A rapidly reversible mutation generates subclonal genetic diversity and unstable drug resistance.

Most genetic changes have negligible reversion rates. As most mutations that confer resistance to an adverse condition (e.g., drug treatment) also confer a growth defect in its absence, it is challenging for cells to genetically adapt to transient environmental changes. Here, we identify a set of rapidly reversible drug-resistance mutations in Schizosaccharomyces pombe that are caused by microhomology-mediated tandem duplication (MTD) and reversion back to the wild-type sequence. Using 10,000× coverage whole-genome sequencing, we identify nearly 6,000 subclonal MTDs in a single clonal population and determine, using machine learning, how MTD frequency is encoded in the genome. We find that sequences with the highest-predicted MTD rates tend to generate insertions that maintain the correct reading frame, suggesting that MTD formation has shaped the evolution of coding sequences. Our study reveals a common mechanism of reversible genetic variation that is beneficial for adaptation to environmental fluctuations and facilitates evolutionary divergence.

Luteimonas galliterrae sp. nov., isolated from poultry farm soil.

Strain SX5T was isolated from the soil of a poultry farm in Shanxi Province, PR China. The isolate was a Gram-stain-negative, rod-shaped, non-flagellated, and yellow bacterium. Growth occurred at 20-37 °C (optimum, 28 °C), pH 6.0-10.0 (optimum, pH 8.0) and 0-1 % NaCl (optimum, 0 %). Phylogenetic analysis based on 16S rRNA gene sequences indicated that strain SX5T was related to members of the genus Luteimonas, and close to Luteimonas gilva H23T (97.9 %), Luteimonas cucumeris Y4T (97.9 %), Luteimonas aquatica RIB1-20T (96.8 %), Luteimonas notoginsengisoli SYP-B804T (96.4 %) and Luteimonas panaciterrae Gsoil 068T (96.1 %). The major cellular fatty acids of strain SX5T were iso-C16 : 0, iso-C17 : 1 ω9c, iso-C15 : 0 and iso-C11 : 0 3OH. The sole isoprenoid quinone was ubiquinone Q-8, and the major polar lipid profile contained diphosphatidylglycerol, phosphatidylglycerol and phosphatidylethanolamine. Genome analyses revealed that strain SX5T had a genome size of 3.6 Mbp with a G+C content of 65.7 mol% and contained abundant carbohydrate-active enzyme genes and three putative distinct biosynthetic gene clusters, suggesting that it may have great potential to degrade and utilize complex biological organic matter and produce special secondary metabolites. Comparative genomic analyses clearly separated strain SX5T from the known species of the genus Luteimonas based on average nucleotide identity and digital DNA-DNA hybridization values below the thresholds for species delineation. Based on its phenotypic, genotypic properties and phylogenetic inference, strain SX5T represents a novel species in the genus Luteimonas, for which the name Luteimonas galliterrae sp. nov. is proposed. The type strain is SX5T (=GDMCC 1.2162T=KCTC 82443T=JCM 34401T).

Polyphase taxonomy and genome analysis reveal the adaptability of Luteolibacter rhizosphaerae sp. nov. to the rhizosphere soil of Ulmus pumila L.

A Gram-stain-negative, rod-shaped, non-flagellated, pale-yellow bacterium, designated GHJ8T, was isolated from the rhizosphere soil of Ulmus pumila L., Shanxi Province, China. Growth occurred at 20-37 °C (optimum, 28 °C), pH 6.0-11.0 (optimum, pH 8.0), and 0-1% NaCl (optimum, 0%). Phylogenetic analysis based on 16S rRNA gene sequences indicated that strain GHJ8T was related to members of the genus Luteolibacter, and close to Luteolibacter flavescens GKXT (98.5%), Luteolibacter luteus G-1-1-1T (97.3%), Luteolibacter arcticus MC 3726T (97.2%), and Luteolibacter marinus NBU1238T (96.0%). The genome size of strain GHJ8T was 6.2 Mbp, with a G + C content of 62.5%. Genomic mining revealed that the strain contained antibiotic resistance genes and secondary metabolic gene clusters, indicating that it had adaptation mechanisms to environmental stress. Comparative genomic analyses clearly separated strain GHJ8T from the recognized species of the genus Luteolibacter based on average nucleotide identity (ANI) and digital DNA-DNA hybridization (dDDH) values below the thresholds for species delineation. The major cellular fatty acids were iso-C14:0 (30.8%), C16:1 ω9c (23.0%), C16:0 (17.3%), and C14:0 (13.4%). The quinone system was composed of the major menaquinones MK-8, MK-9, and MK-10, and the principal polar lipids were diphosphatidylglycerol, phosphatidylethanolamine, phosphatidylglycerol, an unidentified aminophospholipid, an unidentified glycolipid, two unidentified phospholipids, and three unidentified lipids. Based on its phenotypic and genotypic properties and phylogenetic inference, strain GHJ8T is a novel species of the genus Luteolibacter, for which the name Luteolibacter rhizosphaerae sp. nov. is proposed. The type strain is GHJ8T (= GDMCC 1.2160T = KCTC 82452T = JCM 34400T).

Roseomonas populi sp. nov., an acetate-degrading bacteria isolated from the stem of Populus tomentosa.

Strain CN29T, isolated from the stem of 5- to 6-year-old Populus tomentosa in Shandong, China, was characterized using a polyphasic taxonomic approach. Cells of CN29T were Gram-stain negative, aerobic, nonspore-forming, and nonmotile coccoid. Growth occurred at 20-37 °C, pH 4.0-9.0 (optimum, pH 6.0), and with 0-1% NaCl (optimum, 1%). Phylogenetic analysis based on the 16S rRNA gene sequence indicated that strain CN29T was closely related to members of the genus Roseomonas and closest to Roseomonas pecuniae N75T (96.6%). This classification was further supported by phylogenetic analysis using additional core genes. The average nucleotide identity and digital DNA‒DNA hybridization values between strain CN29T and Roseomonas populi CN29T were 82.7% and 27.8%, respectively. The genome size of strain CN29T was 5.87 Mb, with a G + C content of 70.9%. The major cellular fatty acids included summed feature 8 (C18:1 ω7c/C18:1 ω6c), C19:0 cyclo ω8c and C16:0. The major respiratory quinone was Q-10. The polar lipids were phosphatidylcholine, aminolipid, phosphatidylglycerol, and diphosphatidylglycerol. Strain CN29T can utilize acetate as a carbon source for growth and metabolism. Additionally, it contains acid phosphatase (2-naphthyl phosphate), which catalyzes the hydrolysis of phosphoric monoesters. The CN29T strain contains several genes, including maeB, gdhB, and cysJ, involved in carbon, nitrogen, and sulfur cycling. These findings suggest that the strain may actively participate in ecosystem cycling, leading to soil improvement and promoting the growth of poplar trees. Based on the phylogenetic, phenotypic, and genotypic characteristics, strain CN29T is concluded to represent a novel species of the genus Roseomonas, for which the name Roseomonas populi sp. nov. is proposed. The type strain is CN29T (= JCM 35579T = GDMCC 1.3267T).

Differentiation and Identification of Endophytic Bacteria from Populus Based on Mass Fingerprints and Gene Sequences.

Plant endophytic bacteria play important roles in plants' growth and resistance to stress. It is important to characterize endophytic bacteria to be able to understand their benefits. Matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) has become a powerful technique for bacterial identification due to its high throughput and simple procedures. In this study, the endophytic bacteria separated from Populus (the leaves, roots and stems of Populus tomentosa Carrière; stems of Populus nigra Linn. var. nigra; and stems of Populus canadensis Moench) were identified and classified based on MALDI-TOF MS data and 16S rRNA gene sequencing. The sampling and preparation of bacteria were optimized to obtain meaningful protein mass fingerprints. The composite correlation index (CCI) values of the inter-genera and inter-species protein mass fingerprints demonstrated sufficient differences between the strains. In the CCI value matrix for ten species in the same genus, all the CCI values were less than 0.5. Among the species, 95.6% of all the CCI values were less than 0.5. After data processing, the classification capacity of the protein mass fingerprints was verified using inter-specific and inter-generic PCoA. To compare different methods' potential for differentiation and phylogenetic analysis, a dendrogram of the MS profiles and a phylogenetic tree based on the 16S rRNA gene sequences were constructed using 61 endophytic bacteria found in Populus. The clustering and grouping results show that the phylogenetic analysis based on MALDI-TOF MS is similar to that based on 16S rRNA gene sequencing. This study provides a valuable reference for differentiating and identifying endophytic bacteria according to their protein mass fingerprints.

The molecular chaperone Hsp90 regulates heterochromatin assembly through stabilizing multiple complexes in fission yeast.

In the fission yeast Schizosaccharomyces pombe, both RNAi machinery and RNAi-independent factors mediate transcriptional and posttranscriptional silencing and heterochromatin formation. Here, we show that the silencing of reporter genes at major native heterochromatic loci (centromeres, telomeres, mating-type locus and rDNA regions) and an artificially induced heterochromatin locus is alleviated in a fission yeast hsp90 mutant, hsp90-G84C Also, H3K9me2 enrichment at heterochromatin regions, especially at the mating-type locus and subtelomeres, is compromised, suggesting heterochromatin assembly defects. We further discovered that Hsp90 is required for stabilization or assembly of the RNA-induced transcriptional silencing (RITS) and Argonaute siRNA chaperone (ARC) RNAi effector complexes, the RNAi-independent factor Fft3, the shelterin complex subunit Poz1 and the Snf2/HDAC-containing repressor complex (SHREC). Our ChIP data suggest that Hsp90 regulates the efficient recruitment of the methyltransferase/ubiquitin ligase complex CLRC by shelterin to chromosome ends and targeting of the SHREC and Fft3 to mating type locus and/or rDNA region. Finally, our genetic analyses demonstrated that increased heterochromatin spreading restores silencing at subtelomeres in the hsp90-G84C mutant. Thus, this work uncovers a conserved factor critical for promoting RNAi-dependent and -independent heterochromatin assembly and gene silencing through stabilizing multiple effectors and effector complexes.

Roseococcus pinisoli sp. nov., lacking pufL and pufM bacteriochlorophyll a: synthesizing genes.

A Gram-stain-negative, cocci-to-oval-shaped bacterial strain, designated XZZS9T, was isolated from the rhizosphere soil of Pinus sylvestris var. mongolica and characterized taxonomically using a polyphasic approach. Growth occurred at 20-35 °C (optimum, 28 °C), pH 6.0-11.0 (optimum, pH 7.0), and in 0-1% NaCl (optimum, 0%). Phylogenetic analysis based on 16S rRNA gene sequencing indicated that strain XZZS9T was related to members of the genus Roseococcus, with the highest sequence identity to Roseococcus microcysteis NIBR12T (96.9%). The major cellular fatty acids (> 5% of the total) were C18:1 ω7c and C19:0 cyclo ω8c. The major isoprenoid quinone was Q-9 and the polar lipid profile contained diphosphatidylglycerol, phosphatidylglycerol, phosphatidylcholine, phosphatidylethanolamine, an unidentified glycophospholipid, and an unidentified phospholipid. Genome sequencing revealed that had a genome size of 4.79 Mbp with a G + C content of 69.5%. Comparative genomic analyses clearly separated strain XZZS9T from the known species of the genus Roseococcus based on average nucleotide identity (ANI) and digital DNA-DNA hybridization (dDDH) values below the thresholds for species delineation. Genome annotations did not find pufL and pufM genes in strain XZZS9T, suggesting a possible lack of photosynthetic reaction. Based on genotypic and phenotypic characteristics, strain XZZS9T represents a novel species of the genus Roseococcus, for which we propose the name Roseococcus pinisoli sp. nov. The type strain is XZZS9T (= KCTC 82435T = JCM 34402T = GDMCC 1.2158T).

Sphingomonas quercus sp. nov., Isolated from Rhizosphere Soil of Quercus mongolica.

Strain XMGL2T, isolated from rhizosphere soil of Quercus mongolica in China, was characterized using a polyphasic taxonomic approach. Cells were Gram-negative, aerobic, non-spore-forming, and rod-shaped. Growth occurred at 20-37 °C (optimum, 28 °C), pH 5.0-10.0 (optimum, pH 6.0), and with 0-1% NaCl (optimum, 1%). Phylogenetic analysis based on 16S rRNA gene sequences indicated that strain XMGL2T was related to members of the genus Sphingomonas and had the highest 16S rRNA gene sequence identity to Sphingomonas oleivorans FW-11 T (96.4%). The average nucleotide identity and digital DNA-DNA hybridization values between strain XMGL2T and the closely related taxa Sphingomonas oleivorans FW-11 T and Sphingomonas fennica K101T were 75.3/19.8% and 75.8/20.2%, respectively. The major cellular fatty acids were C18:1 ω7c, C14:0 2-OH, and C16:0. The major isoprenoid quinone was Q-10 and the polar lipid profile contained diphosphatidylglycerol, phosphatidylglycerol, phosphatidylcholine, phosphatidylethanolamine, phosphatidyldimethylethanolamine, phosphatidylmonomethylethanolamine, an unidentified glycophospholipid and an unidentified phospholipid. The genomic DNA G + C content was 67.9%. Based on the phenotypic and genotypic properties and phylogenetic inference, strain XMGL2T represents a novel species of the genus Sphingomonas, for which the name Sphingomonas quercus sp. nov. is proposed. The type strain is XMGL2T (= JCM 34441 T = GDMCC 1.2153 T).

Centromere repositioning causes inversion of meiosis and generates a reproductive barrier.

The chromosomal position of each centromere is determined epigenetically and is highly stable, whereas incremental cases have supported the occurrence of centromere repositioning on an evolutionary time scale (evolutionary new centromeres, ENCs), which is thought to be important in speciation. The mechanisms underlying the high stability of centromeres and its functional significance largely remain an enigma. Here, in the fission yeast Schizosaccharomyces pombe, we identify a feedback mechanism: The kinetochore, whose assembly is guided by the centromere, in turn, enforces centromere stability. Upon going through meiosis, specific inner kinetochore mutations induce centromere repositioning-inactivation of the original centromere and formation of a new centromere elsewhere-in 1 of the 3 chromosomes at random. Repositioned centromeres reside asymmetrically in the pericentromeric regions and cells carrying them are competent in mitosis and homozygotic meiosis. However, when cells carrying a repositioned centromere are crossed with those carrying the original centromere, the progeny suffer severe lethality due to defects in meiotic chromosome segregation. Thus, repositioned centromeres constitute a reproductive barrier that could initiate genetic divergence between 2 populations with mismatched centromeres, documenting a functional role of ENCs in speciation. Surprisingly, homozygotic repositioned centromeres tend to undergo meiosis in an inverted order-that is, sister chromatids segregate first, and homologous chromosomes separate second-whereas the original centromeres on other chromosomes in the same cell undergo meiosis in the canonical order, revealing hidden flexibility in the perceived rigid process of meiosis.

A non-symbiotic novel species, Rhizobium populisoli sp. nov., isolated from rhizosphere soil of Populus popularis.

Strain XQZ8T, isolated from the rhizosphere soil of a Populus popularis plant in China, was characterized using a polyphasic taxonomic approach. Cells were Gram-negative, aerobic, non-spore-forming, and rod-shaped. Phylogenetic analysis based on 16S rRNA gene sequences indicated that strain XQZ8T was related to members of the genus Rhizobium and had the highest 16S rRNA gene sequence similarity to Rhizobium smilacinae PTYR-5T (96.6%). The average nucleotide identity and digital DNA-DNA hybridization value between strain XQZ8T and R. smilacinae PTYR-5T were 77.5% and 21.4%, respectively. TYGS whole-genome-based taxonomic and multi-locus sequence analyses of three concatenated housekeeping genes (atpD-recA-glnII) further indicated that strain XQZ8T was a new member of the genus Rhizobium. The major cellular fatty acids included summed feature 8 (C18:1 ω7c/C18:1 ω6c), summed feature 2 (C12:0 aldehyde/unknown 10.928), C16:0, and C19:0 cyclo ω8c. The major respiratory quinones were Q-9 and Q-10. The polar lipids were phosphatidylglycerol, phosphatidylcholine, phosphatidylethanolamine, phosphatidyldimethylethanolamine, phosphatidylmonomethylethanolamine, an unidentified glycophospholipid, and three unidentified lipids. The genomic DNA G + C content of the strain was 60.1 mol%. Based on the phylogenetic, phenotypic, and genotypic characteristics, strain XQZ8T represents a novel species of the genus Rhizobium, for which the name Rhizobium populisoli sp. nov. is proposed. The type strain is XQZ8T (= JCM 34442T = GDMCC 1.2201T).

A Degeneration Gradient of Poplar Trees Contributes to the Taxonomic, Functional, and Resistome Diversity of Bacterial Communities in Rhizosphere Soils.

Bacterial communities associated with roots influence the health and nutrition of the host plant. However, the microbiome discrepancy are not well understood under different healthy conditions. Here, we tested the hypothesis that rhizosphere soil microbial diversity and function varies along a degeneration gradient of poplar, with a focus on plant growth promoting bacteria (PGPB) and antibiotic resistance genes. Comprehensive metagenomic analysis including taxonomic investigation, functional detection, and ARG (antibiotics resistance genes) annotation revealed that available potassium (AK) was correlated with microbial diversity and function. We proposed several microbes, Bradyrhizobium, Sphingomonas, Mesorhizobium, Nocardioides, Variovorax, Gemmatimonadetes, Rhizobacter, Pedosphaera, Candidatus Solibacter, Acidobacterium, and Phenylobacterium, as candidates to reflect the soil fertility and the plant health. The highest abundance of multidrug resistance genes and the four mainly microbial resistance mechanisms (antibiotic efflux, antibiotic target protection, antibiotic target alteration, and antibiotic target replacement) in healthy poplar rhizosphere, corroborated the relationship between soil fertility and microbial activity. This result suggested that healthy rhizosphere soil harbored microbes with a higher capacity and had more complex microbial interaction network to promote plant growing and reduce intracellular levels of antibiotics. Our findings suggested a correlation between the plant degeneration gradient and bacterial communities, and provided insight into the role of high-turnover microbial communities as well as potential PGPB as real-time indicators of forestry soil quality, and demonstrated the inner interaction contributed by the bacterial communities.

Inverted meiosis: an alternative way of chromosome segregation for reproduction.

Canonical meiosis is characterized by two sequential rounds of nuclear divisions following one round of DNA replication-reductional segregation of homologous chromosomes during the first division and equational segregation of sister chromatids during the second division. Meiosis in an inverted order of two nuclear divisions-inverted meiosis has been observed in several species with holocentromeres as an adaptive strategy to overcome the obstacle in executing a canonical meiosis due to the holocentric chromosome structure. Recent findings of co-existence of inverted and canonical meiosis in two monocentric organisms, human and fission yeast, suggested that inverted meiosis could be common and also lead to the puzzle regarding the mechanistic feasibility for executing two meiosis programs simultaneously. Here, we discuss apparent conflicts for concurrent canonical meiosis and inverted meiosis. Furthermore, we attempt to provide a working model that may be compatible for both forms of meiosis.

Replication stress affects the fidelity of nucleosome-mediated epigenetic inheritance.

The fidelity of epigenetic inheritance or, the precision by which epigenetic information is passed along, is an essential parameter for measuring the effectiveness of the process. How the precision of the process is achieved or modulated, however, remains largely elusive. We have performed quantitative measurement of epigenetic fidelity, using position effect variegation (PEV) in Schizosaccharomyces pombe as readout, to explore whether replication perturbation affects nucleosome-mediated epigenetic inheritance. We show that replication stresses, due to either hydroxyurea treatment or various forms of genetic lesions of the replication machinery, reduce the inheritance accuracy of CENP-A/Cnp1 nucleosome positioning within centromere. Mechanistically, we demonstrate that excessive formation of single-stranded DNA, a common molecular abnormality under these conditions, might have correlation with the reduction in fidelity of centromeric chromatin duplication. Furthermore, we show that replication stress broadly changes chromatin structure at various loci in the genome, such as telomere heterochromatin expanding and mating type locus heterochromatin spreading out of the boundaries. Interestingly, the levels of inheritable expanding at sub-telomeric heterochromatin regions are highly variable among independent cell populations. Finally, we show that HU treatment of the multi-cellular organisms C. elegans and D. melanogaster affects epigenetically programmed development and PEV, illustrating the evolutionary conservation of the phenomenon. Replication stress, in addition to its demonstrated role in genetic instability, promotes variable epigenetic instability throughout the epigenome.

Ccp1 modulates epigenetic stability at centromeres and affects heterochromatin distribution in Schizosaccharomyces pombe.

Distinct chromatin organization features, such as centromeres and heterochromatin domains, are inherited epigenetically. However, the mechanisms that modulate the accuracy of epigenetic inheritance, especially at the individual nucleosome level, are not well-understood. Here, using ChIP and next-generation sequencing (ChIP-Seq), we characterized Ccp1, a homolog of the histone chaperone Vps75 in budding yeast that functions in centromere chromatin duplication and heterochromatin maintenance in fission yeast (Schizosaccharomyces pombe). We show that Ccp1 is enriched at the central core regions of the centromeres. Of note, among all histone chaperones characterized, deletion of the ccp1 gene uniquely reduced the rate of epigenetic switching, manifested as position effect variegation within the centromeric core region (CEN-PEV). In contrast, gene deletion of other histone chaperones either elevated the PEV switching rates or did not affect centromeric PEV. Ccp1 and the kinetochore components Mis6 and Sim4 were mutually dependent for centromere or kinetochore association at the proper levels. Moreover, Ccp1 influenced heterochromatin distribution at multiple loci in the genome, including the subtelomeric and the pericentromeric regions. We also found that Gar2, a protein predominantly enriched in the nucleolus, functions similarly to Ccp1 in modulating the epigenetic stability of centromeric regions, although its mechanism remained unclear. Together, our results identify Ccp1 as an important player in modulating epigenetic stability and maintaining proper organization of multiple chromatin domains throughout the fission yeast genome.

Intricate regulation on epigenetic stability of the subtelomeric heterochromatin and the centromeric chromatin in fission yeast.

In eukaryotes, the integrity of chromatin structure and organization is crucial to diverse key cellular processes from development to disease avoidance. To maintain the cell identity through mitotic cell generations, the genome (the genomic DNA sequence) as well as the epigenome (pertaining various forms of epigenetic information carriers, such as histone modifications, nucleosome positioning and the chromatin organization) is inherited with high fidelity. In comparison to the wealth of knowledge on genetic stability, we know much less on what may control the accuracy of epigenetic inheritance. In our recent work in the fission yeast Schizosaccharomyces pombe, by quantifying the epigenetic fidelity of CENP-A/Cnp1 or H3K9me2 nucleosome inheritance through cell divisions, we demonstrated that Ccp1, a homolog of histone chaperone Vps75 in budding yeast, participates in the modulation of centromeric nucleosomal epigenetic stability as well as proper heterochromatin organization. In this essay, we focus on discussing the uniquely high dynamicity of the subtelomeric heterochromatin regions and the complex mechanisms regulating epigenetic stability of centromeric chromatin.

Fission yeast Scm3: A CENP-A receptor required for integrity of subkinetochore chromatin.

The mechanisms ensuring specific incorporation of CENP-A at centromeres are poorly understood. Mis16 and Mis18 are required for CENP-A localization at centromeres and form a complex that is conserved from fission yeast to human. Fission yeast sim1 mutants that alleviate kinetochore domain silencing are defective in Scm3(Sp), the ortholog of budding yeast Scm3(Sc). Scm3(Sp) depends on Mis16/18 for its centromere localization and like them is recruited to centromeres in late anaphase. Importantly, Scm3(Sp) coaffinity purifies with CENP-A(Cnp1) and associates with CENP-A(Cnp1) in vitro, yet localizes independently of intact CENP-A(Cnp1) chromatin and is differentially released from chromatin. While Scm3(Sc) has been proposed to form a unique hexameric nucleosome with CENP-A(Cse4) and histone H4 at budding yeast point centromeres, we favor a model in which Scm3(Sp) acts as a CENP-A(Cnp1) receptor/assembly factor, cooperating with Mis16 and Mis18 to receive CENP-A(Cnp1) from the Sim3 escort and mediate assembly of CENP-A(Cnp1) into subkinetochore chromatin.

DANPOS: dynamic analysis of nucleosome position and occupancy by sequencing.

Recent developments in next-generation sequencing have enabled whole-genome profiling of nucleosome organizations. Although several algorithms for inferring nucleosome position from a single experimental condition have been available, it remains a challenge to accurately define dynamic nucleosomes associated with environmental changes. Here, we report a comprehensive bioinformatics pipeline, DANPOS, explicitly designed for dynamic nucleosome analysis at single-nucleotide resolution. Using both simulated and real nucleosome data, we demonstrated that bias correction in preliminary data processing and optimal statistical testing significantly enhances the functional interpretation of dynamic nucleosomes. The single-nucleotide resolution analysis of DANPOS allows us to detect all three categories of nucleosome dynamics, such as position shift, fuzziness change, and occupancy change, using a uniform statistical framework. Pathway analysis indicates that each category is involved in distinct biological functions. We also analyzed the influence of sequencing depth and suggest that even 200-fold coverage is probably not enough to identify all the dynamic nucleosomes. Finally, based on nucleosome data from the human hematopoietic stem cells (HSCs) and mouse embryonic stem cells (ESCs), we demonstrated that DANPOS is also robust in defining functional dynamic nucleosomes, not only in promoters, but also in distal regulatory regions in the mammalian genome.

Plasticity and epigenetic inheritance of centromere-specific histone H3 (CENP-A)-containing nucleosome positioning in the fission yeast.

Nucleosomes containing the specific histone H3 variant CENP-A mark the centromere locus on each chromatin and initiate kinetochore assembly. For the common type of regional centromeres, little is known in molecular detail of centromeric chromatin organization, its propagation through cell division, and how distinct organization patterns may facilitate kinetochore assembly. Here, we show that in the fission yeast S. pombe, a relatively small number of CENP-A/Cnp1 nucleosomes are found within the centromeric core and that their positioning relative to underlying DNA varies among genetically homogenous cells. Consistent with the flexible positioning of Cnp1 nucleosomes, a large portion of the endogenous centromere is dispensable for its essential activity in mediating chromosome segregation. We present biochemical evidence that Cnp1 occupancy directly correlates with silencing of the underlying reporter genes. Furthermore, using a newly developed pedigree analysis assay, we demonstrated the epigenetic inheritance of Cnp1 positioning and quantified the rate of occasional repositioning of Cnp1 nucleosomes throughout cell generations. Together, our results reveal the plasticity and the epigenetically inheritable nature of centromeric chromatin organization.

Nucleosome fragility reveals novel functional states of chromatin and poises genes for activation.

The structural complexity of nucleosomes underlies their functional versatility. Here we report a new type of complexity-nucleosome fragility, manifested as high sensitivity to micrococcal nuclease, in contrast to the common presumption that nucleosomes are similar in resistance to MNase digestion. Using differential MNase digestion of chromatin and high-throughput sequencing, we have identified a special group of nucleosomes termed "fragile nucleosomes" throughout the yeast genome, nearly 1000 of which were at previously determined "nucleosome-free" loci. Nucleosome fragility is broadly implicated in multiple chromatin processes, including transcription, translocation, and replication, in correspondence to specific physiological states of cells. In the environmental-stress-response genes, the presence of fragile nucleosomes prior to the occurrence of environmental changes suggests that nucleosome fragility poises genes for swift up-regulation in response to the environmental changes. We propose that nucleosome fragility underscores distinct functional statuses of the chromatin and provides a new dimension for portraying the landscape of genome organization.

Enhancing Salt Tolerance in Poplar Seedlings through Arbuscular Mycorrhizal Fungi Symbiosis.

Poplar (Populus spp.) is a valuable tree species with multiple applications in afforestation. However, its growth in saline areas, including coastal regions, is limited. This study aimed to investigate the physiological mechanisms of arbuscular mycorrhizal fungi (AMF) symbiosis with 84K (P. alba × P. tremula var. glandulosa) poplar under salt stress. We conducted pot experiments using NaCl solutions of 0 mM (control), 100 mM (moderate stress), and 200 mM (severe stress) and evaluated the colonization of AMF and various physiological parameters of plants, including photosynthesis, biomass, antioxidant enzyme activity, nutrients, and ion concentration. Partial least squares path modeling (PLS-PM) was employed to elucidate how AMF can improve salt tolerance in poplar. The results demonstrated that AMF successfully colonized the roots of plants under salt stress, effectively alleviated water loss by increasing the transpiration rate, and significantly enhanced the biomass of poplar seedlings. Mycorrhiza reduced proline and malondialdehyde accumulation while enhancing the activity of antioxidant enzymes, thus improving plasma membrane stability. Additionally, AMF mitigated Na+ accumulation in plants, contributing to the maintenance of a favorable ion balance. These findings highlight the effectiveness of using suitable AMF to improve conditions for economically significant tree species in salt-affected areas, thereby promoting their utilization.