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Twenty years after the completion and forty years after the proposal of the Human Genome Project (HGP), genomics, together with its twin field — bioinformatics, has entered a new paradigm, where its bioscience-related, discipline-centric applications have been creating many new research frontiers. Beijing Institute of Genomics (BIG), now also known as China National Center for Bioinformation (CNCB), will play key roles in supporting and participating in these frontier research activities. On the 20th anniversary of the establishment of BIG, we provide a brief retrospective of its historic events and ascertain strategic research directions with a broader vision for future genomics, where digital genome, digital medicine, and digital health are so structured to meet the needs of human life and healthcare, as well as their related metaverses.

Complete and accurate reference genomes and annotations provide fundamental resources for functional genomics and crop breeding. Here we report a de novo assembly and annotation of a pea cultivar ZW6 with contig N50 of 8.98 Mb, which features a 243-fold increase in contig length and evident improvements in the continuity and quality of sequence in complex repeat regions compared with the existing one. Genome diversity of 118 cultivated and wild pea demonstrated that Pisum abyssinicumis a separate species different from P. fulvumand P. sativumwithin Pisum. Quantitative trait locus analyses uncovered two known Mendel’s genes related to stem length (Le/le) and seed shape (R/r) as well as some candidate genes for pod form studied by Mendel. A pan-genome of 116 pea accessions was constructed, and pan-genes preferred in P. abyssinicumand P. fulvumshowed distinct functional enrichment, indicating the potential value of them as pea breeding resources in the future.

Baylisascaris schroederi, a roundworm (ascaridoid) parasite specific to the bamboo-feeding giant panda(Ailuropoda melanoleuca), represents a leading cause of mortality in wild giant panda populations. Here, we present a 293-megabase chromosome-level genome assembly of B. schroederito infer its biology, including host adaptations. Comparative genomics revealed an evolutionary trajectory accompanied by host-shift events in ascaridoid parasite lineages after host separations, suggesting their potential for transmission and rapid adaptation to new hosts. Genomic and anatomical lines of evidence, including expansion and positive selection of genes related to the cuticle and basal metabolisms, indicate that B. schroederiundergoes specific adaptations to survive in the sharp-edged bamboo-enriched gut of giant pandas by structurally increasing its cuticle thickness and efficiently utilizing host nutrients through gut parasitism. Additionally, we characterized the secretome of B. schroederiand predicted potential drug and vaccine targets for new control strategies. Overall, this genome resource provides new insights into the host adaptation of B. schroederito the giant panda as well as the host-shift events in ascaridoid parasite lineages. Our findings on the unique biology of B. schroederiwill also aid in the development of prevention and treatment measures to protect giant panda populations from roundworm parasitism.

Baylisascaris schroederi, a roundworm (ascaridoid) parasite specific to the bamboo-feeding giant panda(Ailuropoda melanoleuca), represents a leading cause of mortality in wild giant panda populations. Here, we present a 293-megabase chromosome-level genome assembly of B. schroederito infer its biology, including host adaptations. Comparative genomics revealed an evolutionary trajectory accompanied by host-shift events in ascaridoid parasite lineages after host separations, suggesting their potential for transmission and rapid adaptation to new hosts. Genomic and anatomical lines of evidence, including expansion and positive selection of genes related to the cuticle and basal metabolisms, indicate that B. schroederiundergoes specific adaptations to survive in the sharp-edged bamboo-enriched gut of giant pandas by structurally increasing its cuticle thickness and efficiently utilizing host nutrients through gut parasitism. Additionally, we characterized the secretome of B. schroederiand predicted potential drug and vaccine targets for new control strategies. Overall, this genome resource provides new insights into the host adaptation of B. schroederito the giant panda as well as the host-shift events in ascaridoid parasite lineages. Our findings on the unique biology of B. schroederiwill also aid in the development of prevention and treatment measures to protect giant panda populations from roundworm parasitism.

Coding regions have complex interactions among multiple selective forces, which are manifested as biases in nucleotide composition. Previous studies have revealed a decreasing GC gradient from the 5′-end to 3′-end of coding regions in various organisms. We confirmed that this gradient is universal in eukaryotic genes, but the decrease only starts from the ∼ 25th codon. This trend is mostly found in nonsynonymous (ns) sites at which the GC gradient is universal across the eukaryotic genome. Increased GC contents at ns sites result in cheaper amino acids, indicating a universal selection for energy efficiencytoward the N-termini of encoded proteins. Within a genome, the decreasing GC gradient is intensified from lowly to highly expressed genes (more and more protein products), further supporting this hypothesis. This reveals a conserved selective constraint for cheaper amino acids at the translation start that drives the increased GC contents at ns sites. Elevated GC contents can facilitate transcriptionbut result in a more stable local secondary structure around the start codon and subsequently impede translation initiation. Conversely, the GC gradients at four-fold and two-fold synonymous sites vary across species. They could decrease or increase, suggesting different constraints acting at the GC contents of different codon sites in different species. This study reveals that the overall GC contents at the translation start are consequences of complex interactions among several major biological processes that shape the nucleotide sequences, especially efficient energy usage.

•Two novel blaKPC-2-bearing plasmids were identified and sequenced.•A novel type of blaKPC-2genetic structure was identified.•Spread of the plasmids was due to clonal dissemination of ST11 KPC-producing K. pneumoniae.

Agrocybe cylindracea, an edible mushroom, is widely cultivated for its abundance of nutrientsand flavor, and many of its metabolites are reported to have beneficial roles, such as medicinal effects on tumors and chronical illnesses. However, the lack of genomic information has hindered further molecular studies on this fungus. Here, we present a genome assembly of A. cylindraceatogether with comparative genomics and pathway analyses of Agaricales species. The draft, generated from both next-generation sequencing (NGS) and single-molecule real-time (SMRT) sequencing platforms to overcome high genetic heterozygosity, is composed of a 56.5 Mb sequence and 15,384 predicted genes. This mushroom possesses a complex reproductive system, including tetrapolar heterothallic and secondary homothallic mechanisms, and harbors several hydrolases and peptidases for gradual and effective degradation of various carbon sources. Our pathway analysis reveals complex processes involved in the biosynthesis of polysaccharides and other active substances, including B vitamins, unsaturated fatty acids, and N-acetylglucosamine. RNA-seq data show that A. cylindraceastipes tend to synthesize carbohydrate for carbon sequestration and energy storage, whereas pilei are more active in carbon utilization and unsaturated fatty acid biosynthesis. These results reflect diverse functions of the two anatomical structures of the fruiting body. Our comprehensive genomic and transcriptomic data, as well as preliminary comparative analyses, provide insights into the molecular details of the medicinal effects in terms of active compounds and nutrient components.

Agrocybe cylindracea, an edible mushroom, is widely cultivated for its abundance of nutrientsand flavor, and many of its metabolites are reported to have beneficial roles, such as medicinal effects on tumors and chronical illnesses. However, the lack of genomic information has hindered further molecular studies on this fungus. Here, we present a genome assembly of A. cylindraceatogether with comparative genomics and pathway analyses of Agaricales species. The draft, generated from both next-generation sequencing (NGS) and single-molecule real-time (SMRT) sequencing platforms to overcome high genetic heterozygosity, is composed of a 56.5 Mb sequence and 15,384 predicted genes. This mushroom possesses a complex reproductive system, including tetrapolar heterothallic and secondary homothallic mechanisms, and harbors several hydrolases and peptidases for gradual and effective degradation of various carbon sources. Our pathway analysis reveals complex processes involved in the biosynthesis of polysaccharides and other active substances, including B vitamins, unsaturated fatty acids, and N-acetylglucosamine. RNA-seq data show that A. cylindraceastipes tend to synthesize carbohydrate for carbon sequestration and energy storage, whereas pilei are more active in carbon utilization and unsaturated fatty acid biosynthesis. These results reflect diverse functions of the two anatomical structures of the fruiting body. Our comprehensive genomic and transcriptomic data, as well as preliminary comparative analyses, provide insights into the molecular details of the medicinal effects in terms of active compounds and nutrient components.

Genome reannotationaims for complete and accurate characterization of gene modelsand thus is of critical significance for in-depth exploration of gene function. Although the availability of massive RNA-seqdata provides great opportunities for gene model refinement, few efforts have been made to adopt these precious data in ricegenome reannotation. Here we reannotate the rice (Oryza sativaL. ssp. japonica) genome based on integration of large-scale RNA-seq data and release a new annotation system IC4R-2.0. In general, IC4R-2.0 significantly improves the completeness of gene structure, identifies a number of novel genes, and integrates a variety of functional annotations. Furthermore, long non-coding RNAs (lncRNAs) and circular RNAs (circRNAs) are systematically characterized in the rice genome. Performance evaluation shows that compared to previous annotation systems, IC4R-2.0 achieves higher integrity and quality, primarily attributable to massive RNA-seq data applied in genome annotation. Consequently, we incorporate the improved annotations into the Information Commons for Rice (IC4R), a database integrating multiple omics data of rice, and accordingly update IC4R by providing more user-friendly web interfaces and implementing a series of practical online tools. Together, the updated IC4R, which is equipped with the improved annotations, bears great promise for comparative and functional genomic studies in rice and other monocotyledonous species. The IC4R-2.0 annotation system and related resources are freely accessible at http://ic4r.org/.

Genome reannotationaims for complete and accurate characterization of gene modelsand thus is of critical significance for in-depth exploration of gene function. Although the availability of massive RNA-seqdata provides great opportunities for gene model refinement, few efforts have been made to adopt these precious data in ricegenome reannotation. Here we reannotate the rice (Oryza sativaL. ssp. japonica) genome based on integration of large-scale RNA-seq data and release a new annotation system IC4R-2.0. In general, IC4R-2.0 significantly improves the completeness of gene structure, identifies a number of novel genes, and integrates a variety of functional annotations. Furthermore, long non-coding RNAs (lncRNAs) and circular RNAs (circRNAs) are systematically characterized in the rice genome. Performance evaluation shows that compared to previous annotation systems, IC4R-2.0 achieves higher integrity and quality, primarily attributable to massive RNA-seq data applied in genome annotation. Consequently, we incorporate the improved annotations into the Information Commons for Rice (IC4R), a database integrating multiple omics data of rice, and accordingly update IC4R by providing more user-friendly web interfaces and implementing a series of practical online tools. Together, the updated IC4R, which is equipped with the improved annotations, bears great promise for comparative and functional genomic studies in rice and other monocotyledonous species. The IC4R-2.0 annotation system and related resources are freely accessible at http://ic4r.org/.

Cold seeps in the deep sea are closely linked to energy exploration as well as global climate change. The alkane-dominated chemical energy-driven model makes cold seeps an oasis of deep-sea life, showcasing an unparalleled reservoir of microbial genetic diversity. Here, by analyzing 113 metagenomes collected from 14 global sites across 5 cold seep types, we present a comprehensive Cold Seep Microbiomic Database (CSMD) to archive the genomic and functional diversity of cold seep microbiomes. The CSMD includes over 49 million non-redundant genes and 3175 metagenome-assembled genomes, which represent 1895 species spanning 105 phyla. In addition, beta diversity analysis indicates that both the sampling site and cold seep type have a substantial impact on the prokaryotic microbiome community composition. Heterotrophic and anaerobic metabolisms are prevalent in microbial communities, accompanied by considerable mixotrophs and facultative anaerobes, highlighting the versatile metabolic potential in cold seeps. Furthermore, secondary metabolic gene cluster analysis indicates that at least 98.81% of the sequences potentially encode novel natural products, with ribosomally synthesized and post-translationally modified peptides being the predominant type widely distributed in archaea and bacteria. Overall, the CSMD represents a valuable resource that would enhance the understanding and utilization of global cold seep microbiomes.

Snapdragon (Antirrhinum majusL.), a member of the Plantaginaceae family, is an important model for plant genetics and molecular studies on plant growth and development, transposon biology and self-incompatibility. Here we report a near-complete genome assembly of A. majuscultivar JI7 (A. majuscv.JI7) comprising 510 Megabases (Mb) of genomic sequence and containing 37,714 annotated protein-coding genes. Scaffolds covering 97.12% of the assembled genome were anchored on eight chromosomes. Comparative and evolutionary analyses revealed that a whole-genome duplication event occurred in the Plantaginaceae around 46–49 million years ago (Ma). We also uncovered the genetic architectures associated with complex traits such as flower asymmetry and self-incompatibility, identifying a unique duplication of TCP family genes dated to around 46–49 Ma and reconstructing a near-complete ψS-locus of roughly 2 Mb. The genome sequence obtained in this study not only provides a representative genome sequenced from the Plantaginaceae but also brings the popular plant model system of Antirrhinuminto the genomic age.

The rapid development of high-throughput sequencing technologies has led to a dramatic decrease in the money and time required for de novogenome sequencing or genome resequencing projects, with new genome sequences constantly released every week. Among such projects, the plethora of updated genome assemblies induces the requirement of version-dependent annotation files and other compatible public dataset for downstream analysis. To handle these tasks in an efficient manner, we developed the reference-based genome assemblyand annotation tool (RGAAT), a flexible toolkit for resequencing-based consensus building and annotation update. RGAAT can detect sequence variants with comparable precision, specificity, and sensitivity to GATK and with higher precision and specificity than Freebayes and SAMtools on four DNA-seq datasets tested in this study. RGAAT can also identify sequence variants based on cross-cultivar or cross-version genomic alignments. Unlike GATK and SAMtools/BCFtools, RGAAT builds the consensus sequence by taking into account the true allele frequency. Finally, RGAAT generates a coordinate conversion file between the reference and query genomes using sequence variants and supports annotation file transfer. Compared to the rapid annotation transfer tool (RATT), RGAAT displays better performance characteristics for annotation transfer between different genome assemblies, strains, and species. In addition, RGAAT can be used for genome modification, genome comparison, and coordinate conversion. RGAAT is available at https://sourceforge.net/projects/rgaat/and https://github.com/wushyer/RGAAT_v2at no cost.

Domestic rice (Oryza sativaL.) is one of the most important cereal crops, feeding a large number of worldwide populations. Along with various high-throughput genome sequencing projects, rice genomics has been making great headway toward direct field applications of basic research advances in understanding the molecular mechanisms of agronomical traits and utilizing diverse germplasm resources. Here, we briefly review its achievements over the past two decades and present the potential for its bright future.

Domestic rice (Oryza sativaL.) is one of the most important cereal crops, feeding a large number of worldwide populations. Along with various high-throughput genome sequencing projects, rice genomics has been making great headway toward direct field applications of basic research advances in understanding the molecular mechanisms of agronomical traits and utilizing diverse germplasm resources. Here, we briefly review its achievements over the past two decades and present the potential for its bright future.

The rapid development of high-throughput sequencing technologies has led to a dramatic decrease in the money and time required for de novogenome sequencing or genome resequencing projects, with new genome sequences constantly released every week. Among such projects, the plethora of updated genome assemblies induces the requirement of version-dependent annotation files and other compatible public dataset for downstream analysis. To handle these tasks in an efficient manner, we developed the reference-based genome assemblyand annotation tool (RGAAT), a flexible toolkit for resequencing-based consensus building and annotation update. RGAAT can detect sequence variants with comparable precision, specificity, and sensitivity to GATK and with higher precision and specificity than Freebayes and SAMtools on four DNA-seq datasets tested in this study. RGAAT can also identify sequence variants based on cross-cultivar or cross-version genomic alignments. Unlike GATK and SAMtools/BCFtools, RGAAT builds the consensus sequence by taking into account the true allele frequency. Finally, RGAAT generates a coordinate conversion file between the reference and query genomes using sequence variants and supports annotation file transfer. Compared to the rapid annotation transfer tool (RATT), RGAAT displays better performance characteristics for annotation transfer between different genome assemblies, strains, and species. In addition, RGAAT can be used for genome modification, genome comparison, and coordinate conversion. RGAAT is available at https://sourceforge.net/projects/rgaat/and https://github.com/wushyer/RGAAT_v2at no cost.

Gastric carcinoma is a heterogeneous malignant disease involving genetic factors. To identify predictive markers for gastric cancertreatment in Chinese patients, we evaluated the association between polymorphismsof the gene encoding cytochrome P450 2A6 (CYP2A6) and outcomes of S-1 plus oxaliplatin (SOX) chemotherapy treatment. Clinical data on 60 consecutive gastric cancer patients receiving SOX regimen were collected prospectively. We sequenced all exons of CYP2A6and a total of 22 different polymorphisms were detected in the present study. Comprehensive analyses of these genetic polymorphisms were performed to determine their association with both safety and efficacy of SOX regimen. Our results showed that polymorphisms of CYP2A6were associated with the safety and efficacy of SOX treatment. Among them, missense mutations CYP2A6rs60823196 and rs138978736 could be possible risk factors (P< 0.05) for severe diarrhea induced by SOX, whereas CYP2A6rs138978736 could be a conceivable predictor for overall survival of patients treated with SOX adjuvant chemotherapy. Further large-scale randomized prospective studies are warranted to confirm these findings.

Gastric carcinoma is a heterogeneous malignant disease involving genetic factors. To identify predictive markers for gastric cancertreatment in Chinese patients, we evaluated the association between polymorphismsof the gene encoding cytochrome P450 2A6 (CYP2A6) and outcomes of S-1 plus oxaliplatin (SOX) chemotherapy treatment. Clinical data on 60 consecutive gastric cancer patients receiving SOX regimen were collected prospectively. We sequenced all exons of CYP2A6and a total of 22 different polymorphisms were detected in the present study. Comprehensive analyses of these genetic polymorphisms were performed to determine their association with both safety and efficacy of SOX regimen. Our results showed that polymorphisms of CYP2A6were associated with the safety and efficacy of SOX treatment. Among them, missense mutations CYP2A6rs60823196 and rs138978736 could be possible risk factors (P<0.05) for severe diarrhea induced by SOX, whereas CYP2A6rs138978736 could be a conceivable predictor for overall survival of patients treated with SOX adjuvant chemotherapy. Further large-scale randomized prospective studies are warranted to confirm these findings.

Rice is one of the most important stable food as well as a monocotyledonous model organism for the plant research community. Here, we present RED (Rice Expression Database; http://expression.ic4r.org), an integrated database of rice gene expression profiles derived entirely from RNA-Seq data. RED features a comprehensive collection of 284 high-quality RNA-Seq experiments, integrates a large number of gene expression profiles and covers a wide range of rice growth stages as well as various treatments. Based on massive expression profiles, RED provides a list of housekeeping and tissue-specific genes and dynamically constructs co-expression networks for gene(s) of interest. Besides, it provides user-friendly web interfaces for querying, browsing and visualizing expression profiles of concerned genes. Together, as a core resource in BIG Data Center, RED bears great utility for characterizing the function of rice genes and better understanding important biological processes and mechanisms underlying complex agronomic traits in rice.

Programmed DNA elimination is a developmentally regulated process leading to the reproducible loss of specific genomic sequences. DNA elimination occurs in unicellular ciliates and a variety of metazoans, including invertebrates and vertebrates. In metazoa, DNA elimination typically occurs in somatic cells during early development, leaving the germline genome intact. Reference genomes for metazoa that undergo DNA elimination are not available. Here, we generated germline and somatic reference genome sequences of the DNA eliminating pig parasitic nematode Ascaris suumand the horse parasite Parascaris univalens.In addition, we carried out in-depth analyses of DNA elimination in the parasitic nematode of humans, Ascaris lumbricoides,and the parasitic nematode of dogs, Toxocara canis. Our analysis of nematode DNA elimination reveals that in all species, repetitive sequences (that differ among the genera) and germline-expressed genes (approximately 1000–2000 or 5%–10% of the genes) are eliminated. Thirty-five percent of these eliminated genes are conserved among these nematodes, defining a core set of eliminated genes that are preferentially expressed during spermatogenesis. Our analysis supports the view that DNA elimination in nematodes silences germline-expressed genes. Over half of the chromosome break sites are conserved between Ascarisand Parascaris, whereas only 10% are conserved in the more divergent T. canis.Analysis of the chromosomal breakage regions suggests a sequence-independent mechanism for DNA breakage followed by telomere healing, with the formation of more accessible chromatin in the break regions prior to DNA elimination. Our genome assemblies and annotations also provide comprehensive resources for analysis of DNA elimination, parasitology research, and comparative nematode genome and epigenome studies.

Transcripts are expressed spatially and temporally and they are very complicated, precise and specific; however, most studies are focused on protein-coding related genes. Recently, massively parallel cDNA sequencing (RNA-seq) has emerged to be a new and promising tool for transcriptome research, and numbers of non-coding RNAs, especially lincRNAs, have been widely identified and well characterized as important regulators of diverse biological processes. In this study, we used ultra-deep RNA-seq data from 15 mouse tissues to study the diversity and dynamic of non-coding RNAs in mouse. Using our own criteria, we identified totally 16,249 non-coding genes (21,569 non-coding RNAs) in mouse. We annotated these non-coding RNAs by diverse properties and found non-coding RNAs are generally shorter, have fewer exons, express in lower level and are more strikingly tissue-specific compared with protein-coding genes. Moreover, these non-coding RNAs show significant enrichment with transcriptional initiation and elongation signals including histone modifications (H3K4me3, H3K27me3 and H3K36me3), RNAPII binding sites and CAGE tags. The gene set enrichment analysis (GSEA) result revealed several sets of lincRNAs associated with diverse biological processes such as immune effector process, muscle development and sexual reproduction. Taken together, this study provides a more comprehensive annotation of mouse non-coding RNAs and gives an opportunity for future functional and evolutionary study of mouse non-coding RNAs.

Twenty years after the completion and forty years after the proposal of the Human Genome Project (HGP), genomics, together with its twin field – bioinformatics, has entered a new paradigm, where its bioscience-related, discipline-centric applications have been creating many new research frontiers. Beijing Institute of Genomics (BIG) and now also known as China National Center for Bioinformation (CNCB), will play key roles in supporting and participating in these frontier research activities. On the 20thanniversary of the establishment of BIG, we provide a brief retrospective of its historic events and ascertain strategic research directions with a broader vision for future genomics, where digital genome, digital medicine, and digital health are so structured to meet the needs of human life and healthcare, as well as their related metaverses.

With the development of genomics and bioinformatics, especially the extensive applications of high-throughput sequencing technology, more transcriptional units with little or no protein-coding potential have been discovered. Such RNA molecules are called non-protein-coding RNAs (npcRNAs or ncRNAs). Among them, long npcRNAs or ncRNAs (lnpcRNAs or lncRNAs) represent diverse classes of transcripts longer than 200 nucleotides. In recent years, the lncRNAs have been considered as important regulators in many essential biological processes. In plants, although a large number of lncRNA transcripts have been predicted and identified in few species, our current knowledge of their biological functions is still limited. Here, we have summarized recent studies on their identification, characteristics, classification, bioinformatics, resources, and current exploration of their biological functions in plants.

With the development of genomics and bioinformatics, especially the extensive applications of high-throughput sequencing technology, more transcriptional units with little or no protein-coding potential have been discovered. Such RNA molecules are called non-protein-coding RNAs (npcRNAs or ncRNAs). Among them, long npcRNAs or ncRNAs (lnpcRNAs or lncRNAs) represent diverse classes of transcripts longer than 200 nucleotides. In recent years, the lncRNAs have been considered as important regulators in many essential biological processes. In plants, although a large number of lncRNA transcripts have been predicted and identified in few species, our current knowledge of their biological functions is still limited. Here, we have summarized recent studies on their identification, characteristics, classification, bioinformatics, resources, and current exploration of their biological functions in plants.

Spermatogenesis is a complicated and poorly understood process that relies on the precise regulation of the self-renewal and differentiation of spermatogonia. In many organisms, microRNAs (miRNAs) are involved in multiple developmental processes as critical regulators of transcriptional and post-transcriptional gene silencing. This study investigated the expression pattern of miRNAs in type B spermatogonia cells (BSc) and primary spermatocytes (PSc) of mice, using a high-throughput small RNA sequencing system. The results revealed that the expression levels of Let-7 family miRNAs were remarkably high in both cell types. Furthermore, the expression levels of miR-21, miR-140-3p, miR-103, miR-30a, miR-101b and miR-99b were decreased during the transformation from BSc to PSc. These miRNAs target vital genes that participate in apoptosis, cell proliferation and differentiation, junction assembly and cell cycle regulation. These results highlight the indispensable role of miRNAs in spermatogenesis.

Ovary development is a complex process involving numerous genes. A well-developed ovary is essential for females to keep fertility and reproduce offspring. In order to gain a better insight into the molecular mechanisms related to the process of mammalian ovary development, we performed a comparative transcriptomic analysis on ovaries isolated from infant and adult mice by using next-generation sequencing technology (SOLiD). We identified 15,454 and 16,646 transcriptionally active genes at the infant and adult stage, respectively. Among these genes, we also identified 7021 differentially expressed genes. Our analysis suggests that, in general, the adult ovary has a higher level of transcriptomic activity. However, it appears that genes related to primordial follicle development, such as those encoding Figla and Nobox, are more active in the infant ovary, whereas expression of genes vital for follicle development, such as Gdf9, Bmp4and Bmp15,is upregulated in the adult. These data suggest a dynamic shift in gene expression during ovary development and it is apparent that these changes function to facilitate follicle maturation, when additional functional gene studies are considered. Furthermore, our investigation has also revealed several important functional pathways, such as apoptosis, MAPK and steroid biosynthesis, that appear to be much more active in the adult ovary compared to those of the infant. These findings will provide a solid foundation for future studies on ovary development in mice and other mammals and help to expand our understanding of the complex molecular and cellular events that occur during postnatal ovary development.

Ovary development is a complex process involving numerous genes. A well-developed ovary is essential for females to keep fertility and reproduce offspring. In order to gain a better insight into the molecular mechanisms related to the process of mammalian ovary development, we performed a comparative transcriptomic analysis on ovaries isolated from infant and adult mice by using next-generation sequencing technology (SOLiD). We identified 15,454 and 16,646 transcriptionally active genes at the infant and adult stage, respectively. Among these genes, we also identified 7021 differentially expressed genes. Our analysis suggests that, in general, the adult ovary has a higher level of transcriptomic activity. However, it appears that genes related to primordial follicle development, such as those encoding Figla and Nobox, are more active in the infant ovary, whereas expression of genes vital for follicle development, such as Gdf9, Bmp4and Bmp15,is upregulated in the adult. These data suggest a dynamic shift in gene expression during ovary development and it is apparent that these changes function to facilitate follicle maturation, when additional functional gene studies are considered. Furthermore, our investigation has also revealed several important functional pathways, such as apoptosis, MAPK and steroid biosynthesis, that appear to be much more active in the adult ovary compared to those of the infant. These findings will provide a solid foundation for future studies on ovary development in mice and other mammals and help to expand our understanding of the complex molecular and cellular events that occur during postnatal ovary development.

Postnatal cerebral development is a complicated biological process precisely controlled by multiple genes. To understand the molecular mechanism of cerebral development, we compared dynamics of mouse cerebrum transcriptome through three developmental stages using high-throughput RNA-seq technique. Three libraries were generated from the mouse cerebrum at infancy, adolescence and adulthood, respectively. Consequently, 44,557,729 (infancy), 59,257,530 (adolescence) and 72,729,636 (adulthood) reads were produced, which were assembled into 15,344, 16,048 and 15,775 genes, respectively. We found that the overall gene expression level increased from infancy to adolescence and decreased later on upon reaching adulthood. The adolescence cerebrum has the most active gene expression, with expression of a large number of regulatory genes up-regulated and some crucial pathways activated. Transcription factor (TF) analysis suggested the similar dynamics as expression profiling, especially those TFs functioning in neurogenesis differentiation, oligodendrocyte lineage determination and circadian rhythm regulation. Moreover, our data revealed a drastic increase in myelin basic protein (MBP)-coding gene expression in adolescence and adulthood, suggesting that the brain myelin may be generated since mouse adolescence. In addition, differential gene expression analysis indicated the activation of rhythmic pathway, suggesting the function of rhythmic movement since adolescence; Furthermore, during infancy and adolescence periods, gene expression related to axon repulsion and attraction showed the opposite trends, indicating that axon repulsion was activated after birth, while axon attraction might be activated at the embryonic stage and declined during the postnatal development. Our results from the present study may shed light on the molecular mechanism underlying the postnatal development of the mammalian cerebrum.

Postnatal cerebral development is a complicated biological process precisely controlled by multiple genes. To understand the molecular mechanism of cerebral development, we compared dynamics of mouse cerebrum transcriptome through three developmental stages using high-throughput RNA-seq technique. Three libraries were generated from the mouse cerebrum at infancy, adolescence and adulthood, respectively. Consequently, 44,557,729 (infancy), 59,257,530 (adolescence) and 72,729,636 (adulthood) reads were produced, which were assembled into 15,344, 16,048 and 15,775 genes, respectively. We found that the overall gene expression level increased from infancy to adolescence and decreased later on upon reaching adulthood. The adolescence cerebrum has the most active gene expression, with expression of a large number of regulatory genes up-regulated and some crucial pathways activated. Transcription factor (TF) analysis suggested the similar dynamics as expression profiling, especially those TFs functioning in neurogenesis differentiation, oligodendrocyte lineage determination and circadian rhythm regulation. Moreover, our data revealed a drastic increase in myelin basic protein (MBP)-coding gene expression in adolescence and adulthood, suggesting that the brain myelin may be generated since mouse adolescence. In addition, differential gene expression analysis indicated the activation of rhythmic pathway, suggesting the function of rhythmic movement since adolescence; Furthermore, during infancy and adolescence periods, gene expression related to axonrepulsion and attraction showed the opposite trends, indicating that axon repulsion was activated after birth, while axon attraction might be activated at the embryonic stage and declined during the postnatal development. Our results from the present study may shed light on the molecular mechanism underlying the postnatal development of the mammalian cerebrum.

Myeloid leukemias are highly diverse diseases and have been shown to be associated with microRNA (miRNA) expression aberrations. The present study involved an in-depth miRNome analysis of two human acute myeloid leukemia (AML) cell lines, HL-60 and THP-1, and one human chronic myeloid leukemia (CML) cell line, K562, via massively parallel signature sequencing. mRNA expression profiles of these cell lines that were established previously in our lab facilitated an integrative analysis of miRNA and mRNA expression patterns. miRNA expression profiling followed by differential expression analysis and target prediction suggested numerous miRNA signatures in AML and CML cell lines. Some miRNAs may act as either tumor suppressors or oncomiRs in AML and CML by targeting key genes in AML and CML pathways. Expression patterns of cell type-specific miRNAs could partially reflect the characteristics of K562, HL-60 and THP-1 cell lines, such as actin filament-based processes, responsiveness to stimulus and phagocytic activity. miRNAs may also regulate myeloid differentiation, since they usually suppress differentiation regulators. Our study provides a resource to further investigate the employment of miRNAs in human leukemia subtyping, leukemogenesis and myeloid development. In addition, the distinctive miRNA signatures may be potential candidates for the clinical diagnosis, prognosis and treatment of myeloid leukemias.

Ribonucleic acid (RNA) deserves not only a dedicated field of biological research — a discipline or branch of knowledge — but also explicit definitions of its roles in cellular processes and molecular mechanisms. Ribogenomics is to study the biology of cellular RNAs, including their origin, biogenesis, structure and function. On the informational track, messenger RNAs (mRNAs) are the major component of ribogenomes, which encode proteins and serve as one of the four major components of the translation machinery and whose expression is regulated at multiple levels by other operational RNAs. On the operational track, there are several diverse types of RNAs — their length distribution is perhaps the most simplistic stratification — involving in major cellular activities, such as chromosomal structure and organization, DNA replication and repair, transcriptional/post-transcriptional regulation, RNA processing and routing, translation and cellular energy/metabolism regulation. An all-out effort exceeding the magnitude of the Human Genome Project is of essence to construct just mammalian transcriptomes in multiple contexts including embryonic development, circadian and seasonal rhythms, defined life-span stages, pathological conditions and anatomy-driven tissue/organ/cell types.

Myeloid leukemias are highly diverse diseases and have been shown to be associated with microRNA (miRNA) expression aberrations. The present study involved an in-depth miRNome analysis of two human acute myeloid leukemia (AML) cell lines, HL-60 and THP-1, and one human chronic myeloid leukemia (CML) cell line, K562, via massively parallel signature sequencing. mRNA expression profiles of these cell lines that were established previously in our lab facilitated an integrative analysis of miRNA and mRNA expression patterns. miRNA expression profiling followed by differential expression analysis and target prediction suggested numerous miRNA signatures in AML and CML cell lines. Some miRNAs may act as either tumor suppressors or oncomiRs in AML and CML by targeting key genes in AML and CML pathways. Expression patterns of cell type-specific miRNAs could partially reflect the characteristics of K562, HL-60 and THP-1 cell lines, such as actin filament-based processes, responsiveness to stimulus and phagocytic activity. miRNAs may also regulate myeloid differentiation, since they usually suppress differentiation regulators. Our study provides a resource to further investigate the employment of miRNAs in human leukemia subtyping, leukemogenesis and myeloid development. In addition, the distinctive miRNA signatures may be potential candidates for the clinical diagnosis, prognosis and treatment of myeloid leukemias.

Ribonucleic acid (RNA) deserves not only a dedicated field of biological research — a discipline or branch of knowledge — but also explicit definitions of its roles in cellular processes and molecular mechanisms. Ribogenomics is to study the biology of cellular RNAs, including their origin, biogenesis, structure and function. On the informational track, messenger RNAs (mRNAs) are the major component of ribogenomes, which encode proteins and serve as one of the four major components of the translation machinery and whose expression is regulated at multiple levels by other operational RNAs. On the operational track, there are several diverse types of RNAs — their length distribution is perhaps the most simplistic stratification — involving in major cellular activities, such as chromosomal structure and organization, DNA replication and repair, transcriptional/post-transcriptional regulation, RNA processing and routing, translation and cellular energy/metabolism regulation. An all-out effort exceeding the magnitude of the Human Genome Project is of essence to construct just mammalian transcriptomes in multiple contexts including embryonic development, circadian and seasonal rhythms, defined life-span stages, pathological conditions and anatomy-driven tissue/organ/cell types.

An organ unique to mammals, the mammary gland develops 90% of its mass after birth and experiences the pregnancylactation-involution cycle (PL cycle) during reproduction. To understand mammogenesis at the transcriptomic level and using a ribo-minus RNA-seq protocol, we acquired greater than 50 million reads each for the mouse mammary gland during pregnancy (day 12 of pregnancy), lactation (day 14 of lactation), and involution (day 7 of involution). The pregnancy-, lactation- and involution-related sequencing reads were assembled into 17344, 10160, and 13739 protein-coding transcripts and 1803, 828, and 1288 non-coding RNAs (ncRNAs), respectively. Differentially expressed genes (DEGs) were defined in the three samples, which comprised 4843 DEGs (749 up-regulated and 4094 down-regulated) from pregnancy to lactation and 4926 DEGs (4706 up-regulated and 220 down-regulated) from lactation to involution. Besides the obvious and substantive up- and down-regulation of the DEGs, we observe that lysosomal enzymes were highly expressed and that their expression coincided with milk secretion. Further analysis of transcription factors such as Trps1, Gtf2i, Tcf7l2, Nupr1, Vdr, Rb1, and Aebp1, and ncRNAs such as mir-125b, Let7, mir-146a, and mir-15has enabled us to identify key regulators in mammary gland development and the PL cycle.

Mammalian testis development is a complex and highly sophisticated process. To study the dynamic change of normal testis development at the transcriptional level, we investigated mouse testes at three postnatal ages: 6 days postnatal, 4 weeks old, and 10 weeks old, representing infant (PN1), juvenile (PN2), and adult (PN3) stages, respectively. Using ultra high-throughput RNA sequencing (RNA-seq) technology, we obtained 211 million reads with a length of 35 bp. We identified 18837 genes that were expressed in mouse testes, and found that genes expressed at the highest level were involved in spermatogenesis. The gene expression pattern in PN1 was distinct from that in PN2 and PN3, which indicates that spermatogenesis has commenced in PN2. We analyzed a large number of genes related to spermatogenesis and somatic development of the testis, which play important roles at different developmental stages. We also found that the MAPK, Hedgehog, and Wnt signaling pathways were significantly involved at different developmental stages. These findings further our understanding of the molecular mechanisms that regulate testis development. Our study also demonstrates significant advantages of RNA-seq technology for studying transcriptome during development.

Immortality and tumorigenicity are two distinct characteristics of cancers. Immortalization has been suggested to precede tumorigenesis. To understand the molecular mechanisms of tumorigenicity and cancer progression in mammary epithelium, we established a tumorigenic cell model by means of heavy-ion radiation of an immortal cell model, which was created by overexpressing the human telomerase reverse transcriptase (hTERT) in normal human mammary epithelial cells. We examined the expression profile of this tumorigenic cell line (T_hMEC) using the hTERT-overexpressing immortal cell line (I_hMEC) as a control. In-depth RNA-seq data was generated by using the next-generation sequencing (NGS) platform (Life Technologies SOLiD3). We found that house-keeping (HK) and tissue-specific (TS) genes were differentially regulated during the tumorigenic process. HK genes tended to be activated while TS genes tended to be repressed. In addition, the HK genes and TS genes tended to contribute differentially to the variation of gene expression at different RPKM (gene expression in reads per exon kilobase per million mapped sequence reads) levels. Based on transcriptome analysis of the two cell lines, we defined 7053 differentially-expressed genes (DEGs) between immortality and tumorigenicity. Differential expression of 20 manually-selected genes was further validated using qRT-PCR. Our observations may help to further our understanding of cellular mechanism(s) in the transition from immortalization to tumorigenesis.

Immortality and tumorigenicity are two distinct characteristics of cancers. Immortalization has been suggested to precede tumorigenesis. To understand the molecular mechanisms of tumorigenicity and cancer progression in mammary epithelium, we established a tumorigenic cell model by means of heavy-ion radiation of an immortal cell model, which was created by overexpressing the human telomerase reverse transcriptase (hTERT) in normal human mammary epithelial cells. We examined the expression profile of this tumorigenic cell line (T_hMEC) using the hTERT-overexpressing immortal cell line (I_hMEC) as a control. In-depth RNA-seq data was generated by using the next-generation sequencing (NGS) platform (Life Technologies SOLiD3). We found that house-keeping (HK) and tissue-specific (TS) genes were differentially regulated during the tumorigenic process. HK genes tended to be activated while TS genes tended to be repressed. In addition, the HK genes and TS genes tended to contribute differentially to the variation of gene expression at different RPKM (gene expression in reads per exon kilobase per million mapped sequence reads) levels. Based on transcriptome analysis of the two cell lines, we defined 7053 differentially-expressed genes (DEGs) between immortality and tumorigenicity. Differential expression of 20 manually-selected genes was further validated using qRT-PCR. Our observations may help to further our understanding of cellular mechanism(s) in the transition from immortalization to tumorigenesis.

Although strand-biased gene distribution (SGD) was described some two decades ago, the underlying molecular mechanisms and their relationship remain elusive. Its facets include, but are not limited to, the degree of biases, the strand-preference of genes, and the influence of background nucleotide composition variations. Using a dataset composed of 364 non-redundant bacterial genomes, we sought to illustrate our current understanding of SGD. First, when we divided the collection of bacterial genomes into non-polCand polCgroups according to their possession of DnaE isoforms that correlate closely with taxonomy, the SGD of the polCgroup stood out more significantly than that of the non-polCgroup. Second, when examining horizontal gene transfer, coupled with gene functional conservation (essentiality) and expressivity (level of expression), we realized that they all contributed to SGD. Third, we further demonstrated a weaker G-dominance on the leading strand of the non-polCgroup but strong purine dominance (both G and A) on the leading strand of the polCgroup. We propose that strand-biased nucleotide composition plays a decisive role for SGD since the polC-bearing genomes are not only AT-rich but also have pronounced purine-rich leading strands, and we believe that a special mutation spectrum that leads to a strong purine asymmetry and a strong strand-biased nucleotide composition coupled with functional selections for genes and their functions are both at work.

Although strand-biased gene distribution (SGD) was described some two decades ago, the underlying molecular mechanisms and their relationship remain elusive. Its facets include, but are not limited to, the degree of biases, the strand-preference of genes, and the influence of background nucleotide composition variations. Using a dataset composed of 364 non-redundant bacterial genomes, we sought to illustrate our current understanding of SGD. First, when we divided the collection of bacterial genomes into non-polCand polCgroups according to their possession of DnaE isoforms that correlate closely with taxonomy, the SGD of the polCgroup stood out more significantly than that of the non-polCgroup. Second, when examining horizontal gene transfer, coupled with gene functional conservation (essentiality) and expressivity (level of expression), we realized that they all contributed to SGD. Third, we further demonstrated a weaker G-dominance on the leading strand of the non-polCgroup but strong purine dominance (both G and A) on the leading strand of the polCgroup. We propose that strand-biased nucleotide composition plays a decisive role for SGD since the polC-bearing genomes are not only AT-rich but also have pronounced purine-rich leading strands, and we believe that a special mutation spectrum that leads to a strong purine asymmetry and a strong strand-biased nucleotide composition coupled with functional selections for genes and their functions are both at work.

Histone H3 lysine 4 trimethylation (H3K4me3) is well known to occur in the promoter region of genes for transcription activation. However, when investigating the H3K4me3 profiles in the mouse cerebrum and testis, we discovered that H3K4me3 also has a significant enrichment at the 3′ end of actively transcribed (sense) genes, named as 3′-H3K4me3. 3′-H3K4me3 is associated with ∼15% of protein-coding genes in both tissues. In addition, we examined the transcriptional initiation signals including RNA polymerase II (RNAPII) binding sites and 5′-CAGE-tag that marks transcriptional start sites. Interestingly, we found that 3′-H3K4me3 is associated with the initiation of antisense transcription. Furthermore, 3′-H3K4me3 modification levels correlate positively with the antisense expression levels of the associated sense genes, implying that 3′-H3K4me3 is involved in the activation of antisense transcription. Taken together, our findings suggest that H3K4me3 may be involved in the regulation of antisense transcription that initiates from the 3′ end of sense genes. In addition, a positive correlation was also observed between the expression of antisense and the associated sense genes with 3′-H3K4me3 modification. More importantly, we observed the 3′-H3K4me3 enrichment among genes in human, fruitfly and Arabidopsis, and found that the sequences of 3′-H3K4me3-marked regions are highly conserved and essentially indistinguishable from known promoters in vertebrate. Therefore, we speculate that these 3′-H3K4me3-marked regions may serve as potential promoters for antisense transcription and 3′-H3K4me3 appear to be a universal epigenetic feature in eukaryotes. Our results provide a novel insight into the epigenetic roles of H3K4me3 and the regulatory mechanism of antisense transcription.

The global features of H3K4 and H3K27 trimethylations (H3K4me3 and H3K27me3) have been well studied in recent years, but most of these studies were performed in mammalian cell lines. In this work, we generated the genome-wide maps of H3K4me3 and H3K27me3 of mouse cerebrum and testis using ChIP-seq and their high-coverage transcriptomes using ribominus RNA-seq with SOLiD technology. We examined the global patterns of H3K4me3 and H3K27me3 in both tissues and found that modifications are closely-associated with tissue-specific expression, function and development. Moreover, we revealed that H3K4me3 and H3K27me3 rarely occur in silent genes, which contradicts the findings in previous studies. Finally, we observed that bivalent domains, with both H3K4me3 and H3K27me3, existed ubiquitously in both tissues and demonstrated an invariable preference for the regulation of developmentally-related genes. However, the bivalent domains tend towards a “winner-takes-all” approach to regulate the expression of associated genes. We also verified the above results in mouse ES cells. As expected, the results in ES cells are consistent with those in cerebrum and testis. In conclusion, we present two very important findings. One is that H3K4me3 and H3K27me3 rarely occur in silent genes. The other is that bivalent domains may adopt a “winner-takes-all” principle to regulate gene expression.

Histone H3 lysine 4 trimethylation (H3K4me3) is well known to occur in the promoter region of genes for transcription activation. However, when investigating the H3K4me3 profiles in the mouse cerebrum and testis, we discovered that H3K4me3 also has a significant enrichment at the 3′ end of actively transcribed (sense) genes, named as 3′-H3K4me3. 3′-H3K4me3 is associated with ∼15% of protein-coding genes in both tissues. In addition, we examined the transcriptional initiation signals including RNA polymerase II (RNAPII) binding sites and 5′-CAGE-tag that marks transcriptional start sites. Interestingly, we found that 3′-H3K4me3 is associated with the initiation of antisense transcription. Furthermore, 3′-H3K4me3 modification levels correlate positively with the antisense expression levels of the associated sense genes, implying that 3′-H3K4me3 is involved in the activation of antisense transcription. Taken together, our findings suggest that H3K4me3 may be involved in the regulation of antisense transcription that initiates from the 3′ end of sense genes. In addition, a positive correlation was also observed between the expression of antisense and the associated sense genes with 3′-H3K4me3 modification. More importantly, we observed the 3′-H3K4me3 enrichment among genes in human, fruitfly and Arabidopsis, and found that the sequences of 3′-H3K4me3-marked regions are highly conserved and essentially indistinguishable from known promoters in vertebrate. Therefore, we speculate that these 3′-H3K4me3-marked regions may serve as potential promoters for antisense transcription and 3′-H3K4me3 appear to be a universal epigenetic feature in eukaryotes. Our results provide a novel insight into the epigenetic roles of H3K4me3 and the regulatory mechanism of antisense transcription.

The global features of H3K4 and H3K27 trimethylations (H3K4me3 and H3K27me3) have been well studied in recent years, but most of these studies were performed in mammalian cell lines. In this work, we generated the genome-wide maps of H3K4me3 and H3K27me3 of mouse cerebrum and testis using ChIP-seq and their high-coverage transcriptomes using ribominus RNA-seq with SOLiD technology. We examined the global patterns of H3K4me3 and H3K27me3 in both tissues and found that modifications are closely-associated with tissue-specific expression, function and development. Moreover, we revealed that H3K4me3 and H3K27me3 rarely occur in silent genes, which contradicts the findings in previous studies. Finally, we observed that bivalent domains, with both H3K4me3 and H3K27me3, existed ubiquitously in both tissues and demonstrated an invariable preference for the regulation of developmentally-related genes. However, the bivalent domains tend towards a “winner-takes-all” approach to regulate the expression of associated genes. We also verified the above results in mouse ES cells. As expected, the results in ES cells are consistent with those in cerebrum and testis. In conclusion, we present two very important findings. One is that H3K4me3 and H3K27me3 rarely occur in silent genes. The other is that bivalent domains may adopt a “winner-takes-all” principle to regulate gene expression.