Subtelomeric Chromatin in the Fission Yeast S. pombe

Telomeres play important roles in safeguarding the genome. The specialized repressive chromatin that assembles at telomeres and subtelomeric domains is key to this protective role. However, in many organisms, the repetitive nature of telomeric and subtelomeric sequences has hindered research efforts. The fission yeast S. pombe has provided an important model system for dissection of chromatin biology due to the relative ease of genetic manipulation and strong conservation of important regulatory proteins with higher eukaryotes. Telomeres and the telomere-binding shelterin complex are highly conserved with mammals, as is the assembly of constitutive heterochromatin at subtelomeres. In this review, we seek to summarize recent work detailing the assembly of distinct chromatin structures within subtelomeric domains in fission yeast. These include the heterochromatic SH subtelomeric domains, the telomere-associated sequences (TAS), and ST chromatin domains that assemble highly condensed chromatin clusters called knobs. Specifically, we review new insights into the sequence of subtelomeric domains, the distinct types of chromatin that assemble on these sequences and how histone H3 K36 modifications influence these chromatin structures. We address the interplay between the subdomains of chromatin structure and how subtelomeric chromatin is influenced by both the telomere-bound shelterin complexes and by euchromatic chromatin regulators internal to the subtelomeric domain. Finally, we demonstrate that telomere clustering, which is mediated via the condensed ST chromatin knob domains, does not depend on knob assembly within these domains but on Set2, which mediates H3K36 methylation.

More than causing (epi)genomic instability: emerging physiological implications of transposable element modulation

Transposable elements (TEs) initially attracted attention because they comprise a major portion of the genomic sequences in plants and animals. TEs may jump around the genome and disrupt both coding genes as well as regulatory sequences to cause disease. Host cells have therefore evolved various epigenetic and functional RNA-mediated mechanisms to mitigate the disruption of genomic integrity by TEs. TE associated sequences therefore acquire the tendencies of attracting various epigenetic modifiers to induce epigenetic alterations that may spread to the neighboring genes. In addition to posting threats for (epi)genome integrity, emerging evidence suggested the physiological importance of endogenous TEs either as cis-acting control elements for controlling gene regulation or as TE-containing functional transcripts that modulate the transcriptome of the host cells. Recent advances in long-reads sequence analysis technologies, bioinformatics and genetic editing tools have enabled the profiling, precise annotation and functional characterization of TEs despite their challenging repetitive nature. The importance of specific TEs in preimplantation embryonic development, germ cell differentiation and meiosis, cell fate determination and in driving species specific differences in mammals will be discussed.

iCAT: diagnostic assessment tool of immunological history using high-throughput T-cell receptor sequencing

The pathogen exposure history of an individual is recorded in their T-cell repertoire and can be accessed through the study of T-cell receptors (TCRs) if the tools to identify them were available. For each T-cell, the TCR loci undergoes genetic rearrangement that creates a unique DNA sequence. In theory these unique sequences can be used as biomarkers for tracking T-cell responses and cataloging immunological history. We developed the immune Cell Analysis Tool (iCAT), an R software package that analyzes TCR sequencing data from exposed (positive) and unexposed (negative) samples to identify TCR sequences statistically associated with positive samples. The presence and absence of associated sequences in samples trains a classifier to diagnose pathogen-specific exposure. We demonstrate the high accuracy of iCAT by testing on three TCR sequencing datasets. First, iCAT successfully diagnosed smallpox vaccinated versus naïve samples in an independent cohort of mice with 95% accuracy. Second, iCAT displayed 100% accuracy classifying naïve and monkeypox vaccinated mice. Finally, we demonstrate the use of iCAT on human samples before and after exposure to SARS-CoV-2, the virus behind the COVID-19 global pandemic. We were able to correctly classify the exposed samples with perfect accuracy. These experimental results show that iCAT capitalizes on the power of TCR sequencing to simplify infection diagnostics. iCAT provides the option of a graphical, user-friendly interface on top of usual R interface allowing it to reach a wider audience.

iCAT: diagnostic assessment tool of immunological history using high-throughput T-cell receptor sequencing

The pathogen exposure history of an individual is recorded in their T-cell repertoire and can be accessed through the study of T-cell receptors (TCRs) if the tools to identify them were available. For each T-cell, the TCR loci undergoes genetic rearrangement that creates a unique DNA sequence. In theory these unique sequences can be used as biomarkers for tracking T-cell responses and cataloging immunological history. We developed the immune Cell Analysis Tool (iCAT), an R software package that analyzes TCR sequencing data from exposed (positive) and unexposed (negative) samples to identify TCR sequences statistically associated with positive samples. The presence and absence of associated sequences in samples trains a classifier to diagnose pathogen-specific exposure. We demonstrate the high accuracy of iCAT by testing on three TCR sequencing datasets. First, iCAT successfully diagnosed smallpox vaccinated versus naïve samples in an independent cohort of mice with 95% accuracy. Second, iCAT displayed 100% accuracy classifying naïve and monkeypox vaccinated mice. Finally, we demonstrate the use of iCAT on human samples before and after exposure to SARS-CoV-2, the virus behind the COVID-19 global pandemic. We were able to correctly classify the exposed samples with perfect accuracy. These experimental results show that iCAT capitalizes on the power of TCR sequencing to simplify infection diagnostics. iCAT provides the option of a graphical, user-friendly interface on top of usual R interface allowing it to reach a wider audience.

Coronavirus-associated molecular mimicry through homology to a SARS-CoV-2 peptide could be leading to susceptibility in patients with HLA-A*02:01 and HLA-A*24:02 serotypes

AimThis study aims to predict autoimmunity-related pathological mechanisms that posses risk for individuals with certain HLA serotypes and are common to the pathogenicity of certain coronaviruses including SARS-CoV-2, based on homology to a SARS-CoV-2 peptide.MethodsCoronavirus-associated sequences, which are homologous to the SARS-CoV-2 peptide CFLGYFCTCYFGLFC, are obtained. Human peptides that have at least 7 residue matches with those coronavirus sequences, and the SARS-CoV-2 peptide, are obtained. Then, epitope pairs, which are sourced by those coronavirus and human sequences’ alignments, are identified. There, epitope pairs that are predicted to bind strongly not only to the same HLA allele with each other but also to the same HLA allele with the respective alignment of the SARS-CoV-2 peptide are selected.ResultsFollowing is the list of proteins (or regions) with predicted HLA-A*02:01 or HLA-A*24:02 epitopes, which are not only common but also homologous to the epitopes that are sourced by the SARS-CoV-2 peptide and its homologous coronavirus peptides: Immunoglobulin heavy chain junction regions, CRB1 isoform I precursor, slit homolog 2 protein, hCG1995581, hCG2028737, phospholipid phosphatase-related protein type 2. Among those, CRB1 isoform I precursor sequence with the predicted HLA-A*24:02 epitope aligns with the highest number of different coronavirus sequences.ConclusionResults imply autoimmunity risk in COVID-19 patients with HLA-A*02:01 and HLA-A*24:02 serotypes, through molecular mimicry, as a common pathogenicity risk that can be prevalent upon getting infected with certain other coronaviruses. These results can pave the way to improved risk groups’ assessment and autoimmunity treatment options, to be used in COVID-19 and its associated diseases.

Complete mitochondrial genome sequence of Lepus yarkandensis Günther, 1875 (Lagomorpha, Leporidae): characterization and phylogenetic analysis

Lepus yarkandensis is a national second-class protected animal endemic to China and distributed only in the hot and arid Tarim Basin in Xinjiang. We sequenced and described the complete mitogenome of L. yarkandensis to analyze its characteristics and phylogeny. The species’ DNA is a 17,047 bp circular molecule that includes 13 protein-coding genes (PCGs), two rRNA genes, 22 tRNA genes, and one control region. The overall base composition was as follows: A, 31.50%; T, 29.40%; G, 13.30% and C, 25.80%, with a high A+T bias of 60.9%. In the PCGs, ND6 had deviation ranges for AT skew (–0.303) and GC skew (0.636). The Ka/Ks values of ND1 (1.067) and ND6 (1.352) genes were >1, indicating positive selection, which might play an important role in the adaptation of L. yarkandensis to arid and hot environments. The conserved sequence block, the central conserved domain, and the extended termination-associated sequences of the control region and their features were identified and described. The phylogenetic tree based on the complete mitogenome showed that L. yarkandensis was closely related to the sympatric Lepus tibetanus pamirensis. These novel datasets of L. yarkandensis can supply basic data for phylogenetic studies of Lepus spp., apart from providing essential and important resource for further genetic research and the protection of this species.

Complete Mitochondrial Genome of Three Species of the Genus Microtus (Arvicolinae, Rodentia)

The 65 species of the genus Microtus have unusual sex-related genetic features and a high rate of karyotype variation. However, only nine complete mitogenomes for these species are currently available. We describe the complete mitogenome sequences of three Microtus, which vary in length from 16,295 bp to 16,331 bp, contain 13 protein-coding genes (PCGs), two ribosomal RNA genes, 22 transfer RNA genes and a control region. The length of the 13 PCGs and the coded proteins is the same in all three species, and the start and stop codons are conserved. The non-coding regions include the L-strand origin of replication, with the same sequence of 35 bp, and the control region, which varies between 896 bp and 930 bp in length. The control region includes three domains (Domains I, II and III) with extended termination-associated sequences (ETAS-1 and ETAS-2) in Domain I. Domain II and Domain III include five (CSB-B, C, D, E and F) and three (CSB-1, CSB-2, and CSB-3) conserved sequence blocks, respectively. Phylogenetic reconstructions using the mitochondrial genomes of all the available Microtus species and one representative species from another genus of the Arvicolinae subfamily reproduced the established phylogenetic relationships for all the Arvicolinae genera that were analyzed.

Sequence Composition Underlying Centromeric and Heterochromatic Genome Compartments of the Pacific Oyster Crassostrea gigas

Segments of the genome enriched in repetitive sequences still present a challenge and are omitted in genome assemblies. For that reason, the exact composition of DNA sequences underlying the heterochromatic regions and the active centromeres are still unexplored for many organisms. The centromere is a crucial region of eukaryotic chromosomes responsible for the accurate segregation of genetic material. The typical landmark of centromere chromatin is the rapidly-evolving variant of the histone H3, CenH3, while DNA sequences packed in constitutive heterochromatin are associated with H3K9me3-modified histones. In the Pacific oyster Crassostrea gigas we identified its centromere histone variant, Cg-CenH3, that shows stage-specific distribution in gonadal cells. In order to investigate the DNA composition of genomic regions associated with the two specific chromatin types, we employed chromatin immunoprecipitation followed by high-throughput next-generation sequencing of the Cg-CenH3- and H3K9me3-associated sequences. CenH3-associated sequences were assigned to six groups of repetitive elements, while H3K9me3-associated-ones were assigned only to three. Those associated with CenH3 indicate the lack of uniformity in the chromosomal distribution of sequences building the centromeres, being also in the same time dispersed throughout the genome. The heterochromatin of C. gigas exhibited general paucity and limited chromosomal localization as predicted, with H3K9me3-associated sequences being predominantly constituted of DNA transposons.

Cloning and preliminary verification of telomere-associated sequences in upland cotton

Telomeres are structures enriched in repetitive sequences at the end of chromosomes. In this study, using the telomere primer AA(CCCTAAA)3CCC for the single primer PCR, two DNA sequences were obtained from Gossypium hirsutum (Linnaeus, 1753) accession (acc.) TM-1. Sequence analysis showed that the two obtained sequences were all rich in A/T base, which was consistent with the characteristic of the telomere-associated sequence (TAS). They were designated as GhTAS1 and GhTAS2 respectively. GhTAS1 is 489 bp long, with 57.6% of A/T, and GhTAS2 is 539 bp long, with 63.9% of A/T. Fluorescence in situ hybridization results showed that both of the cloned TASs were located at the ends of the partial chromosomes of G. hirsutum, with the strong signals, which further confirmed that GhTAS1 and GhTAS2 were telomere-associated sequences including highly tandemly repetitive sequences. Results of blast against the assembled genome of G. hirsutum showed that GhTAS sequences may be missed on some assembled chromosomes. The results provide important evidence for the evaluation of the integrity of assembled chromosome end sequences, and will also contribute to the further perfection of the draft genomes of cotton.

A strategy for large-scale comparison of evolutionary- and reaction-based classifications of enzyme function

Determining the molecular function of enzymes discovered by genome sequencing represents a primary foundation for understanding many aspects of biology. Historically, classification of enzyme reactions has used the enzyme nomenclature system developed to describe the overall reactions performed by biochemically characterized enzymes, irrespective of their associated sequences. In contrast, functional classification and assignment for the millions of protein sequences of unknown function now available is largely done in two computational steps, first by similarity-based assignment of newly obtained sequences to homologous groups, followed by transferring to them the known functions of similar biochemically characterized homologs. Due to the fundamental differences in their etiologies and practice, `how’ these chemistry- and evolution-centric functional classification systems relate to each other has been difficult to explore on a large scale. To investigate this issue in a new way, we integrated two published ontologies that had previously described each of these classification systems independently. The resulting infrastructure was then used to compare the functional assignments obtained from each classification system for the well-studied and functionally diverse enolase superfamily. Mapping these function assignments to protein structure and reaction similarity networks shows a profound and complex disconnect between the homology- and chemistry-based classification systems. This conclusion mirrors previous observations suggesting that except for closely related sequences, facile annotation transfer from small numbers of characterized enzymes to the huge number uncharacterized homologs to which they are related is problematic. Our extension of these comparisons to large enzyme superfamilies in a computationally intelligent manner provides a foundation for new directions in protein function prediction for the huge proportion of sequences of unknown function represented in major databases. Interactive sequence, reaction, substrate and product similarity networks computed for this work for the enolase and two other superfamilies are freely available for download from the Structure Function Linkage Database Archive (http://sfld.rbvi.ucsf.edu).