Adaptive Selection of Cis-regulatory Elements in the Han Chinese.

Cis-regulatory elements have an important role in human adaptation to the living environment. However, the lag in population genomic cohort studies and epigenomic studies, hinders the research in the adaptive analysis of cis-regulatory elements in human populations. In this study, we collected 4,013 unrelated individuals and performed a comprehensive analysis of adaptive selection of genome-wide cis-regulatory elements in the Han Chinese. In total, 12.34% of genomic regions are under the influence of adaptive selection, where 1.00% of enhancers and 2.06% of promoters are under positive selection, and 0.06% of enhancers and 0.02% of promoters are under balancing selection. Gene ontology enrichment analysis of these cis-regulatory elements under adaptive selection reveals that many positive selections in the Han Chinese occur in pathways involved in cell-cell adhesion processes, and many balancing selections are related to immune processes. Two classes of adaptive cis-regulatory elements related to cell adhesion were in-depth analyzed, one is the adaptive enhancers derived from neanderthal introgression, leads to lower hyaluronidase level in skin, and brings better performance on UV-radiation resistance to the Han Chinese. Another one is the cis-regulatory elements regulating wound healing, and the results suggest the positive selection inhibits coagulation and promotes angiogenesis and wound healing in the Han Chinese. Finally, we found that many pathogenic alleles, such as risky alleles of type 2 diabetes or schizophrenia, remain in the population due to the hitchhiking effect of positive selections. Our findings will help deepen our understanding of the adaptive evolution of genome regulation in the Han Chinese.

Exploring noncoding variants in genetic diseases: from detection to functional insights.

Previous studies on genetic diseases predominantly focused on protein-coding variations, overlooking the vast noncoding regions in the human genome. The development of high-throughput sequencing technologies and functional genomics tools has enabled the systematic identification of functional noncoding variants. These variants can impact gene expression, regulation, and chromatin conformation, thereby contributing to disease pathogenesis. Understanding the mechanisms that underlie the impact of noncoding variants on genetic diseases is indispensable for the development of precisely targeted therapies and the implementation of personalized medicine strategies. The intricacies of noncoding regions introduce a multitude of challenges and research opportunities. In this review, we introduce a spectrum of noncoding variants involved in genetic diseases, along with research strategies and advanced technologies for their precise identification and in-depth understanding of the complexity of the noncoding genome. We will delve into the research challenges and propose potential solutions for unraveling the genetic basis of rare and complex diseases.

Recent positive selection signatures reveal phenotypic evolution in the Han Chinese population.

Characterizing natural selection signatures and relationships with phenotype spectra is important for understanding human evolution and both biological and pathological mechanisms. Here, we identified 24 genetic loci under recent selection by analyzing rare singletons in 3946 high-depth whole-genome sequencing data of Han Chinese. The loci include immune-related gene regions (MHC cluster, IGH cluster, STING1, and PSG), alcohol metabolism-related gene regions (ADH1B, ALDH2, and ALDH3B2), and the olfactory perception gene OR4C16, in which the MHC cluster, ADH1B, and ALDH2 were also identified by TOPMed and WestLake Biobank. Among the signals, the IGH cluster is particularly interesting, in which the favored allele of variant 14_105737776_C_T (rs117518546, IgG1-G396R) promotes immune response, but also increases the risk of an autoimmune disease systemic lupus erythematosus (SLE). It is also surprising that our newly discovered ALDH3B2 evolved in the opposite direction to ALDH2 for alcohol metabolism. Besides monogenic traits, we found that multiple complex traits experienced polygenic adaptation. Particularly, multi-methods consistently revealed that lower blood pressure was favored in natural selection. Finally, we built a database named RePoS (recent positive selection, http://bigdata.ibp.ac.cn/RePoS/) to integrate and display multi-population selection signals. Our study extended our understanding of natural evolution and phenotype adaptation in Han Chinese as well as other populations.

Noncoding RNA mutations in cancer.

Cancer is driven by both germline and somatic genetic changes. Efforts have been devoted to characterizing essential genetic variations in cancer initiation and development. Most attention has been given to mutations in protein-coding genes and associated regulatory elements such as promoters and enhancers. The development of sequencing technologies and in silico and experimental methods has allowed further exploration of cancer predisposition variants and important somatic mutations in noncoding RNAs, mainly for long noncoding RNAs and microRNAs. Association studies including GWAS have revealed hereditary variations including SNPs and indels in lncRNA or miRNA genes and regulatory regions. These mutations altered RNA secondary structures, expression levels, and target recognition and then conferred cancer predisposition to carriers. Whole-exome/genome sequencing comparing cancer and normal tissues has revealed important somatic mutations in noncoding RNA genes. Mutation hotspots and somatic copy number alterations have been identified in various tumor-associated noncoding RNAs. Increasing focus and effort have been devoted to studying the noncoding region of the genome. The complex genetic network of cancer initiation is being unveiled. This article is categorized under: RNA in Disease and Development > RNA in Disease.

Characterization of genome-wide STR variation in 6487 human genomes.

Short tandem repeats (STRs) are abundant and highly mutagenic in the human genome. Many STR loci have been associated with a range of human genetic disorders. However, most population-scale studies on STR variation in humans have focused on European ancestry cohorts or are limited by sequencing depth. Here, we depicted a comprehensive map of 366,013 polymorphic STRs (pSTRs) constructed from 6487 deeply sequenced genomes, comprising 3983 Chinese samples (~31.5x, NyuWa) and 2504 samples from the 1000 Genomes Project (~33.3x, 1KGP). We found that STR mutations were affected by motif length, chromosome context and epigenetic features. We identified 3273 and 1117 pSTRs whose repeat numbers were associated with gene expression and 3'UTR alternative polyadenylation, respectively. We also implemented population analysis, investigated population differentiated signatures, and genotyped 60 known disease-causing STRs. Overall, this study further extends the scale of STR variation in humans and propels our understanding of the semantics of STRs.

NPInter v5.0: ncRNA interaction database in a new era.

Noncoding RNAs (ncRNAs) play key regulatory roles in biological processes by interacting with other biomolecules. With the development of high-throughput sequencing and experimental technologies, extensive ncRNA interactions have been accumulated. Therefore, we updated the NPInter database to a fifth version to document these interactions. ncRNA interaction entries were doubled from 1 100 618 to 2 596 695 by manual literature mining and high-throughput data processing. We integrated global RNA-DNA interactions from iMARGI, ChAR-seq and GRID-seq, greatly expanding the number of RNA-DNA interactions (from 888 915 to 8 329 382). In addition, we collected different types of RNA interaction between SARS-CoV-2 virus and its host from recently published studies. Long noncoding RNA (lncRNA) expression specificity in different cell types from tumor single cell RNA-seq (scRNA-seq) data were also integrated to provide a cell-type level view of interactions. A new module named RBP was built to display the interactions of RNA-binding proteins with annotations of localization, binding domains and functions. In conclusion, NPInter v5.0 (http://bigdata.ibp.ac.cn/npinter5/) provides informative and valuable ncRNA interactions for biological researchers.

Long noncoding RNA MAGI2-AS3 regulates the H2O2 level and cell senescence via HSPA8.

The redox homeostasis system regulates many biological processes, intracellular antioxidant production and redox signaling. However, long noncoding RNAs (lncRNAs) involved in redox regulation have rarely been reported. Herein, we reported that downregulation of MAGI2-AS3 decreased the superoxide level in Human fibroblasts (Fbs), a replicative aging model, as detected by the fluorescent probes dihydroethidium (DHE) and MitoSOX™ Red. RNA pulldown combined with mass spectrometry showed that HSPA8 is a novel interacting protein of MAGI2-AS3, which was further confirmed by photoactivatable ribonucleoside-enhanced crosslinking and immunoprecipitation (PAR-CLIP). Downregulation of MAGI2-AS3 decreased the hydrogen peroxide (H2O2) content by stabilizing the HSPA8 protein level via inhibiting the protesome degradation of HSPA8. Further evidence showed that MAGI2-AS3 interacted with the C-terminal domain (CTD) of HSPA8. Downregulation of MAGI2-AS3 delayed cell senescence, while this antiaging effect was abolished by HSPA8 knockdown. The underlying molecular mechanism by which MAGI2-AS3 knockdown inhibited cell senescence was mediated via suppression of the ROS/MAP2K6/p38 signaling pathway. Taken together, these findings revealed that downregulation of lncRNA MAGI2-AS3 decreased the H2O2 content and delayed cell senescence by stabilizing the HSPA8 protein level, identifying a potential antiaging application.

Mutational Pattern Induced by 5-Fluorouracil and Oxaliplatin in the Gut Microbiome.

Chemotherapeutic agents, such as 5-fluorouracil (5-FU) and oxaliplatin (Oxi), can not only kill the cancer cell but also influence the proliferation of gut microbiota; however, the interaction between these drugs and gut microbiota remains poorly understood. In this study, we developed a powerful framework for taxonomy composition and genomic variation analysis to investigate the mutagenesis effect and proliferation influence of chemotherapeutic agents, such as 5-FU and Oxi, on gut microbiota and the interaction between these drugs and gut microbiota during chemotherapy. Using the gut microbiome data, we detected 1.45 million variations among the chemotherapy groups and found the drugs significantly affected mutation signatures of gut microbiota. Oxi notably increased transversion rate, whereas 5-FU reduced the rate. Traits related to cell division and nutrient mobilization showed evidence of strong selection pressure from chemotherapeutic agents. In addition, drug-associated bacteriome shift patterns and functional alterations were found: the metabolism changes in the 5-FU group implied that gut microbiota could provide additional nicotinamide adenine dinucleotide (NAD+) to inhibit cancer cell autophagy; in the Oxi group, the ribosome and lysine biosynthesis genes were obviously enriched. Our study provides a blueprint for characterizing the role of microbes and drug-microbe interaction in the gut microbiota response to chemotherapy.

Novel MDM2 Inhibitor XR-2 Exerts Potent Anti-Tumor Efficacy and Overcomes Enzalutamide Resistance in Prostate Cancer.

Background: The inactivation of tumor-suppressor p53 plays an important role in second generation anti-androgens (SGAs) drug resistance and neuroendocrine differentiation in castration-resistant prostate cancer (CRPC). The reactivation of p53 by blocking the MDM2-p53 interaction represents an attractive therapeutic remedy in cancers with wild-type or functional p53. Whether MDM2-p53 inhibitor could overcome SGAs drug resistance in CRPC is still needed further research. Here, we investigated the anti-tumor efficacy and mechanisms of a novel MDM2-p53 inhibitor XR-2 in CRPC. Methods: To investigate the functions and mechanisms of XR-2 in prostate cancer, in vitro and in vivo biofunctional assays were performed. Western blot and qRT-PCR assay were performed to detect the protein and mRNA expression levels of indicated genes. CCK8, colony formation, flow cytometry and senescence assays were performed for cell function identifications. RNA-sequencing and bioinformatics analysis were mainly used to identify the influence of XR-2 on prostate cancer cells transcriptome. Subcutaneous 22Rv1 derived xenografts mice model was used to investigate the in vivo anti-tumor activity of XR-2. In addition, the broad-spectrum anti-tumor activities in vivo of XR-2 were evaluated by different xenografts mice models. Results: XR-2 could directly bind to MDM2, potently reactivate the p53 pathway and thus induce cell cycle arrest and apoptosis in wild-type p53 CRPC cell lines. XR-2 also suppresses the AR pathway as p53 regulates AR transcription inhibition and MDM2 participates in AR degradation. As a result, XR-2 efficiently inhibited CRPC cell viability, showed a synergistic effect with enzalutamide and overcame enzalutamide resistance both in vitro and in vivo. Moreover, results illustrated that XR-2 possesses broad-spectrum anti-tumor activities in vivo with favourable safety. Conclusion: MDM2-p53 inhibitor (XR-2) possesses potently prostate cancer progresses inhibition activity both in vitro and in vivo. XR-2 shows a synergistic effect with enzalutamide and overcomes enzalutamide resistance.

Characterizing mobile element insertions in 5675 genomes.

Mobile element insertions (MEIs) are a major class of structural variants (SVs) and have been linked to many human genetic disorders, including hemophilia, neurofibromatosis, and various cancers. However, human MEI resources from large-scale genome sequencing are still lacking compared to those for SNPs and SVs. Here, we report a comprehensive map of 36 699 non-reference MEIs constructed from 5675 genomes, comprising 2998 Chinese samples (∼26.2×, NyuWa) and 2677 samples from the 1000 Genomes Project (∼7.4×, 1KGP). We discovered that LINE-1 insertions were highly enriched in centromere regions, implying the role of chromosome context in retroelement insertion. After functional annotation, we estimated that MEIs are responsible for about 9.3% of all protein-truncating events per genome. Finally, we built a companion database named HMEID for public use. This resource represents the latest and largest genomewide study on MEIs and will have broad utility for exploration of human MEI findings.

piRBase: integrating piRNA annotation in all aspects.

Piwi-interacting RNAs are a type of small noncoding RNA that have various functions. piRBase is a manually curated resource focused on assisting piRNA functional analysis. piRBase release v3.0 is committed to providing more comprehensive piRNA related information. The latest release covers >181 million unique piRNA sequences, including 440 datasets from 44 species. More disease-related piRNAs and piRNA targets have been collected and displayed. The regulatory relationships between piRNAs and targets have been visualized. In addition to the reuse and expansion of the content in the previous version, the latest version has additional new content, including gold standard piRNA sets, piRNA clusters, piRNA variants, splicing-junction piRNAs, and piRNA expression data. In addition, the entire web interface has been redesigned to provide a better experience for users. piRBase release v3.0 is free to access, browse, search, and download at http://bigdata.ibp.ac.cn/piRBase.

NyuWa Genome resource: A deep whole-genome sequencing-based variation profile and reference panel for the Chinese population.

The lack of haplotype reference panels and whole-genome sequencing resources specific to the Chinese population has greatly hindered genetic studies in the world's largest population. Here, we present the NyuWa genome resource, based on deep (26.2×) sequencing of 2,999 Chinese individuals, and construct a NyuWa reference panel of 5,804 haplotypes and 19.3 million variants, which is a high-quality publicly available Chinese population-specific reference panel with thousands of samples. Compared with other panels, the NyuWa reference panel reduces the Han Chinese imputation error rate by a margin ranging from 30% to 51%. Population structure and imputation simulation tests support the applicability of one integrated reference panel for northern and southern Chinese. In addition, a total of 22,504 loss-of-function variants in coding and noncoding genes are identified, including 11,493 novel variants. These results highlight the value of the NyuWa genome resource in facilitating genetic research in Chinese and Asian populations.

SmProt: A Reliable Repository with Comprehensive Annotation of Small Proteins Identified from Ribosome Profiling.

Small proteins specifically refer to proteins consisting of less than 100 amino acids translated from small open reading frames (sORFs), which were usually missed in previous genome annotation. The significance of small proteins has been revealed in current years, along with the discovery of their diverse functions. However, systematic annotation of small proteins is still insufficient. SmProt was specially developed to provide valuable information on small proteins for scientific community. Here we present the update of SmProt, which emphasizes reliability of translated sORFs, genetic variants in translated sORFs, disease-specific sORF translation events or sequences, and remarkably increased data volume. More components such as non-ATG translation initiation, function, and new sources are also included. SmProt incorporated 638,958 unique small proteins curated from 3,165,229 primary records, which were computationally predicted from 419 ribosome profiling (Ribo-seq) datasets or collected from literature and other sources from 370 cell lines or tissues in 8 species (Homo sapiens, Mus musculus, Rattus norvegicus, Drosophila melanogaster, Danio rerio, Saccharomyces cerevisiae, Caenorhabditis elegans, and Escherichia coli). In addition, small protein families identified from human microbiomes were also collected. All datasets in SmProt are free to access, and available for browse, search, and bulk downloads at http://bigdata.ibp.ac.cn/SmProt/.

Global profiling of RNA-binding protein target sites by LACE-seq.

RNA-binding proteins (RBPs) have essential functions during germline and early embryo development. However, current methods are unable to identify the in vivo targets of a RBP in these low-abundance cells. Here, by coupling RBP-mediated reverse transcription termination with linear amplification of complementary DNA ends and sequencing, we present the LACE-seq method for identifying RBP-regulated RNA networks at or near the single-oocyte level. We determined the binding sites and regulatory mechanisms for several RBPs, including Argonaute 2 (Ago2), Mili, Ddx4 and Ptbp1, in mature mouse oocytes. Unexpectedly, transcriptomics and proteomics analysis of Ago2-/- oocytes revealed that Ago2 interacts with endogenous small interfering RNAs (endo-siRNAs) to repress mRNA translation globally. Furthermore, the Ago2 and endo-siRNA complexes fine-tune the transcriptome by slicing long terminal repeat retrotransposon-derived chimeric transcripts. The precise mapping of RBP-binding sites in low-input cells opens the door to studying the roles of RBPs in embryonic development and reproductive diseases.

Genomes of 12 fig wasps provide insights into the adaptation of pollinators to fig syconia.

Figs and fig pollinators are one of the few classic textbook examples of obligate pollination mutualism. The specific dependence of fig pollinators on the relatively safe living environment with sufficient food sources in the enclosed fig syconia implies that they are vulnerable to habitat changes. However, there is still no extensive genomic evidence to reveal the evolutionary footprint of this long-term mutually beneficial symbiosis in fig pollinators. In fig syconia, there are also non-pollinator species. The non-pollinator species differ in their evolutionary and life histories from pollinators. We conducted comparative analyses on 11 newly sequenced fig wasp genomes and one previously published genome. The pollinators colonized the figs approximately 66.9 million years ago, consistent with the origin of host figs. Compared with non-pollinators, many more genes in pollinators were subject to relaxed selection. Seven genes were absent in pollinators in response to environmental stress and immune activation. Pollinators had more streamlined gene repertoires in the innate immune system, chemosensory toolbox, and detoxification system. Our results provide genomic evidence for the differentiation between pollinators and nonpollinators. The data suggest that owing to the long-term adaptation to the fig, some genes related to functions no longer required are absent in pollinators.

NONCODEV6: an updated database dedicated to long non-coding RNA annotation in both animals and plants.

NONCODE (http://www.noncode.org/) is a comprehensive database of collection and annotation of noncoding RNAs, especially long non-coding RNAs (lncRNAs) in animals. NONCODEV6 is dedicated to providing the full scope of lncRNAs across plants and animals. The number of lncRNAs in NONCODEV6 has increased from 548 640 to 644 510 since the last update in 2017. The number of human lncRNAs has increased from 172 216 to 173 112. The number of mouse lncRNAs increased from 131 697 to 131 974. The number of plant lncRNAs is 94 697. The relationship between lncRNAs in human and cancer were updated with transcriptome sequencing profiles. Three important new features were also introduced in NONCODEV6: (i) updated human lncRNA-disease relationships, especially cancer; (ii) lncRNA annotations with tissue expression profiles and predicted function in five common plants; iii) lncRNAs conservation annotation at transcript level for 23 plant species. NONCODEV6 is accessible through http://www.noncode.org/.

Active retrotransposons help maintain pericentromeric heterochromatin required for faithful cell division.

Retrotransposons are populated in vertebrate genomes, and when active, are thought to cause genome instability with potential benefit to genome evolution. Retrotransposon-derived RNAs are also known to give rise to small endo-siRNAs to help maintain heterochromatin at their sites of transcription; however, as not all heterochromatic regions are equally active in transcription, it remains unclear how heterochromatin is maintained across the genome. Here, we address these problems by defining the origins of repeat-derived RNAs and their specific chromatin locations in Drosophila S2 cells. We demonstrate that repeat RNAs are predominantly derived from active gypsy elements and processed by Dcr-2 into small RNAs to help maintain pericentromeric heterochromatin. We also show in cultured S2 cells that synthetic repeat-derived endo-siRNA mimics are sufficient to rescue Dcr-2-deficiency-induced defects in heterochromatin formation in interphase and chromosome segregation during mitosis, demonstrating that active retrotransposons are required for stable genetic inheritance.

RIC-seq for global in situ profiling of RNA-RNA spatial interactions.

Highly structured RNA molecules usually interact with each other, and associate with various RNA-binding proteins, to regulate critical biological processes. However, RNA structures and interactions in intact cells remain largely unknown. Here, by coupling proximity ligation mediated by RNA-binding proteins with deep sequencing, we report an RNA in situ conformation sequencing (RIC-seq) technology for the global profiling of intra- and intermolecular RNA-RNA interactions. This technique not only recapitulates known RNA secondary structures and tertiary interactions, but also facilitates the generation of three-dimensional (3D) interaction maps of RNA in human cells. Using these maps, we identify noncoding RNA targets globally, and discern RNA topological domains and trans-interacting hubs. We reveal that the functional connectivity of enhancers and promoters can be assigned using their pairwise-interacting RNAs. Furthermore, we show that CCAT1-5L-a super-enhancer hub RNA-interacts with the RNA-binding protein hnRNPK, as well as RNA derived from the MYC promoter and enhancer, to boost MYC transcription by modulating chromatin looping. Our study demonstrates the power and applicability of RIC-seq in discovering the 3D structures, interactions and regulatory roles of RNA.