Chromosome-level Subgenome-aware de novo Assembly of Saccharomyces bayanus Provides Insight into Genome Divergence after Hybridization.

Interspecies hybridization is prevalent in various eukaryotic lineages and plays important roles in phenotypic diversification, adaption, and speciation. To better understand the changes that occurred in the different subgenomes of a hybrid species and how they facilitated adaptation, we completed chromosome-level de novo assemblies of all 16 pairs chromosomes for a recently formed hybrid yeast, Saccharomyces bayanus strain CBS380 (IFO11022), using Nanopore MinION long-read sequencing. Characterization of S. bayanus subgenomes and comparative analysis with the genomes of its parent species, S. uvarum and S. eubayanus, provide several new insights into understanding genome evolution after a relatively recent hybridization. For instance, multiple recombination events between the two subgenomes have been observed in each chromosome, followed by loss of heterozygosity (LOH) in most chromosomes in nine chromosome pairs. In addition to maintaining nearly all gene content and synteny from its parental genomes, S. bayanus has acquired many genes from other yeast species, primarily through the introgression of S. cerevisiae, such as those involved in the maltose metabolism. In addition, the patterns of recombination and LOH suggest an allotetraploid origin of S. bayanus. The gene acquisition and rapid LOH in the hybrid genome probably facilitated its adaption to maltose brewing environments and mitigated the maladaptive effect of hybridization.

PhyloVelo enhances transcriptomic velocity field mapping using monotonically expressed genes.

Single-cell RNA sequencing (scRNA-seq) is a powerful approach for studying cellular differentiation, but accurately tracking cell fate transitions can be challenging, especially in disease conditions. Here we introduce PhyloVelo, a computational framework that estimates the velocity of transcriptomic dynamics by using monotonically expressed genes (MEGs) or genes with expression patterns that either increase or decrease, but do not cycle, through phylogenetic time. Through integration of scRNA-seq data with lineage information, PhyloVelo identifies MEGs and reconstructs a transcriptomic velocity field. We validate PhyloVelo using simulated data and Caenorhabditis elegans ground truth data, successfully recovering linear, bifurcated and convergent differentiations. Applying PhyloVelo to seven lineage-traced scRNA-seq datasets, generated using CRISPR-Cas9 editing, lentiviral barcoding or immune repertoire profiling, demonstrates its high accuracy and robustness in inferring complex lineage trajectories while outperforming RNA velocity. Additionally, we discovered that MEGs across tissues and organisms share similar functions in translation and ribosome biogenesis.

A Genetic Bottleneck of Mitochondrial DNA During Human Lymphocyte Development.

Mitochondria are essential organelles in eukaryotic cells that provide critical support for energetic and metabolic homeostasis. Although the elimination of pathogenic mitochondrial DNA (mtDNA) mutations in somatic cells has been observed, the mechanisms to maintain proper functions despite their mtDNA mutation load are poorly understood. In this study, we analyzed somatic mtDNA mutations in more than 30,000 single human peripheral and bone marrow mononuclear cells. We observed a significant overrepresentation of homoplasmic mtDNA mutations in B, T, and natural killer (NK) lymphocytes. Intriguingly, their overall mutational burden was lower than that in hematopoietic progenitors and myeloid cells. This characteristic mtDNA mutational landscape indicates a genetic bottleneck during lymphoid development, as confirmed with single-cell datasets from multiple platforms and individuals. We further demonstrated that mtDNA replication lags behind cell proliferation in both pro-B and pre-B progenitor cells, thus likely causing the genetic bottleneck by diluting mtDNA copies per cell. Through computational simulations and approximate Bayesian computation (ABC), we recapitulated this lymphocyte-specific mutational landscape and estimated the minimal mtDNA copies as <30 in T, B, and NK lineages. Our integrative analysis revealed a novel process of a lymphoid-specific mtDNA genetic bottleneck, thus illuminating a potential mechanism used by highly metabolically active immune cells to limit their mtDNA mutation load.

TSSr: an R package for comprehensive analyses of TSS sequencing data.

Transcription initiation is regulated in a highly organized fashion to ensure proper cellular functions. Accurate identification of transcription start sites (TSSs) and quantitative characterization of transcription initiation activities are fundamental steps for studies of regulated transcriptions and core promoter structures. Several high-throughput techniques have been developed to sequence the very 5'end of RNA transcripts (TSS sequencing) on the genome scale. Bioinformatics tools are essential for processing, analysis, and visualization of TSS sequencing data. Here, we present TSSr, an R package that provides rich functions for mapping TSS and characterizations of structures and activities of core promoters based on all types of TSS sequencing data. Specifically, TSSr implements several newly developed algorithms for accurately identifying TSSs from mapped sequencing reads and inference of core promoters, which are a prerequisite for subsequent functional analyses of TSS data. Furthermore, TSSr also enables users to export various types of TSS data that can be visualized by genome browser for inspection of promoter activities in association with other genomic features, and to generate publication-ready TSS graphs. These user-friendly features could greatly facilitate studies of transcription initiation based on TSS sequencing data. The source code and detailed documentations of TSSr can be freely accessed at https://github.com/Linlab-slu/TSSr.

Delineating the longitudinal tumor evolution using organoid models.

Cancer is an evolutionary process fueled by genetic or epigenetic alterations in the genome. Understanding the evolutionary dynamics that are operative at different stages of tumor progression might inform effective strategies in early detection, diagnosis, and treatment of cancer. However, our understanding on the dynamics of tumor evolution through time is very limited since it is usually impossible to sample patient tumors repeatedly. The recent advances in in vitro 3D organoid culture technologies have opened new avenues for the development of more realistic human cancer models that mimic many in vivo biological characteristics in human tumors. Here, we review recent progresses and challenges in cancer genomic evolution studies and advantages of using tumor organoids to study cancer evolution. We propose to establish an experimental evolution model based on continuous passages of patient-derived organoids and longitudinal sampling to study clonal dynamics and evolutionary patterns over time. Development and integration of population genetic theories and computational models into time-course genomic data in tumor organoids will help to pinpoint the key cellular mechanisms underlying cancer evolutionary dynamics, thus providing novel insights on therapeutic strategies for highly dynamic and heterogeneous tumors.

The origin and evolution of a distinct mechanism of transcription initiation in yeasts.

The molecular process of transcription by RNA Polymerase II is highly conserved among eukaryotes ("classic model"). A distinct way of locating transcription start sites (TSSs) has been identified in a budding yeast Saccharomyces cerevisiae ("scanning model"). Herein, we applied genomic approaches to elucidate the origin of the scanning model and its underlying genetic mechanisms. We first identified TSSs at single-nucleotide resolution for 12 yeast species using the nAnT-iCAGE technique, which significantly improved the annotations of these genomes by providing accurate 5' boundaries for protein-coding genes. We then inferred the initiation mechanism of each species based on its TSS maps and genome sequences. We discovered that the scanning model likely originated after the split of Yarrowia lipolytica and the other budding yeasts. Species that use the scanning model showed an adenine-rich region immediately upstream of the TSS that might facilitate TSS selection. Both initiation mechanisms share a strong preference for pyrimidine-purine dinucleotides surrounding the TSS. Our results suggest that the purine is required to accurately recruit the first nucleotide, thereby increasing the chances of a messenger RNA of being capped during mRNA maturation, which is critical for efficient translation initiation during protein biosynthesis. Based on our findings, we propose a model for TSS selection in the scanning-model species, as well as a model for the stepwise process responsible for the origin and evolution of the scanning model.

Distinct roles of nucleosome sliding and histone modifications in controlling the fidelity of transcription initiation.

Regulation of gene expression starts from the transcription initiation. Regulated transcription initiation is critical for generating correct transcripts with proper abundance. The impact of epigenetic control, such as histone modifications and chromatin remodelling, on gene regulation has been extensively investigated, but their specific role in regulating transcription initiation is far from well understood. Here we aimed to better understand the roles of genes involved in histone H3 methylations and chromatin remodelling on the regulation of transcription initiation at a genome-scale using the budding yeast as a study system. We obtained and compared maps of transcription start site (TSS) at single-nucleotide resolution by nAnT-iCAGE for a strain with depletion of MINC (Mot1-Ino80C-Nc2) by Mot1p and Ino80p anchor-away (Mot1&Ino80AA) and a strain with loss of histone methylation (set1Δset2Δdot1Δ) to their wild-type controls. Our study showed that the depletion of MINC stimulated transcription initiation from many new sites flanking the dominant TSS of genes, while the loss of histone methylation generates more TSSs in the coding region. Moreover, the depletion of MINC led to less confined boundaries of TSS clusters (TCs) and resulted in broader core promoters, and such patterns are not present in the ssdΔ mutant. Our data also exhibits that the MINC has distinctive impacts on TATA-containing and TATA-less promoters. In conclusion, our study shows that MINC is required for accurate identification of bona fide TSSs, particularly in TATA-containing promoters, and histone methylation contributes to the repression of transcription initiation in coding regions.

Toxicity and metabolism of 3-bromopyruvate in Caenorhabditis elegans.

In this study, we aimed to evaluate the toxic effects, changes in life span, and expression of various metabolism-related genes in Caenorhabditis elegans, using RNA interference (RNAi) and mutant strains, after 3-bromopyruvate (3-BrPA) treatment. C. elegans was treated with various concentrations of 3-BrPA on nematode growth medium (NGM) plates, and their survival was monitored every 24 h. The expression of genes related to metabolism was measured by the real-time fluorescent quantitative polymerase chain reaction (qPCR). Nematode survival in the presence of 3-BrPA was also studied after silencing three hexokinase (HK) genes. The average life span of C. elegans cultured on NGM with 3-BrPA was shortened to 5.7 d compared with 7.7 d in the control group. hxk-1, hxk-2, and hxk-3 were overexpressed after the treatment with 3-BrPA. After successfully interfering hxk-1, hxk-2, and hxk-3, the 50% lethal concentration (LC50) of all mutant nematodes decreased with 3-BrPA treatment for 24 h compared with that of the control. All the cyp35 genes tested were overexpressed, except cyp-35B3. The induction of cyp-35A1 expression was most obvious. The LC50 values of the mutant strains cyp-35A1, cyp-35A2, cyp-35A4, cyp-35B3, and cyp-35C1 were lower than that of the control. Thus, the toxicity of 3-BrPA is closely related to its effect on hexokinase metabolism in nematodes, and the cyp-35 family plays a key role in the metabolism of 3-BrPA.

Origins and Stepwise Expansion of R2R3-MYB Transcription Factors for the Terrestrial Adaptation of Plants.

The R2R3-MYB transcription factors play critical roles in various processes in embryophytes (land plants). Here, we identified genes encoding R2R3-MYB proteins from rhodophytes, glaucophytes, Chromista, chlorophytes, charophytes, and embryophytes. We classified the R2R3-MYB genes into three subgroups (I, II, and III) based on their evolutionary history and gene structure. The subgroup I is the most ancient group that includes members from all plant lineages. The subgroup II was formed before the divergence of charophytes and embryophytes. The subgroup III genes form a monophyletic group and only comprise members from land plants with conserved exon-intron structure. Each subgroup was further divided into multiple clades. The subgroup I can be divided into I-A, I-B, I-C, and I-D. The I-A, I-B, and I-C are the most basal clades that have originated before the divergence of Archaeplastida. The I-D with the II and III subgroups form a monophyletic group, containing only green plants. The II and III subgroups form another monophyletic group with Streptophyta only. Once on land, the subgroup III genes have experienced two rounds of major expansions. The first round occurred before the origin of land plants, and the second round occurred after the divergence of land plants. Due to significant gene expansion, the subgroup III genes have become the predominant group of R2R3-MYBs in land plants. The highly unbalanced pattern of birth and death evolution of R2R3-MYB genes indicates their important roles in the successful adaptation and massive radiation of land plants to occupy a multitude of terrestrial environments.

Parallel Concerted Evolution of Ribosomal Protein Genes in Fungi and Its Adaptive Significance.

Ribosomal protein (RP) genes encode structural components of ribosomes, the cellular machinery for protein synthesis. A single functional copy has been maintained in most of 78-80 RP families in animals due to evolutionary constraints imposed by gene dosage balance. Some fungal species have maintained duplicate copies in most RP families. The mechanisms by which the RP genes were duplicated and maintained and their functional significance are poorly understood. To address these questions, we identified all RP genes from 295 fungi and inferred the timing and nature of gene duplication events for all RP families. We found that massive duplications of RP genes have independently occurred by different mechanisms in three distantly related lineages: budding yeasts, fission yeasts, and Mucoromycota. The RP gene duplicates in budding yeasts and Mucoromycota were mainly created by whole genome duplication events. However, duplicate RP genes in fission yeasts were likely generated by retroposition, which is unexpected considering their dosage sensitivity. The sequences of most RP paralogs have been homogenized by repeated gene conversion in each species, demonstrating parallel concerted evolution, which might have facilitated the retention of their duplicates. Transcriptomic data suggest that the duplication and retention of RP genes increased their transcript abundance. Physiological data indicate that increased ribosome biogenesis allowed these organisms to rapidly consume sugars through fermentation while maintaining high growth rates, providing selective advantages to these species in sugar-rich environments.

Pervasive and dynamic transcription initiation in Saccharomyces cerevisiae.

Transcription initiation is finely regulated to ensure proper expression and function of genes. The regulated transcription initiation in response to various environmental stimuli in a classic model organism Saccharomyces cerevisiae has not been systematically investigated. In this study, we generated quantitative maps of transcription start sites (TSSs) at a single-nucleotide resolution for S. cerevisiae grown in nine different conditions using no-amplification nontagging Cap analysis of gene expression (nAnT-iCAGE) sequencing. We mapped ∼1 million well-supported TSSs, suggesting highly pervasive transcription initiation in the compact genome of the budding yeast. The comprehensive TSS maps allowed us to identify core promoters for ∼96% verified protein-coding genes. We corrected misannotation of translation start codon for 122 genes and suggested an alternative start codon for 57 genes. We found that 56% of yeast genes are controlled by multiple core promoters, and alternative core promoter usage by a gene is widespread in response to changing environments. Most core promoter shifts are coupled with altered gene expression, indicating that alternative core promoter usage might play an important role in controlling gene transcriptional activities. Based on their activities in responding to environmental cues, we divided core promoters into constitutive class (55%) and inducible class (45%). The two classes of core promoters display distinctive patterns in transcriptional abundance, chromatin structure, promoter shape, and sequence context. In summary, our study improved the annotation of the yeast genome and demonstrated a much more pervasive and dynamic nature of transcription initiation in yeast than previously recognized.

YeasTSS: an integrative web database of yeast transcription start sites.

The transcription initiation landscape of eukaryotic genes is complex and highly dynamic. In eukaryotes, genes can generate multiple transcript variants that differ in 5' boundaries due to usages of alternative transcription start sites (TSSs), and the abundance of transcript isoforms are highly variable. Due to a large number and complexity of the TSSs, it is not feasible to depict details of transcript initiation landscape of all genes using text-format genome annotation files. Therefore, it is necessary to provide data visualization of TSSs to represent quantitative TSS maps and the core promoters (CPs). In addition, the selection and activity of TSSs are influenced by various factors, such as transcription factors, chromatin remodeling and histone modifications. Thus, integration and visualization of functional genomic data related to these features could provide a better understanding of the gene promoter architecture and regulatory mechanism of transcription initiation. Yeast species play important roles for the research and human society, yet no database provides visualization and integration of functional genomic data in yeast. Here, we generated quantitative TSS maps for 12 important yeast species, inferred their CPs and built a public database, YeasTSS (www.yeastss.org). YeasTSS was designed as a central portal for visualization and integration of the TSS maps, CPs and functional genomic data related to transcription initiation in yeast. YeasTSS is expected to benefit the research community and public education for improving genome annotation, studies of promoter structure, regulated control of transcription initiation and inferring gene regulatory network.

Detection and Identification of Serum Peptides Biomarker in Papillary Thyroid Cancer.

BACKGROUND Papillary thyroid cancer (PTC) is currently the most commonly diagnosed endocrine malignancy. In addition, the sex- and age-adjusted incidence of PTC has exhibited a greater increase over the last 2 decades than in many other malignancies. Thus, discovering noninvasive specific serum biomarker to distinguish PTC from cancer-free controls in its early stages remains an important goal. MATERIAL AND METHODS Serum samples from 88 PTC patients and 80 cancer-free controls were randomly allocated into training or validation sets. Serum peptide profiling was performed by matrix-assisted laser desorption/ionization-time of flight mass spectrometry (MALDI-TOF-MS) after using weak cation exchange magnetic beads (WCX-MB), and the results were evaluated by use of ClinProTools™ Software. To distinguish PTC from cancer-free controls, quick classifier (QC), supervised neural network (SNN), and genetic algorithm (GA) models were established. The models were blindly validated to verify their diagnostic capabilities. The most discriminative peaks were subsequently identified with a nano-liquid chromatography-electrospray ionization-tandem mass spectrometry system. RESULTS Six peptide ions were identified as the most discriminative peaks between the PTC and cancer-free control samples. The QC model exhibited satisfactory sensitivity and specificity among the 3 models that were validated. Two peaks, at m/z 2671.17 and m/z 1464.68, were identified as fragments of the alpha chain of fibrinogen, while a peak at m/z 1738.92 was a fragment of complement component 4A/B. CONCLUSIONS MS combined with ClinProTools™ software was able to detect peptide biomarkers in PTC patients. In addition, the constructed classification models provided a serum peptidome pattern for distinguishing PTC from cancer-free controls. Both fibrinogen a and complement C4A/B were identified as potential markers for diagnosis of PTC.

Diagnostic accuracy of MALDI-TOF mass spectrometry for non-small cell lung cancer: a meta-analysis.

Objective: To assess the overall accuracy of matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) in identifying non-small cell lung cancer (NSCLC).Methods: A comprehensive search of PubMed, EMBASE and CNKI databases as well as the reference lists from relevant articles was performed prior to July 2017. Two authors independently screened articles based on inclusion and exclusion criteria and assessed the quality of each study using the Quality Assessment of Diagnostic Accuracy Studies 2 (QADAS-2) tool. Meta-disc 1.4 and Stata12.0 software programs were used for the statistical analysis.Results: Eleven eligible articles comprising 16 studies and representing 935 subjects were included in this meta-analysis. The pooled sensitivity and specificity were 0.84 (95% CI: 0.80-0.87) and 0.77 (95% CI: 0.74-0.80), respectively. The overall diagnostic performance as measured by the area under the curve (AUC) for the summary receiver-operating characteristic (SROC) curve was 0.9380.Conclusions: MALDI-TOF MS has a high diagnostic accuracy for NSCLC.

The Analysis of Gene Expression on Fertility Decline in Caenorhabditis elegans after the Treatment with 5-Fluorouracil.

BACKGROUND: 5-Fluorouracil could lead to a decline in fertility in Caenorhabditis elegans. The aim of this study was to describe the mechanisms underlying such an altered fertility phenotype and to illustrate the specific genes and pathways that are involved in the related phenotypic changes in C. elegans. METHODS: We isolated total RNA from the samples and used a new method called Digital Gene Expression (DGE), which can rapidly identify genes with altered transcript levels. The random genes were confirmed by real-time RT-PCR. RESULTS: We analyzed the results of two methods to draw conclusions based on a comparison between C. elegans and other harmful parasites. Compared with controls, 1147 genes were up-regulated, and 1067 were down-regulated. Overall, 101 up-regulated genes had a log2 ratio higher than 8, whereas the log2 ratio of 141 down-regulated genes was higher than 8. After mapping to the reference database, 4 pathways were confirmed to be involved in this phenomenon, with statistically significant participation from 19 genes. CONCLUSION: For the first time, the transcript sequence of 5-Fu-treated worms and controls was detected. We found that 4 possible pathways, i.e., ECM-receptor interaction pathway, TGF-beta signaling pathway, Focal adhesion and Hypertrophic cardiomyopathy, may be involved in the number decline in the embryos of C. elegans. Specifically, the ECM-receptor interaction pathway and Focal adhesion may be very important pathways that alter the reproduction of C. elegans.

Diagnostic value of total plasma lysophosphatidic acid in ovarian cancer: a meta-analysis.

OBJECTIVE: This study aimed to assess the diagnostic value of lysophosphatidic acid (LPA) in ovarian cancer. METHODS: A systematic review of related studies was performed; sensitivity, specificity, and other measures about the accuracy of serum LPA in the diagnosis of ovarian cancer were pooled using random-effects models. Summary receiver operating characteristic curve analysis was used to summarize the overall test performance. RESULTS: Six studies involving 363 patients with ovarian cancer and 273 healthy control women met the inclusion criteria. The summary estimates for LPA in diagnosing ovarian cancer in the included studies were as follows: sensitivity, 0.94 [95% confidence interval (CI), 0.91-0.96]; specificity, 0.88 (95% CI, 0.83-0.91); and diagnostic odds ratio, 141.59 (95% CI, 52.1-384.63). The area under the curve and Q value for summary receiver operating characteristic curves were 0.97 and 0.92, respectively. CONCLUSIONS: The LPA assay showed high accuracy and sensitivity for the diagnosis of ovarian cancer. The present study was limited by the small number of available studies and sample size; therefore, additional studies with a better design and larger samples are needed to further assess the diagnostic accuracy of LPA.

Incidence of Japanese encephalitis, visceral leishmaniasis and malaria before and after the Wenchuan earthquake, in China.

The objective of this study was to investigate the incidence of insect-borne diseases from 2005 to 2011, before and after the 2008 Wenchuan earthquake in Longnan City, Gansu Province, China. The data include Japanese encephalitis, Kala-azar and malaria cases from 2005 to 2011 that occurred in Longnan City. We calculated the incidence rates and analyzed the epidemiological characteristics of the diseases before and after the Wenchuan earthquake. During 2005-2011, 212 Japanese encephalitis cases were reported in Longnan City, and the average incidence was 1.11/100,000. Compared with any year from 2005 to 2010 the incidence of Japanese encephalitis in Longnan City in 2011 was not significantly different (P≥0.05). From 2005 to 2011, there were 719 Kala-azar cases in Longnan City, the annual incidence was 3.77/100,000, and the incidence in males was higher than females (P<0.001). Compared with 2011, there was no significant difference in incidence of Kala-azar in 2009 or 2010 (P≥0.05). There were seven total cases of malaria from 2005 to 2011, and the annual incidence was 0.07/100,000. Wudu District and Wen County were the main endemic areas of insect-borne diseases in Longnan City. The results showed that Japanese encephalitis and Kala-azar were common insect-borne infectious diseases in Longnan City, and that the incidence of insect-borne disease did not increase after the Wenchuan earthquake. It is possible that vector control measures implemented after the earthquake prevented an increase in such diseases.

Hypoglycemic activities of A- and B-type procyanidin oligomer-rich extracts from different Cinnamon barks.

Procyanidin oligomers in Cinnamon are thought to be responsible for the biological activity in the treatment of diabetes mellitus (DM). To clarify types of procyanidin oligomers in different Cinnamon species and investigate their different effects, the present study investigated procyanidin oligomers in polyphenolic oligomer-rich extracts of three Cinnamon samples by LC-MS methods, and their hypoglycemic activities were detected in vivo and in vitro. The results showed that two of the three samples from Cinnamomum cassia were rich in B-type procyanidin oligomers, and the other sample was rich in A-type procyanidin oligomers. The Cinnamon extracts were administered at doses of 200 and 300 mg/kg body wt. in high-fat diet-fed and low-dose streptozotocin (STZ)-induced diabetic mice for 14 days. The results showed that blood glucose concentrations were significantly decreased in all Cinnamon extract groups compared with the control group (p<0.05). Administration of the Cinnamon extracts significantly increased the consumption of extracellular glucose in insulin-resistant HepG2 cells and normal HepG2 cells compared with the control group. These results suggest that both A- and B-type procyanidin oligomers in different Cinnamon species have hypoglycemic activities and may improve insulin sensitivity in type 2 DM.