Ethological-trophic groups of bivalve mollusks and their distribution in the Phanerozoic

Based on their adaptations to substrates, degree of mobility, and feeding modes, bivalve mollusks are subdivided into 30 ethological-trophic groups. This paper presents an account of their morphological characteristics, distribution in the Phanerozoic, and taxonomic changes through time.     

Oocyte-specific linker histone H1foo is an epigenomic modulator that decondenses chromatin and impairs pluripotency

Mammalian oocytes contain the histone H1foo, a distinct member with low sequence similarity to other members in the H1 histone family. Oocyte-specific H1foo exists until the second embryonic cell stage. H1foo is essential for oocyte maturation in mice; however, the molecular function of this H1 subtype is unclear. To explore the function of H1foo, we generated embryonic stem (ES) cells ectopically expressing H1foo fused to an EGFP (H1foo-ES). Interestingly, ectopic expression of H1foo prevented normal differentiation into embryoid bodies (EBs). The EB preparations from H1foo-ES cells maintained the expression of pluripotent marker genes, including Nanog, Myc and Klf9, and prevented the shift of the DNA methylation profile. Because the short hairpin RNA-mediated knockdown of H1foo-EGFP recovered the differentiation ability, H1foo was involved in preventing differentiation. Furthermore, ChIP analysis revealed that H1foo-EGFP bound selectively to a set of hypomethylated genomic loci in H1foo-ES, clearly indicating that these loci were targets of H1foo. Finally, nuclease sensitivity assay suggested that H1foo made these target loci decondensed. We concluded that H1foo has an impact on the genome-wide, locus-specific epigenetic status.     

Oocyte-specific linker histone H1foo is an epigenomic modulator that decondenses chromatin and impairs pluripotency

Mammalian oocytes contain the histone H1foo, a distinct member with low sequence similarity to other members in the H1 histone family. Oocyte-specific H1foo exists until the second embryonic cell stage. H1foo is essential for oocyte maturation in mice; however, the molecular function of this H1 subtype is unclear. To explore the function of H1foo, we generated embryonic stem (ES) cells ectopically expressing H1foo fused to an EGFP (H1foo-ES). Interestingly, ectopic expression of H1foo prevented normal differentiation into embryoid bodies (EBs). The EB preparations from H1foo-ES cells maintained the expression of pluripotent marker genes, including Nanog, Myc and Klf9, and prevented the shift of the DNA methylation profile. Because the short hairpin RNA-mediated knockdown of H1foo-EGFP recovered the differentiation ability, H1foo was involved in preventing differentiation. Furthermore, ChIP analysis revealed that H1foo-EGFP bound selectively to a set of hypomethylated genomic loci in H1foo-ES, clearly indicating that these loci were targets of H1foo. Finally, nuclease sensitivity assay suggested that H1foo made these target loci decondensed. We concluded that H1foo has an impact on the genome-wide, locus-specific epigenetic status.     

Coexistence of two chromatin structures in sperm nuclei of the bivalve molluscProtothaca thaca

The chromatin of the spermatozoa from the bivalve molluscProtothaca thaca, has a peculiar composition in which coexist core histones with sperm-specific proteins H1 and Pt1, the latter being a protein exhibiting features intermediate between histones and protamines. In this paper, we report an analysis of chromatin organization using micrococcal nuclease digestion, salt fractionation of soluble chromatin derived from nuclease digestion and crosslinking experiments. The results obtained indicate that it is possible to obtain two types of chromatin, one which is soluble, more accessible to micrococcal nuclease action and which does not contain Pt1, and another insoluble type, more resistant to micrococcal nuclease and enriched in protein Pt1. The crosslinking experiments show that the protein Pt1 interacts with itself and with core histones but not with sperm-specific H1. These results have led us to propose a special structural arrangement for this chromatin. Based in the data reported here we propose the coexstence in the genome ofP. thaca of two interspersed chromatin domains, one nucleosomal and the other nonnucleosomal containing protein Pt1.     

Nucleosomal organization of chromatin in sperm nuclei of the bivalve mollusc Aulacomya ater

The sperm nuclei of Aulacomya ater, family Mitylidae, contain three proteins (X, Aa5 and Aa6) which are specific to this cell type coexisting with a set of five somatic-type histones. Information about the chromatin structure resulting from this kind of association is scarce. Therefore, we have probed the structure of this sperm chromatin through digestion with micrococcal nuclease in combination with salt fractionation. The data obtained have allowed us to propose a nucleosomal arrangement for this chromatin. However, two types of nucleosomes would be present in agreement with their protein components.     

An H1-like protein from the sperm chromatin of Mytilus galloprovincialis

We have isolated and purified a sperm-specific protein (S3) from the mussel M. galloprovincialis. Antibodies against S3 were raised in rabbits and used for its immunological comparison to somatic histones. The results showed that S3 did not share common immunological determinants with H2b or any other core histone-contrary to the suggestion that it was an H2b-like protein (Ausio and Subirana, 1982). With H1 there was a crossreaction between S3 and anti-H1 as well as with H1 and anti-S3. Although similar to somatic H1, S3 is not identical with it. This fact makes S3 an interesting example of another protein of the H1-H5 type, present in a completely inactive chromatin.     

Molecular characterization and expression of a tobacco histone H1 cDNA

We have isolated a 1104 bp tobacco cDNA clone (H1c12) which includes an 846 bp open reading frame. This encodes a polypeptide of 282 amino acid residues and represents the largest plant H1 histone identified so far. The structure of the deduced protein shows the classical tripartite organization of the H1-type linker histones. The expression of the tobacco H1 histone gene(s) corresponding to the H1c12 cDNA clone was examined during different developmental stages. We found that, at the level of steadystate mRNA, expression of gene(s) encoding this H1 histone was rapidly induced in germinating seeds. The H1 gene was expressed in all tissues examined. However, its expression was higher in tissues known to contain meristematic cells. Furthermore, in the leaves of mature plants accumulation of the H1 mRNA exhibits a very characteristic oscillation. This latter finding indicates that, at least in fully developed plants, the expression of this type of H1 histone gene(s) is modulated by a diurnal cycle.     

Possible role of histone H1 in the regulation of furin-dependent proprotein processing

Histone H1 and its C-terminal lysine rich fragments were recently found to be potent inhibitorsof furin,a mammalian proprotein convertase.However,its role in the regulation of furin-dependent proproteinprocessing remains unclear.Here we report that histone H1 efficiently blocks furin-dependent pro-yonWillebrand factor(pro-vWF)processing in a dose-dependent manner.Coimmunoprecipitation and immunof-luorescence studies confirmed that histone H1 could interact with furin,and the interaction mainly took placeon the cell surface.We noted that histone H1 was released from cells undergoing necrosis and apoptosisinduced by H_2O_2.Our findings suggested that histone H1 might be involved in extracellular and/or intracellu-lar furin regulation.     

Distinct effects of histone H1 and non-histone chromosomal proteins on the structure of nucleosomes and the chromatin fibre in solution

In this study we attempt to differentiate between the effects of the non-histone chromosomal proteins and histone H1 on the structure of the nucleosomes and the chromatin fibre in solution. The properties of chromatin preparations with different histone H1 and non-histone protein compositions were compared using circular dichroism and flow linear dichroism and the following conclusions were drawn. When histone H1 is absent the non-histone proteins partially prevent the unfolding of the nucleosomes at low ionic strength. The complete blocking of this unfolding, however, is accomplished only in the presence of histone H1. The non-histone proteins do not affect the orientation of the nucleosomes along the fibre axis. Only histone H1 can maintain the positive anisotropy of the chromatin fibre.     

Structure of transfection-active histone H1/DNA complexes

Relationships between the structure of transfecting complexes of histone H1 and DNA and their transfection efficiency were studied. Transfection activity proved to be connected to complex aggregates. Low speed centrifugation of the complexes resulted in loss of the transfection activity. The complexes/aggregates were active with high efficiency in a broad range of weight input ratios r _i (0.1<r _i<30). Using atomic force microscopy (AFM), the complexes were imaged at negative, nearly electroneutral and positive charge conditions. Electroneutral complexes at r _i=1 showed a multitude of different complex forms. Fibrillar, network-like and branched structures were frequently present in one complex. Strongly positive charged complexes had a toroidal appearance. All these different forms contributed to the high transfection efficiency. Cellular uptake is supposed to be by phagocytosis.     

Unambiguous identification of histone H1 in Trypanosoma cruzi

The existence of histone H₁ has been questioned in Trypanosomatids. We report here the presence of a histone H₁ in the chromatin of Trypanosoma cruzi. This protein was purified by narrow-bore reversed phase HPLC and its amino acid composition analyzed and compared with histones H₁ from other species. Furthermore, the purified chromosomal protein was digested with proteases and the amino acid sequences of the resulting peptides were analyzed by the automated Edman degradation. The sequences obtained were found to present a high degree of homology when compared to the carboxy terminal domain of other known histones H₁.     

Sequence specific binding of chlamydial histone H1-like protein.

Chlamydia trachomatis is one of the few prokaryotic organisms known to contain proteins that bear homology to eukaryotic histone H1. Changes in macromolecular conformation of DNA mediated by the histone H1-like protein (Hc1) appear to regulate stage specific differentiation. We have developed a cross-linking immunoprecipitation protocol to examine in vivo protein-DNA interaction by immune precipitating chlamydial Hc1 cross linked to DNA. Our results strongly support the presence of sequence specific binding sites on the chlamydial plasmid and hc1 gene upstream of its open reading frame. The preferential binding sites were mapped to 520 bp BamHI-XhoI and 547 bp BamHI-DraI DNA fragments on the plasmid and hc1 respectively. Comparison of these two DNA sequences using Bestfit program has identified a 24 bp region with >75% identity that is unique to the chlamydial genome. Double-stranded DNA prepared by annealing complementary oligonucleotides corresponding to the conserved 24 bp region bind Hc1, in contrast to control sequences with similar A+T ratios. Further, Hc1 binds to DNA in a strand specific fashion, with preferential binding for only one strand. The site specific affinity to plasmid DNA was also demonstrated by atomic force microscopy data images. Binding was always followed by coiling, shrinking and aggregation of the affected DNA. Very low protein-DNA ratio was required if incubations were carried out in solution. However, if DNA was partially immobilized on mica substrate individual strands with dark foci were still visible even after the addition of excess Hc1.     

Phosphorylation of histone H1 during the cell cycle of artichoke

Abstract. Histones have been extracted from tuber and cultured tuber explant material and separated by gel electrophoresis. Histone H1 is heterogenous with 3–4 components in addition to the widely recognized histone H1a and H1b. Using labelling procedures and alkaline phosphatase treatment, histone H1 has been shown to be phosphorylated on both serine and threonine residues and possibly other acid-labile linkages. Variations in histone H1 phosphorylation have been measured through the cell cycle and the evidence indicates enhanced phosphorylation occurring during the G2/M phase as in animal systems.     

Structure and expression of the human oocyte-specific histone H1 gene elucidated by direct RT-nested PCR of a single oocyte

Oocyte-specific histone H1 is expressed during oogenesis and early embryogenesis. It has been described in mice and some nonmammalian species, but not in humans. Here, we identified the cDNA in unfertilized human oocytes using direct RT-nested PCR of a single cell. Sequencing of this cDNA indicated an open reading frame encoding a 347-amino acid protein. Expression was oocyte-specific. Homology was closest with the corresponding gene of mouse (H1oo; 42.3%), and, to lesser extent, with that of Xenopus laevis (B4; 25.0%). The gene, named osH1, included five exons as predicted by the NCBI annotation project of the human genome, although the actual splicing site at the 3(') end of exon 3 was different by 48 nucleotides from the prediction. The presence of polyadenylation signals and successful amplification of cDNA by RT-PCR using an oligo(dT) primer suggested that the osH1 mRNA is polyadenylated unlike somatic H1 mRNA. Our technique and findings should facilitate investigation of human fertilization and embryogenesis.     

The H1 histone variant of tomato, H1-S, is targeted to the nucleus and accumulates in chromatin in response to water-deficit stress

 Water deficit has a significant impact on patterns of gene expression. Based on the deduced amino acid sequence, it has been proposed that the drought and abscisic acid-induced gene (his1-s) of tomato (Lycopersicon esculentum Mill.) encodes an H1 histone variant. To study the role of H1-S it is important to understand the expression characteristics of the protein. To identify the his1-s product in vivo the his1-s cDNA was fused to a (His)₆ tag and overexpressed in Escherichiacoli. The H1-S fusion protein was used to generate an antibody that recognized a protein with an apparent molecular weight of 31 kDa that accumulates in response to water deficit in the whole plant and detached leaves. A time course of his1-s expression showed that protein accumulation is delayed compared to the mRNA accumulation in both the whole plant and detached leaves. Cellular fractionation, immunofluorescence and H1-S::β-glucuronidase fusion analyses in transgenic tissues were used to determine the cellular localization of H1-S. The results showed that H1-S accumulates in nuclei and is associated with chromatin of wilted tomato leaves. The drought- and abscisic acid-induced gene his1-s encodes a linker-histone subtype specifically accumulated in the nuclei and chromatin of tomato leaves subjected to water-deficit conditions. Although the molecular mechanism of H1-S function is still unclear, the expression characteristics of H1-S are consistent with a potential role of this protein in the regulation of gene expression in response to water deficit. Received: 1 October 1999 / Accepted: 3 December 1999