The description of genetic alterations in tumours is of increasing importance. In human genetics, and in pathology reports, sequence alterations are given using the human genome variation society (HGVS) guidelines for the description of such variants. However, there is less adherence to these guidelines for sequence variations in histone genes. Due to early cleavage of the N-terminal methionine in most histones, the description of histone sequence alterations follows their own nomenclature and differs from the HGVS-compliant numbering by omitting this first amino acid. Next generation sequencing reports, however, follow the HGVS guidelines and as a result, an unambiguous description of sequence variants in histones cannot be provided. The coexistence of these two nomenclatures leads to confusions for pathologists, oncologists, and researchers. This review provides an overview of tumour entities with sequence alterations of the H3-3A gene (HGNC ID = HGNC:4764), highlights the problems associated with the coexistence of these two nomenclatures, and proposes a standard for the reporting of histone sequence variants that allows an unambiguous description of these variants according to HGVS principles. We hope that scientific journals will adopt the new notation, and that both geneticists and pathologists will include it in their reports. © 2021 The Authors. The Journal of Pathology published by John Wiley & Sons, Ltd. on behalf of The Pathological Society of Great Britain and Ireland.

Consensus-based protein engineering strategy has been applied to various proteins and it can lead to the design of proteins with enhanced biological performance. Histone-like HUs comprise a protein family with sequence variety within a highly conserved 3D-fold. HU function includes compacting and regulating bacterial DNA in a wide range of biological conditions in bacteria. To explore the possible impact of consensus-based design in the thermodynamic stability of HU proteins, the approach was applied using a dataset of sequences derived from a group of 40 mesostable, thermostable, and hyperthermostable HUs. The consensus-derived HU protein was named HUBest, since it is expected to perform best. The synthetic HU gene was overexpressed in E. coli and the recombinant protein was purified. Subsequently, HUBest was characterized concerning its correct folding and thermodynamic stability, as well as its ability to interact with plasmid DNA. A substantial increase in HUBest stability at high temperatures is observed. HUBest has significantly improved biological performance at ambience temperature, presenting very low Kd values for binding plasmid DNA as indicated from the Gibbs energy profile of HUBest. This Kd may be associated to conformational changes leading to decreased thermodynamic stability and, therefore, higher flexibility at ambient temperature.

The nervous system may include more than 100 residue-specific posttranslational modifications of histones forming the nucleosome core that are often regulated in cell-type-specific manner. On a genome-wide scale, some of the histone posttranslational modification landscapes show significant overlap with the genetic risk architecture for several psychiatric disorders, fueling PsychENCODE and other large-scale efforts to comprehensively map neuronal and nonneuronal epigenomes in hundreds of specimens. However, practical guidelines for efficient generation of histone chromatin immunoprecipitation followed by deep sequencing (ChIP-seq) datasets from postmortem brains are needed.
Protocols and quality controls are given for the following: 1) extraction, purification, and NeuN neuronal marker immunotagging of nuclei from adult human cerebral cortex; 2) fluorescence-activated nuclei sorting; 3) preparation of chromatin by micrococcal nuclease digest; 4) ChIP for open chromatin-associated histone methylation and acetylation; and 5) generation and sequencing of ChIP-seq libraries.
We present a ChIP-seq pipeline for epigenome mapping in the neuronal and nonneuronal nuclei from the postmortem brain. This includes a stepwise system of quality controls and user-friendly data presentation platforms.
Our practical guidelines will be useful for projects aimed at histone posttranslational modification mapping in chromatin extracted from hundreds of postmortem brain samples in cell-type-specific manner.

Histone post-translational modifications (PTMs) have pivotal functions in many chromatin processes, which makes their detection and characterization an imperative in chromatin biology. The established approaches for histone PTM characterization are generally based on affinity reagents specific for modified histone tails such as antibodies and, most recently, recombinant reading domains. Hence, the proper performance of these reagents is a critical precondition for the validity of the generated experimental data. In this review, we evaluate and update the quality criteria for assessment of the binding specificity of histone PTM affinity reagents. In addition, we discuss in detail the advantages and pitfalls of using antibodies and recombinant reading domains in chromatin biology research. Reading domains provide key advantages, such as consistent quality and recombinant production, but the future will tell if this emerging technology keeps its promises.

Chromatin immunoprecipitation (ChIP) followed by high-throughput DNA sequencing (ChIP-seq) has become a valuable and widely used approach for mapping the genomic location of transcription-factor binding and histone modifications in living cells. Despite its widespread use, there are considerable differences in how these experiments are conducted, how the results are scored and evaluated for quality, and how the data and metadata are archived for public use. These practices affect the quality and utility of any global ChIP experiment. Through our experience in performing ChIP-seq experiments, the ENCODE and modENCODE consortia have developed a set of working standards and guidelines for ChIP experiments that are updated routinely. The current guidelines address antibody validation, experimental replication, sequencing depth, data and metadata reporting, and data quality assessment. We discuss how ChIP quality, assessed in these ways, affects different uses of ChIP-seq data. All data sets used in the analysis have been deposited for public viewing and downloading at the ENCODE (http://encodeproject.org/ENCODE/) and modENCODE (http://www.modencode.org/) portals.

The han1A gene, encoding a subunit of the histone-like protein HAN1 from the Thermococcus species AN1, has been cloned and sequenced. Sequence analysis of the translation product of the gene demonstrates homology with other archaeal histone-like proteins of the \'HMf family\' and eukaryal consensus sequences, particularly H4. The region of highest homology between the AN1 histone subunit, termed the HAN1A1 subunit, and the H4 consensus is suggested, by the 3-dimensional structure of the histone octamer, to interact with the minor groove of DNA. The results presented add further weight to the notion that the \'archaeal histones\' and the eukaryal histones are indeed related and that the approximate 65 amino acid residue length of the archaeal histones represents the archaeal equivalent of the histone fold structural building block common to all eukaryal histones.

32P-Labeled histone H1 was isolated from synchronized Chinese hamster (line CHO) cells, subjected to tryptic digestion, and fractionated into 15 phosphopeptides by high performance liquid chromatography. These phosphopeptides were grouped into five classes having different cell cycle phosphorylation kinetics: 1) peptides reaching a maximum phosphorylation rate in S and then declining in G2 and M, 2) peptides reaching a maximum phosphorylation rate in G2 and then remaining constant or declining in M, 3) peptides with increasing phosphorylation throughout S and G2 and reaching a maximum in M, 4) one peptide that was phosphorylated only in M, and 5) peptides that had low levels of phosphorylation that remained constant throughout the cell cycle. Amino acid analysis and sequencing demonstrated that the mitotic specific H1 phosphopeptide was the 16-amino acid, N-terminal, tryptic peptide Ac-SETAPAAPAAAPPAEK of the H1-1 class. This peptide, which is phosphorylated on both the Ser and Thr, does not contain the consensus sequence (S/T)PXZ (where X is any amino acid and Z is a basic amino acid). This sequence is thought to be required by the p34cdc2/cyclin B kinase that has maximum phosphorylating activity in mitosis. These data indicate that this kinase either does not have an obligatory requirement for the consensus sequence in vivo as generally believed or that it is not the enzyme responsible for the mitotic specific H1 phosphorylation.

We have previously identified an autonomously replicating segment (ARS) near the 3\' end of the histone H4 gene at the copy-I H3-H4 locus. We have now searched for additional autonomously replicating segments and sequences homologous with the ARS core consensus sequence near the copy-II histone H4 gene and both of the histone H3 genes. No new ARS elements were identified by functional cloning assays. However, several matches to the ARS core consensus element were found within the DNA sequences of the copy-I and copy-II genes. An exact match to the ARS core consensus was identified in the region downstream from the copy-I histone H3 gene and a set of sequences with weak homology was also located within the copy-II region. However, restriction fragments including these sequences did not demonstrate ARS activity on a plasmid in transformed cells.

Mammalian histone gene transcription is increased approximately fivefold during the transition from the G1 phase to the S phase of the cell cycle. In this study, we present a detailed in vivo analysis of the human histone H2b promoter, which establishes that transcriptional regulation of this gene is mediated by a subtype-specific consensus element containing the core octanucleotide ATTTGCAT. Our results demonstrate that the activity of this sequence is specific for S phase. Comparative analysis of different replication variant mammalian histone gene promoters and our knowledge of the transcription factors interacting with the human histone H2b and H4 promoters allow us to conclude that coordinate regulation of histone gene transcription in higher eukaryotes is mediated by distinct factors. We propose a simple model for transcriptional regulation of mammalian histone gene expression, which incorporates both the distinct features of the individual histone gene promoters and the apparent functional equivalence of the specific sequence elements regulating transcription of each histone gene subtype.

Definition of mechanisms regulating human histone H1 gene transcription during the cell cycle requires the isolation and biochemical characterization of protein factors which interact with specific promoter elements. Two distinct binding activities have been identified in nuclear extracts from HeLa cells and mapped within a 180-base-pair (bp) region of a cell cycle-regulated H1 gene promoter. H1TF1 bound to an H1-specific A + C-rich sequence (AC box), 100 bp upstream of the cap site; H1TF2 interacted with the H1 subtype-specific consensus element and was dependent on the presence of an intact CCAAT box for binding. H1TF2 was purified through a combination of ion-exchange and oligonucleotide affinity chromatographies. Analysis of purified fractions by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and UV crosslinking showed that H1TF2 was a single polypeptide of 47 kilodaltons. This factor was distinct from previously characterized CCAAT-binding proteins in both molecular size and binding properties. Fractions containing H1TF2 activity activated transcription in vitro only if programmed with an H1 DNA template carrying an intact H1TF2-binding site.