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Whereas the genomes of many organisms contain several nonallelic types of linker histone genes, one single histone H1 type is known in Drosophila melanogaster that occurs in about 100 copies per genome. Amplification of H1 gene sequences from genomic DNA of wild type strains of D. melanogaster from Oregon, Australia, and central Africa yielded numerous clones that all exhibited restriction patterns identical to each other and to those of the known H1 gene sequence. Nucleotide sequences encoding the evolutionarily variable domains of H1 were determined in two gene copies of strain Niamey from central Africa and were found to be identical to the known H1 sequence. Most likely therefore, the translated sequences of D. melanogaster H1 genes do not exhibit intragenomic or intergenomic variations. In contrast, three different histone H1 genes were isolated from D. virilis and found to encode proteins that differ remarkably from each other and from the H1 of D. melanogaster and D. hydei. About 40 copies of H1 genes are organized in the D. virilis genome with copies of core histone genes in gene quintets that were found to be located in band 25F of chromosome 2. Another type of histone gene cluster is present in about 15 copies per genome and contains a variable intergenic sequence instead of an H1 gene. The H1 heterogeneity in D. virilis may have arisen from higher recombination rates than occur near the H1 locus in D. melanogaster and might provide a basis for formation of different chromatin subtypes.

1. Seven subfractions of histone H1 have been isolated and purified from larvae of Chironomus thummi (Diptera). They have been denominated 1-1, 11-1, 11-2, 11-3, 111-1, 111-2, and 111-3, according to the order of migration in two steps of preparative electrophoresis.2. The amino acid compositions are similar to those of other H1 histones. Subfractions 1-1 and 11-1 were found to contain one methionine and two tyrosine residues, 11-2 contained two methionine and three tyrosine residues, and 111-1 one methionine and three tyrosine residues. The other subfractions contained one or two methionine and two or three tyrosine residues. For subfractions 1-1 and 11-1 a chain length of about 252 amino acids was estimated.3. Peptide pattern analyses after chemical cleavage at the methionine and tyrosine residues, and enzymatic cleavage with thrombin and chymotrypsin, respectively, showed that all subfractions have different individual primary structures. A comparison of peptide sizes and of the positions in the peptide patterns of epitopes recognized by monoclonal antibodies was made to check whether some of the subfractions could arise by proteolytic degradation of others. This possibility can be excluded for five of the subfractions and is very improbable for the two others. Treatment of C. thummi H1 with alkaline phosphatase did not change the pattern of subfractions, while the phosphorylated subfraction of histone H2A disappeared after this treatment. Most and very probably all subfractions are thus H1 sequence variants.4. Inbred strains and individual larvae of C. thummi were found to comprise all seven' variants. The HI heterogeneity can therefore not be due to allelic polymorphism. Salivary gland nuclei were found to contain variant 1-1 and at least some of the other variants.5. H1 from Drosophilu melunogaster and from calf thymus were used as reference molecules in all cleavage experiments and yielded the peptide patterns expected from the sequence. The comparison discriminates the group of C. thummi H1 histones clearly from Drosophilu and calf thymus H1. Limited trypsin digestion yielded a protected peptide of uniform size in six of the seven variants which was considerably smaller than the protected central domain of calf thymus H1.6. Two other species of Chironomidae, C. pullidivittatus and Glyptotendipes barhipes were found to contain five and three H 1 subfractions, respectively.7. The results provide a basis for the localization of H1 variants in specific chromatin regions within the giant chromosomes by means of monoclonal antibodies and may lead to an understanding of the functional significance of H1 heterogeneity.Histone H1 binds to the nucleosome chain and mediates its folding into the 30-nm fibre of eukaryote chromatin [l]. The central globular domain of the molecule probably binds to a specific nucleosome site [2] and contains sequences which are largely conserved within a wide range of organisms [3]. The amino acid sequences of the N-and C-terminal domains of HI, on the other hand, are more variable an...

Chromatin of Chironomus thummi (Diptera) contains seven sequence variants of histone Hi. A structurally divergent Hi, variant I-1, accounts for about 20% of the total Hi in C. th. piger and for about 30% in C. th. thummi.Monoclonal antibodies against this protein have been induced and have revealed its restriction to the centromeres and to a limited number of other bands in the salivary gland chromosomes. Indirect inmunofluorescence of the somatically paired homologous chromosomes of F1 hybrids indicates that the difference between the two subspecies in Hi I-1 content largely depends on differences situated at a number of distinct homologous chromosome bands. These bands were intensely decorated by antibodies against Hi I-1 in C. th. thummi but appeared virtually black in C. th. piger. The same bands, however, were decorated equally in both subspecies by an antibody that reacts with other Hi variants but does not recognize Hi I-1 and by a polyclonal anti-Hi antibody. The results suggest that Hi variant I-1 is characteristic of a specific type of chromatin that is confined to distinct chromosome segments and that is more frequent in the subspecies C. th. thummi, which has a 27% larger genome.reported earlier, larvae of C. thummi contain seven different sequence variants of histone H1 (12-14). Salivary gland nuclei contain several different and possibly all seven H1 variants. Antibodies that recognize either a subset of the H1 complement or one single H1 variant have been induced and have been used in the present study to decorate larval salivary gland chromosomes. We find that a specific H1 variant, I-1, is abundant in a limited number of chromosome bands, while it is rare or virtually absent in the chromatin of other chromosome regions. A comparison of two subspecies, C. th. thummi and C. th. piger, which differ remarkably in their total content of H1 variant I-1, revealed that this difference is largely confined to differences between specific homologous chromosome bands. Many of the C. th. thummi chromosome regions with a high content of H1 I-1 differ from their homologous C. th. piger counterparts also in that they contain more and repetitive DNA (15,16), replicate late in S phase (17), and/or react positively in a C-banding staining procedure (18). It appears that a high percentage of H1 variant I-1 is characteristic of a specific type of chromatin.In the periodic structural element of chromatin, the nucleosome, DNA is associated with octameric protein complexes containing pairs of molecules of each of four types of histones. The folding ofthis chain of nucleosomes into the 30-nm fiber of eukaryote chromatin is mediated by the binding of another type ofhistone, H1 (1). Numerous organisms and cell types have been found to contain several different subtypes of H1 (for reviews, see refs. 2, 3), but the functional role of this heterogeneity is not known.In the course of spermiohistogenesis in mammals (4-6) and erythrocyte maturation (7-9), most of the nuclear H1 molecules are replaced by other members of...