DNA- and chromatin-condensing properties of rat testes H1a and H1t compared to those of rat liver H1bdec: H1t is a poor condenser of chromatin

Histones H1a and H1t are two major linker histone variants present at the pachytene interval of mammalian spermatogenesis. The DNA- and chromatin-condensing properties of these two variants isolated from rat testes were studied and compared with those from rat liver. For this purpose, the histone H1 subtypes were purified from the respective tissues using bath acid and salt extraction procedures, Circular dichroism studies revealed that acid exposure during isolation affects the alpha-helical structure of both the globular domain (in the presence of 1 M NaCl) and the C-terminal lambda-tail (in the presence of 60% trifluoroethanol). The condensation of rat oligonucleosomal DNA, as measured by circular dichroism spectroscopy, by the salt-extracted histone H1 was at least 10 times more efficient than condensation by the acid-extracted histone H1. A site size of 16-20 base pairs was calculated for the salt-extracted histone H1. Among the different histone H1 subtypes, somatic histone H1bdec had the highest DNA-condensing property, followed by histone H1a and histone H1t. All the salt-extracted histones condensed rat oligonucleosomal DNA more efficiently than linear pBR-322 DNA, Histones H1bdec and H1a condensed histone H1-depleted chromatin, prepared from rat liver nuclei, with relatively equal efficiency. On the other hand, there was no condensation of histone H1-depleted chromatin with the testes specific histone H1t. A comparison of the amino acid sequences of histone H1d (rat) and histone H1t (rat) revealed several interesting differences in the occurrence of DNA-binding motifs at the C-terminus. A striking observation is the presence of a direct repeat of an octapeptide motif K(A)T(S)PKKA(S)K(T)K(A) in histone H1d that is absent in histone H1t

Condensation of DNA and chromatin by an SPKK-containing octapeptide repeat motif present in the c-terminus of histone H1

Several DNA binding motifs have been described in the C-terminus of histone H1 (Churchill & Travers, 1991), of these the S/TPKK repeat (Suzuki, 1989) often occurs as a part of an octapeptide repeat of the type XTPKKXKK. We have studied in detail the DNA and chromatin condensing properties of a consensus octapeptide KSPKKAKK (8 mer) present in many histone H1 subtypes and its imperfect repeat ATPKKSTKKTPKKAKK (16 mer TPKK) as it occurs in the C-terminus of rat histone H1d. The 16 mer TPKK peptide containing two S/TPKK motifs was able to condense both rat oligonucleosomal (2-5 kbp) DNA and histone H1-depleted chromatin as revealed by circular dichroism spectroscopy. The 8 mer peptide, however, was unable to condense either the DNA or the histone H1-depleted chromatin. Both the 8 mer peptide and the 16 mer TPKK peptide displaced distamycin A from the drug-DNA complex, although with different efficiency, indicating that while these two peptides could bind DNA, only the 16 mer (TPKK) peptide could bring about condensation of DNA and histone H1-depleted chromatin. A mutant 16 mer (TAKK) peptide wherein two proline residues are replaced by alanine, was ineffective in bringing about condensation of both DNA and histone H1-depleted chromatin. These results suggest that the two β-turn structures present in the 16 mer (TPKK) peptide could be important in facilitating binding to different regions of duplex DNA thereby bringing about close packing and condensation. The condensation property of the 16 mer (TPKK) peptide was very similar to that of histone H1 in terms of (a) its preference for AT rich DNA, (b) cooperativity of condensation, and (c) salt dependence of condensation. The 16 mer (TPKK) peptide, but not the 8 mer peptide or the 16 mer (TAKK) peptide, could form complexes with a polynucleosomal 5S DNA core resulting in retarded mobility similar to the complexes formed with histone H1 on agarose gel electrophoresis

Best practice data life cycle approaches for the life sciences

Throughout history, the life sciences have been revolutionised by technological advances; in our era this is manifested by advances in instrumentation for data generation, and consequently researchers now routinely handle large amounts of heterogeneous data in digital formats. The simultaneous transitions towards biology as a data science and towards a ‘life cycle’ view of research data pose new challenges. Researchers face a bewildering landscape of data management requirements, recommendations and regulations, without necessarily being able to access data management training or possessing a clear understanding of practical approaches that can assist in data management in their particular research domain. Here we provide an overview of best practice data life cycle approaches for researchers in the life sciences/bioinformatics space with a particular focus on ‘omics’ datasets and computer-based data processing and analysis. We discuss the different stages of the data life cycle and provide practical suggestions for useful tools and resources to improve data management practices

Low-Frequency and Rare-Coding Variation Contributes to Multiple Sclerosis Risk

Multiple sclerosis is a complex neurological disease, with ∼20% of risk heritability attributable to common genetic variants, including >230 identified by genome-wide association studies. Multiple strands of evidence suggest that much of the remaining heritability is also due to additive effects of common variants rather than epistasis between these variants or mutations exclusive to individual families. Here, we show in 68,379 cases and controls that up to 5% of this heritability is explained by low-frequency variation in gene coding sequence. We identify four novel genes driving MS risk independently of common-variant signals, highlighting key pathogenic roles for regulatory T cell homeostasis and regulation, IFNγ biology, and NFκB signaling. As low-frequency variants do not show substantial linkage disequilibrium with other variants, and as coding variants are more interpretable and experimentally tractable than non-coding variation, our discoveries constitute a rich resource for dissecting the pathobiology of MS.status: publishe

New gene functions in megakaryopoiesis and platelet formation

Platelets are the second most abundant cell type in blood and are essential for maintaining haemostasis. Their count and volume are tightly controlled within narrow physiological ranges, but there is only limited understanding of the molecular processes controlling both traits. Here we carried out a high-powered meta-analysis of genome-wide association studies (GWAS) in up to 66,867 individuals of European ancestry, followed by extensive biological and functional assessment. We identified 68 genomic loci reliably associated with platelet count and volume mapping to established and putative novel regulators of megakaryopoiesis and platelet formation. These genes show megakaryocyte-specific gene expression patterns and extensive network connectivity. Using gene silencing in Danio rerio and Drosophila melanogaster, we identified 11 of the genes as novel regulators of blood cell formation. Taken together, our findings advance understanding of novel gene functions controlling fate-determining events during megakaryopoiesis and platelet formation, providing a new example of successful translation of GWAS to function

Maps of open chromatin highlight cell type-restricted patterns of regulatory sequence variation at hematological trait loci

<p>Nearly three-quarters of the 143 genetic signals associated with platelet and erythrocyte phenotypes identified by meta-analyses of genome-wide association (GWA) studies are located at non-protein-coding regions. Here, we assessed the role of candidate regulatory variants associated with cell type-restricted, closely related hematological quantitative traits in biologically relevant hematopoietic cell types. We used formaldehyde-assisted isolation of regulatory elements followed by next-generation sequencing (FAIRE-seq) to map regions of open chromatin in three primary human blood cells of the myeloid lineage. In the precursors of platelets and erythrocytes, as well as in monocytes, we found that open chromatin signatures reflect the corresponding hematopoietic lineages of the studied cell types and associate with the cell type-specific gene expression patterns. Dependent on their signal strength, open chromatin regions showed correlation with promoter and enhancer histone marks, distance to the transcription start site, and ontology classes of nearby genes. Cell type-restricted regions of open chromatin were enriched in sequence variants associated with hematological indices. The majority (63.6%) of such candidate functional variants at platelet quantitative trait loci (QTLs) coincided with binding sites of five transcription factors key in regulating megakaryopoiesis. We experimentally tested 13 candidate regulatory variants at 10 platelet QTLs and found that 10 (76.9%) affected protein binding, suggesting that this is a frequent mechanism by which regulatory variants influence quantitative trait levels. Our findings demonstrate that combining large-scale GWA data with open chromatin profiles of relevant cell types can be a powerful means of dissecting the genetic architecture of closely related quantitative traits.</p>