characteristic differences in primary structure. Consensus sequences noted in many other eukaryotic genes, as well as histone-specific consensus sequences, have been identified. An unusual feature of the spacer region between the H4 and H2B genes is the presence of a duplicated sequence 87 bp in length. The 5 and 3 ends of each repeat are complementary, and each repeat contains smaller repeated sequences internally, as well as a possible cruciform structure.     

Further genetic variation at the esterase loci of Drosophila virilis

Reexamination of the electrophoretic mobilities of esterases encoded by the Est- and the Est- alleles of Drosophila virilis was carried out in detail using both thin-layer agar gel and polyacrylamide slab gel electrophoresis. Many allelic products with fine differences in their electrophoretic mobilities were found and designated by a new system. Some esterases separable by the agar gel method were indistinguishable using the polyacrylamide gel method. But the polyacrylamide gel method uncovered two multiband homozygotes, ^(d).77 and ^(d)1.28. Some allelic frequencies on the basis of the new designation were estimated in two natural populations. As a result, it is proposed that the total scope of allelic variation at the two esterase loci of Drosophila virilis is composed of discrete distribution patterns of gene frequencies, each histogram of which shows a bell-shaped pattern.     

Evolutionary divergence of the cytochrome b5 gene of Drosophila

Cytochrome proteins perform a broad spectrum of biological functions ranging from oxidative metabolism to electron transport and are thus essential to all organisms. The b-type cytochrome proteins bind heme noncovalently, are expressed in many different forms and are localized to various cellular compartments. We report the characterization of the cytochrome b₅ (Cyt-b) gene of Drosophila virilis and compare its structure to the Cyt-b gene of Drosophila melanogaster. As in D. melanogaster, the D. virilis gene is nuclear encoded and single copy. Although the intron/exon structures of these homologues differ, the Cyt-b proteins of D. melanogaster and D. virilis are approximately 75% identical and share the same size coding regions (1,242 nucleotides) and protein products (414 amino acids). The Drosophila Cyt-b proteins show sequence similarity to other b-type cytochromes, especially in the N-terminal heme-binding domain, and may be targeted to the mitochondrial membrane. The greatest levels of similarity are observed in areas of potential importance for protein structure and function. The exon sequences of the D. virilis Cyt-b gene differ by a total of 292 base changes. However, 62% of these changes are silent. The high degree of conservation between species separated by 60 million years of evolution in both the DNA and amino acid sequences suggests this nuclear cytochrome b₅ locus encodes an essential product of the Drosophila system.Correspondence to: C.E. Rozek     

The genetics of esterases in Drosophila. VIII. The gene regulating the activity of JH-esterase in D. virilis

The content of JH-esterase was assayed by radial immunodiffusion in Drosophila virilis pupae under normal conditions and under the effects of extreme factors. It was found that JH-esterase content is the same (not different from the control) in pupae showing a high activity of the enzyme and in those not showing it. These data are evidence for a gene controlling JH-esterase activity. It was also shown that a regulatory factor converts inactive into active JH-esterase when homogenates of pupae, with active and inactive forms, were mixed and incubated together. It was demonstrated that the source of the activating factor is the larval brain. Sublines 147-R and 147-I were produced by introducing the second chromosome pair of stocks 103 and 101, which are heat resistant, into the genome of individuals of stock 147, which is heat sensitive. Sublines 160-III, 160-IV, 160-V, and 160-VI were produced by introducing the third, fourth, fifth, and sixth chromosome pairs of stock 147 into the genome of stock 160S, which is heat-resistant. The results of analysis of JH-esterase activity and the viability of individuals of these sublines at high temperatures indicated that the gene regulating the activity of JH-esterase is located in the sixth chromosome of D. virilis.     

Variability and inheritance of histone genes H3 and H4 in Vicia faba

Summary We have compared copy numbers and blothybridization patterns of histone genes (H3 plus H4) between and within individuals of broad bean (Vicia faba). Copy number differences among individuals in the population of 200 individuals were as great as 27 fold, and as much as 3.2 fold among separate leaves of the same plant. Among F₂ progeny from genetic crosses, up to a 5.4-fold range was seen (mean=3.5 fold), and among F₁ progeny of self-pollinated plants, up to a 5.9-fold range was observed (mean=2.3 fold). Histone gene blot-hybridization patterns for EcoRI and HindIII were also variable among individuals and indicated that the genes are probably clustered in only a few chromosomal loci. The degree of variation in histone gene copy number per haploid genome (2–55 copies, or 27 fold) was similar to that found previously for ribosomal RNA genes (230–22000, or 95 fold) of V. faba. However, the two gene families change independently, since individuals with a high or low copy number for one gene can have either a high or low copy number for the other. The mechanisms(s) for rapid gene copy number change may be similar for these gene families.     

Conserved organization of an avian histone gene cluster with inverted duplications of H3 and H4 genes

Summary The organization of histone gene clusters of the duckCairina moschata was studied in the DNA inserts of two recombinant phage that overlap and feature identical histone gene arrangements but differ in sequence details and in the extent of repetition of an AT-rich motif in one of the nontranscribed spacer regions. These few but substantial differences between otherwise nearly identical histone gene groups suggest that we have independently isolated alleles of the same site of the duck genome or that this gene arrangement occurs (with slight variations) more than once per haploid genome. Within the histone gene cluster described, H3 and H4 genes are duplicated (with inverted orientation), whereas one H1 gene is flanked by single H2A and H2B genes. The arrangement of duck histone genes described here is identical to a subsection of the chicken genome but differs from any other published histone gene cluster.     

Sequence evolution of theGpdh gene in theDrosophila virilis species group

The nucleotide sequence of theGpdh gene from six taxa,D. virilis, D. lummei, D. novamexicana, D. a. americana, D. a. texana andD. ezoana, belonging to thevirilis species group was determined to examine details of evolutionary change in the structure of theGpdh gene. TheGpdh gene is comprised of one 5 non-translated region, eight exons, seven introns and three 3 non-translated regions. Exon/intron organization was identical in all the species examined, but different from that of mammals. Interspecific nucleotide divergence in the entireGpdh gene followed the common pattern: it was low in the exon, high in the intron and intermediate in the non-translated regions. The degree of nucleotide divergence differed within these regions, suggesting that selection exerts constraints differentially on nucleotide change of theGpdh gene. A phylogenetic tree of thevirilis phylad constructed from nucleotide variation of total sequence was consistent with those obtained from other data.Nucleotide sequences for theGpdh gene ofD. lummei, D. novamexicana, D. a. americana, D. a. texana andD. ezoana have been submitted to GenBank with accession numbers D50087, D50088, D50089, D50090 and D50091.     

Nucleotide sequence of the histone gene cluster in the coral acropora formosa (cnidaria; scleractinia): Features of histone gene structure and organization are common to diploblastic and triploblastic metazoans

We report the nucleotide sequence of the core histone gene cluster from the Cnidarian Acropora formosa. This is the first histone gene cluster to be sequenced from a diploblastic organism and the predicted amino acid sequences most resemble those of sea urchin equivalents. Each of the Cnidarian histone genes has two conserved regions 3 of the coding sequences and these closely resemble those of the metazoan a-class histone genes. In A. formosa the core histone genes are arranged as opposed (H3/H4 and H2A/H2B) pairs, a pattern common to the nondeuterostome metazoa, and tandem repetition is the predominant pattern of organization in the Cnidarian. With the recent identification of several classes of homeobox genes in Cnidarians these features clearly align the Cnidaria with triploblastic metazoans, supporting a monophyletic origin of the metazoa.     

Tyrosine Decarboxylase and Dopa Decarboxylase in Drosophila virilis Under Normal Conditions and Heat Stress: Genetic and Physiological Aspects

The activity of tyrosine decarboxylase (TDC) and dopa decarboxylase (DDC) was studied in adults of two lines of Drosophila virilis,contrasting in their reaction to stress conditions. Differences were found in the activity of both enzymes between individuals of the examined lines. Genetic analysis of these differences was made. Each of the two enzymes was found to be controlled by a single gene or, possibly, by a block of closely linked genes. The gene responsible for TDC activity is located on one of the autosomes (excluding chromosome II). DDC activity in D. virilisis regulated by a gene located, apparently, on chromosome II. Adults of the line responding to stress by a stress reaction (r-line) were shown to react to a short-term heat stress (38°C, 60 min) by a decrease in TDC activity. TDC activity in flies of the line incapable of the stress reaction (nr-line) did not alter in such conditions. DDC activity of adults of both lines was found to be unchangeable under stress conditions.     

Organization of the histone H3 genes in soybean, barley and wheat

Several variants of the replacement histone H3 genes from soybean, barley and wheat have been cloned and sequenced. Analysis of segregating populations in barley and soybean, as well as analysis of clones isolated from a soybean genomic library, suggested that these genes are dispersed throughout the genome. Several genes contain introns located in similar positions, but of different lengths and sequence. Comparison of mRNA levels in different tissues revealed that the intron-containing and intronless genes have different expression patterns. The distribution of the introns in the histone H3 genes across several plant species suggests that some of the introns might have been lost during the evolution of the gene family. Sequence divergence among introns and gene-flanking sequences in cloned gene variants allowed us to use them as specific probes for localizing individual gene copies and analyzing the genomic distribution of these variants across a range of genotypes.Journal paper No. J-16127 of the Iowa Agriculture and Home Economics Experiment Station, Ames, IowaMention of a trademark or proprietary product does not constitute a guarantee or warranty of the product by the United States Department of Agriculture and does not imply its approval to the exclusion of other products that may be suitable     

Biochemical differences between cytoplasmic malate dehydrogenase allozymes of Drosophila virilis

Two allozymes (MDH^f and MDH^s) of cytoplasmic malate dehydrogenase of Drosophila virilis were partially purified and their biochemical properties were compared. MDH^f has a pH optimum of 9.75 and MDH^s one of 9.25 for malate oxidation. Optimal pH for oxaloacetate reduction is 6.75 and 8.0 for MDH^f and MDH^s, respectively. The K_m value for oxaloacetate of MDH^s is approximately twice as that of MDH^f. No differences were found with respect to thermostability and K_m's for malate, NAD⁺, or NADH. These results are discussed in terms of the physiological role of cytoplasmic malate dehydrogenase of D. virilis.This work was supported in part by grants from the Ministry of Education, Japan, Nos. 134050 and 154205.     

In vivo andin vitro methylation of lysine residues ofEuglena gracilis histone H1

We have earlier identified and purified two protein-lysine N-methyltransferases (Protein methylase III) fromEuglena gracilis [J. Biol. Chem.,260, 7114 (1985)]. The enzymes were highly specific toward histone H1 (lysine-rich), and the enzymatic products were identified as -N-mono-, di- and trimethyllysines. These earlier studies, however, were carried out with rat liver histone H1 as thein vitro substrate. Presently, histone H1 has been purified fromEuglena gracilis through Bio-Rex 70 and Bio-Gel P-100 column chromatography. TheEuglena histone H1 showed a single band on SDS-polyacrylamide gel electrophoresis and behaved like other histone H1 of higher animals, whereas it had a much higherR _f value than the other histones H1 in acid/urea gel electrophoresis. When theEuglena histone H1 was [methyl-³H]-labeledin vitro by a homologous enzyme (one of the twoEuglena protein methylase III) and analyzed on two-dimensional gel electrophoresis, three distinctive subtypes of histone H1 were shown to be radiolabeled, whereas five subtypes of rat liver histone H1 were found to be labeled. Finally, by the combined use of a strong cation exchange and reversed-phase Resolve C₁₈ columns on HPLC, we demonstrated thatEuglena histone H1 contains approximately 9 mol% of -N-methyllysines (1.40, 1.66, and 5.62 mol% for -N-mono-, di- and trimethyllysines, respectively). This is the first demonstration of the natural occurrence of -N-methyllysines in histone H1.     

Polymorphism and stability of histone gene clusters in Drosophila melanogaster cultured cells

In a previous communication (Saigo, K., Millstein, L. and Thomas, C.A., Jr. (1981) Cold Spring Harbor Symp. Quant. Biol. 45, 815–827), the overall structure of histone genes of Schneider line 2 cells was shown to extensively differ from that of Oregon-R embryo from which the cell line was established, and it was speculated that the histone genes might be reshuffled extensively during either the periods of the establishment, or maintenance of cell lines, or both. To establish the validity of this notion the structure of histone genes was examined in Drosophila melanogaster cultured cells. The overall organization of histone gene clusters was found to be stably maintained in both the periods for the establishment and maintenance of cultured cells, indicating that the previous assumption is inadequate. Instead of an extensive rearrangement, minor structural changes were found to occasionally occur probably by simple base substitutions and/or, deletion or insertion of very short DNA pieces. It was also shown that the extensive variation in structures of histone genes in cultured cells such as Schneider line 2 are attributable to polymorphism on the level of individual flies.     

Effects of dopamine on juvenile hormone metabolism and fitness in Drosophila virilis

The effects of dopamine (DA) on juvenile hormone (JH) metabolism and fitness (estimated as fecundity and viability levels under heat stress (38 °C)) in Drosophila virilis have been studied. An increase of DA level obtained by feeding with DA reduced fitness of wild-type (wt) flies under stress, and decreased JH degradation in young wt females while increasing it in sexually mature wt females. A decrease in DA levels resulted from 3-iodo-tyrosine treatment and caused a decrease in JH degradation in sexually mature wt and heat sensitive (hs) mutant females (DA level in hs females is twice as high in wt females). A dramatic decrease in viability under stress and fecundity under normal conditions in wt, but not hs, females was observed. 3-iodo-tyrosine treatment also reduced the number of oocytes at stages 8-14, delayed oocyte transition to stage 10 and resulted in the accumulation of mature eggs in wt females. It delayed maturation of wt, but not hs, males as well, but did not affect their fertility. This advances our understanding of the regulation of JH metabolism by DA in Drosophila and suggests a crucial role for the basal DA level in fitness.     

Drosophila mitochondrial DNA: Conserved sequences in the A+T-rich region and supporting evidence for a secondary structure model of the small ribosomal RNA

Summary The sequence of a segment of theDrosophila virilis mitochondrial DNA (mtDNA) molecule that contains the A+T-rich region, the small rRNA gene, the tRNA^f-met, tRNA^gln, and tRNA^ile genes, and portions of the ND2 and tRNA^val genes is presented and compared with the corresponding segment of theD. yakuba mtDNA molecule. The A+T-rich regions ofD. virilis andD. yakuba contain two correspondingly located sequences of 49 and 276/274 nucleotides that appear to have been conserved during evolution. In each species the replication origin of the mtDNA molecule is calculated to lie within a region that overlaps the larger conserved sequence, and within this overlap is found a potential hairpin structure. Substitutions between the larger conserved sequences of the A+T-rich regions, the small mt-rRNA genes, and the ND2 genes are biased in favor of transversions, 71–97% of which are AT changes. There is a 13.8 times higher frequency of nucleotide differences between the 5 halves than between the 3 halves of theD. virilis andD. yakuba small mt-rRNA genes. Considerations of the effects of observed substitutions and deletion/insertions on possible nucleotide pairing within the small mt-rRNA genes ofD. virilis andD. yakuba strongly support the secondary structure model for theDrosophila small mt-rRNA that we previously proposed.     

Human histone acetyltransferase 1 protein preferentially acetylates H4 histone molecules in H3.1-H4 over H3.3-H4

In mammalian cells, canonical histone H3 (H3.1) and H3 variant (H3.3) differ by five amino acids and are assembled, along with histone H4, into nucleosomes via distinct nucleosome assembly pathways. H3.1-H4 molecules are assembled by histone chaperone CAF-1 in a replication-coupled process, whereas H3.3-H4 are assembled via HIRA in a replication-independent pathway. Newly synthesized histone H4 is acetylated at lysine 5 and 12 (H4K5,12) by histone acetyltransferase 1 (HAT1). However, it remains unclear whether HAT1 and H4K5,12ac differentially regulate these two nucleosome assembly processes. Here, we show that HAT1 binds and acetylates H4 in H3.1-H4 molecules preferentially over H4 in H3.3-H4. Depletion of Hat1, the catalytic subunit of HAT1 complex, results in reduced H3.1 occupancy at H3.1-enriched genes and reduced association of Importin 4 with H3.1, but not H3.3. Finally, depletion of Hat1 or CAF-1p150 leads to changes in expression of a H3.1-enriched gene. These results indicate that HAT1 differentially impacts nucleosome assembly of H3.1-H4 and H3.3-H4.     

Mytilus edulis Histone Gene Clusters Containing Only H1 Genes

We isolated five different phage clones containing histone gene clusters with up to five H1 genes per phage clone from a Mytilus edulis genomic library. Among these H1 genes, nine gene types coding for five different H1 proteins have been identified. All H1 histone genes were located on repetitive restriction fragments with only slightly different sizes. The H1 coding regions show highly related sequences, suggesting that the multitude of H1 genes has evolved by gene duplication events. Core histone genes could not be found on these five Mytilus edulis genome fragments. Received: 28 July 1998 / Accepted: 17 May 1999     

Male diploid embryonic cell line ofDrosophila virilis

Summay A new established cell line 79f7Dv3g, ofDrosophila virilis consisting initially of male and female cells and represented now, after 6 yr of cultivation, only by male cells is described. The population doubling time is 36 h at 25° C. The cell culture is also able to grow in serum-free media for an indefinite time without special selection and has a population doubling time of 2 d.