Transcriptional Orientation Research Articles

Genomic and transcriptomic insights into the cytochrome P450 monooxygenase gene repertoire in the rice pest brown planthopper, Nilaparvata lugens

The cytochrome P450 monooxygenase (P450) gene family is one of the most abundant eukaryotic gene families that encode detoxification enzymes. In this study, we identified an abundance of P450 gene repertoire through genome- and transcriptome-wide analysis in the brown planthopper (Nilaparvata lugens), the most destructive rice pest in Asia. Detailed gene information including the exon–intron organization, size, transcription orientation and distribution in the genome revealed that many P450 loci were closely situated on the same scaffold, indicating frequent occurrence of gene duplications. Insecticide-response expression profiling revealed that imidacloprid significantly increased NlCYP6CS1v2, NLCYP4CE1v2, NlCYP4DE1, NlCYP417A1v2 and NlCYP439A1 expression; while triazophos and deltamethrin notably enhanced NlCYP303A1 expression. Expression analysis at the developmental stage showed the egg-, nymph-, male- and female-specific expression patterns of N. lugens P450 genes. These novel findings will be helpful for clarifying the P450 functions in physiological processes including development, reproduction and insecticide resistance in this insect species.

Phylogenetic analysis of the complete mitogenome sequence of the raspberry weevil, Aegorhinus superciliosus (Coleoptera: Curculionidae), supports monophyly of the tribe Aterpini

The superfamily Curculionoidea is one of the most diverse groups of insects in the world, including many species which are crop pests. Within this group, the native raspberry weevil, Aegorhinus superciliosus (Guérin, 1830), is an important pest in blueberry and raspberry fields in southern South America. Using a 454 sequencing approach, we sequenced and annotated the mitogenome of A. superciliosus, it, providing the first such information for any species in the tribe Aterpini, subfamily Cyclominae. The assembled mitogenome is a circular DNA molecule 15,121bp in length containing all 37 genes normally found in metazoans. Mitogenome organization and transcriptional orientation in A. superciliosus showed the same pattern that characterizes the suborder Polyphaga. Bayesian and Maximum Likelihood phylogenetic analyses supported the monophyly of the tribe Aterpini and the subfamily Cyclominae, recovering this clade in a sister group relationship with Entiminae and Hyperinae. The monophyly of these three subfamilies defines a critical transition to an ectophagous lifestyle in the larvae, from an ancestrally endophagous larval lifestyle in all other lineages. The sequenced mitogenome of A. superciliosus can provide basic data for future studies investigating population history, molecular systematics, stress ecophysiology and phylogeography.

Immunoglobulin genomics in the prairie vole (Microtus ochrogaster)

In science, the prairie voles are ideal models for studying the regulatory mechanisms of social behavior in humans. The utility of the prairie vole as a biology model can be further enhanced by characterization of the genes encoding components of the immune system. Here, we report the genomic organization of the prairie vole immunoglobulin heavy and light chain genes. The prairie vole IgH locus on chromosome 1 spans over 1600kb, and consists of at least 79 VH segments (28 potentially functional genes, 2 ORFs and 49 pseudogenes), 7 DH segments, 4 JH segments, four constant region genes (μ, γ, ɛ, and α), and two transmembrane regions of δ gene. The Igκ locus, found on three scaffolds (JH996430, JH996605 and JH996566), contains a totle of 124 Vκ segments (47 potentially functional genes, 1 ORF and 76 pseudogenes), 5 Jκ segments and a single Cκ gene. Two different transcriptional orientations were determined for these Vκ gene segments. In contrast, the Igλ locus on scaffold JH996473 and JH996489 includes 21 Vλ gene segments (14 potentially functional genes, 1 ORF and 6 pseudogenes), all with the same transcriptional polarity as the downstream Jλ-Cλ cluster. Phylogenetic analysis and sequence alignments suggested the prairie vole's large germline VH, Vκ and Vλ gene segments appear to form limited gene families. Therefore, this species may generate antibody diversity via a gene conversion-like mechanism associated with its pseudogene reserves.

26. HIV-1 Mediated Insertional Mutagenesis Increase the Persistence of Infected T Cells in Patients Under ART by Triggering Their Differentiation Into Long Lived T-Regulatory and T-Central Memory Cells

It has been recently suggested that HIV-1 by integrating near cancer-associated genes could promote the expansion and persistence of infected cells in patients under Anti Retroviral Therapy (ART). However, the molecular mechanism/s of insertional mutagenesis used and the physiological impact on the cells harboring these integrations are completely unknown.Here, we found that in the peripheral blood mononuclear cells from 54 HIV-1 infected patients under ART, BACH2 and STAT5B were targeted by a significant number of integrations compared to other lentiviral integration datasets (p<0.0001) and with the same orientation of gene transcription. Furthermore, we observed that in the blood of 34% of HIV-1 patients under ART (30/87) there are cell clones expressing aberrant chimeric transcripts containing viral sequences fused to the first protein coding exon of BACH2 or STAT5B and predicted to encode for unaltered full-length proteins. However, it is unlikely that the ectopic expression of these transcription factors in T cells per se would trigger cell transformation. Indeed, accordingly with their roles in the regulation of T-cell identity and homeostasis, forced expression of STAT5B and BACH2 in naive CD4+ T-cells significantly skewed their differentiation towards functional T-regulatory cells with a marked immunosuppressive potential. Moreover, STAT5B ectopic expression promoted the IL-2 independent proliferation of T-regulatory cells that was however blocked by TGFB treatment, indicating that these cells are able to respond to external differentiating cues.Importantly, tracking the expression of HIV-1/STAT5B transcripts in T cell subpopulations and monocytes purified from the blood of patients under ART, we found that, in all patients tested (N=6), chimeric mRNAs were present only in T-regulatory and T-central memory cells but not in CD8+ T-cells, T naive, T stem cell memory, T effector memory nor monocytes.Our findings provide novel evidence that HIV-1 takes advantage of insertional mutagenesis to favor its persistence in the host by activating STAT5B and possibly BACH2. However, the selective advantage conferred by these integrations does not involve T-cell transformation but rather their differentiation into functional T-regulatory and T-central memory cells which are long lived, potentially able to diminish the immune surveillance against infected cells thus favoring the escape from the immune system and long-term viral persistence.Hence, new targeted therapies aimed at interfering with BACH2 and STAT5B regulated pathways could provide the general means for the immunological re-sensitization towards HIV-1 infected cells to reduce long lived cellular reservoirs and the eradication of the viral infection in HIV-1 patients.

Proteogenomics analysis reveals specific genomic orientations of distal regulatory regions composed by non-canonical histone variants.

BackgroundHistone variants play further important roles in DNA packaging and controlling gene expression. However, our understanding about their composition and their functions is limited.ResultsIntegrating proteomic and genomic approaches, we performed a comprehensive analysis of the epigenetic landscapes containing the four histone variants H3.1, H3.3, H2A.Z, and macroH2A. These histones were FLAG-tagged in HeLa cells and purified using chromatin immunoprecipitation (ChIP). By adopting ChIP followed by mass spectrometry (ChIP-MS), we quantified histone post-translational modifications (PTMs) and histone variant nucleosomal ratios in highly purified mononucleosomes. Subsequent ChIP followed by next-generation sequencing (ChIP-seq) was used to map the genome-wide localization of the analyzed histone variants and define their chromatin domains. Finally, we included in our study large datasets contained in the ENCODE database. We newly identified a group of regulatory regions enriched in H3.1 and the histone variant associated with repressive marks macroH2A. Systematic analysis identified both symmetric and asymmetric patterns of histone variant occupancies at intergenic regulatory regions. Strikingly, these directional patterns were associated with RNA polymerase II (PolII). These asymmetric patterns correlated with the enhancer activities measured using global run-on sequencing (GRO-seq) data.ConclusionsOur studies show that H2A.Z and H3.3 delineate the orientation of transcription at enhancers as observed at promoters. We also showed that enhancers with skewed histone variant patterns well facilitate enhancer activity. Collectively, our study indicates that histone variants are deposited at regulatory regions to assist gene regulation.Electronic supplementary materialThe online version of this article (doi:10.1186/s13072-015-0005-9) contains supplementary material, which is available to authorized users.

Effect of orientation of transcription of a gene in an inverted transferred DNA repeat on transcriptional gene silencing in rice transgenics-a case study.

We studied transgene silencing in two transgenic rice plants, OSM25 and COT-OSM4, which harboured two different types of right border (RB)-centered inverted transferred DNA (T-DNA) repeats (IRs). The T-DNA in OSM25 has three genes gus, OSM and hph, all under the transcriptional control of the Cauliflower mosaic virus 35S promoter (P35S). The gus gene, which is proximal to the RB, is in a convergent orientation of transcription in the IR. OSM25 displayed silencing of all three transgenes. Nuclear run-on transcription analysis revealed that silencing of gus, OSM and hph genes in OSM25 operates at the transcriptional level. P35S showed hypermethylation in OSM25 plants. COT-OSM4 has P35S-driven gus and hph genes in its T-DNA. The hph gene, which is proximal to the RB, is in a divergent orientation of transcription in the IR. Unlike in OSM25, the transgenes in COT-OSM4 showed no silencing. These findings show that convergent orientation of transcription of a gene at the origin of an IR is important for transgene silencing.

Complete mitochodrial genome of Vanmanenia pingchowensis (Cypriniformes, Cyprinidae)

The complete mitochondrial genome of Vanmanenia pingchowensis was determined in this study. The gene composition, arrangement and transcriptional orientation in V. pingchowensis mitogenome were identical to most vertebrates. The complete mitogenome of V. pingchowensis was 15,560 bp in size with 13 protein coding genes, 22 tRNA genes, and a control region. Two start codon patterns and three stop codon patterns were found in protein-coding genes. Only the tRNA-Ser2 could not fold into a typical clover-leaf secondary structure for lacking the dihydrouridine arm. Sequences alignment results suggest that the complete mitogenome of V. pingchowensis is an efficient tool to study molecular phylogenetics, biogeography and adaptive evolution of this lineage.

Complete mitochodrial genome of Bangana decora (Cypriniformes, Cyprinidae)

The complete mitochondrial genome of Bangana decora was determined in this study. The gene composition, arrangement and transcriptional orientation in B. decora mitogenome were identical to most vertebrates. The complete mitogenome of B. decora was 16,607 bp in size with 13 protein-coding genes, 22 tRNA genes and a control region. Two start codon patterns and three stop codon patterns were found in protein-coding genes. Only the tRNA-Ser2 could not fold into a typical clover-leaf secondary structure due to the absence of the dihydrouridine arm. Sequence alignment results suggest that the complete mitogenome of B. decora is an efficient tool to study molecular phylogenetics, biogeography and adaptive evolution of this lineage.

Complete mitochodrial genome of Parasinilabeo assimilis (Cypriniformes, Cyprinidae)

The complete mitochondrial genome of Parasinilabeo assimilis has been determined in this study. The gene composition, arrangement and transcriptional orientation in P. assimilis mitogenome were identical to most vertebrates. The complete mitogenome of P. assimilis was 16,602 bp in size with 13 protein coding genes, 22 tRNA genes and a control region. Two start codon patterns and three stop codon patterns were found in protein-coding genes. Only the tRNA-Ser2 could not fold into a typical clover-leaf secondary structure due to the lack of the dihydrouridine arm. Sequence alignment results suggest that the complete mitogenome of P. assimilis is an efficient tool to study molecular phylogenetics, biogeography and adaptive evolution of this lineage.

Complete mitochondrial genome of the Freshwater Whipray Himantura dalyensis

The complete mitochondrial genome of the Freshwater Whipray Himantura dalyensis is presented in this study. It is 17,693 bp in length and contains 37 genes in typical gene order and transcriptional orientation observed in vertebrates. There were a total of 86 bp short intergenic spacers and 22 bp overlaps in the genome. The overall base composition was 31.4% A, 25.5% C, 13.2% G and 29.9% T. Two start codons (GTG and ATG) and two stop codons (TAG and TAA/T) were found in 13 protein-coding genes. The length of 22 tRNA genes ranged from 68 (tRNA-Cys and tRNA-Ser2) to 75 bp (tRNA-Leu1). The origin of L-strand replication (OL) was found between the tRNA-Asn and tRNA-Cys genes. The base composition of the control region (1940 bp) was similar to the whole mitogenome.

Complete mitochodrial genome of Elopichthys bambusa (Cypriniformes, Cyprinidae)

The complete mitochondrial genome of Elopichthys bambusa was determined in this study. The gene composition, arrangement and transcriptional orientation in E. bambusa mitogenome were identical to most vertebrates. Two start codon patterns (ATG and GTG) and three stop codon patterns (TAG, TAA and T) were found in protein-coding genes. Only the tRNA-Ser2 could not fold into a typical clover-leaf secondary structure for lacking the dihydrouridine arm. Sequences alignment results suggest that the complete mitogenome of E. bambusa is an efficient tool to check species identification by comparing different gene sequences.

Annotation and sequence diversity of transposable elements in common bean (Phaseolus vulgaris)

Common bean (Phaseolus vulgaris) is an important legume crop grown and consumed worldwide. With the availability of the common bean genome sequence, the next challenge is to annotate the genome and characterize functional DNA elements. Transposable elements (TEs) are the most abundant component of plant genomes and can dramatically affect genome evolution and genetic variation. Thus, it is pivotal to identify TEs in the common bean genome. In this study, we performed a genome-wide transposon annotation in common bean using a combination of homology and sequence structure-based methods. We developed a 2.12-Mb transposon database which includes 791 representative transposon sequences and is available upon request or from www.phytozome.org. Of note, nearly all transposons in the database are previously unrecognized TEs. More than 5,000 transposon-related expressed sequence tags (ESTs) were detected which indicates that some transposons may be transcriptionally active. Two Ty1-copia retrotransposon families were found to encode the envelope-like protein which has rarely been identified in plant genomes. Also, we identified an extra open reading frame (ORF) termed ORF2 from 15 Ty3-gypsy families that was located between the ORF encoding the retrotransposase and the 3′LTR. The ORF2 was in opposite transcriptional orientation to retrotransposase. Sequence homology searches and phylogenetic analysis suggested that the ORF2 may have an ancient origin, but its function is not clear. These transposon data provide a useful resource for understanding the genome organization and evolution and may be used to identify active TEs for developing transposon-tagging system in common bean and other related genomes.

Complete mitochondrial genome of Anguilla japonica (Anguilliformes, Anguillidae)

The complete mitochondrial genome of Anguilla japonica have been determined in this study. The gene composition, arrangement and transcriptional orientation in A. japonica mitogenome were identical to those of most vertebrates. The complete mitogenome of A. japonica was 16,615 bp in size wih 13 protein-coding genes, 22 tRNA genes, 2 rRNA genes and a control region. Two start codon patterns and three stop codon patterns were found in protein-coding genes. Only the tRNA-Ser2 could not fold into a typical clover-leaf secondary structure for lacking the dihydrouridine arm. Sequences alignment results suggest that the complete mitogenome of A. japonica is an efficient tool to study molecular phylogenetics, biogeography and adaptive evolution of this lineage.

Complete mitochondrial genome of the hardnose shark Carcharhinus macloti (Carcharhiniformes: Carcharhinidae)

The complete mitochondrial genome of Carcharhinus macloti was determined in this study. It is 16,701 bp in length and contains 37 genes with the typical gene order and transcriptional orientation in vertebrates. A total of 29 bp overlaps and 29 bp short intergenic spaces located in 22 gene junctions. The overall base composition is 31.6% A, 26.2% C, 13.0% G and 29.2% T. Two start codons (ATG and GTG) and three stop codons (AGG, TAG and TAA/T) were found in 13 protein-coding genes. The length of 22 tRNA genes ranged from 66 bp (tRNA-Ser2) to 75 bp (tRNA-Leu1). The tRNA-Ser2 (GCU) lacks the dihydrouridine arm by a simple loop and can not be folded into the typical cloverleaf structure. The control region is 1066 bp in length with high A+T content (68.2%).

Complete mitochondrial genome of the whitetip reef shark Triaenodon obesus (Carcharhiniformes: Carcharhinidae)

The complete mitochondrial genome of the whitetip reef shark Triaenodon obesus is determined in this study. It is 16,700 bp in length, with the typical gene composition, arrangement and transcriptional orientation in vertebrates. The overall base composition is 31.4% A, 25.8% C, 13.2% G and 29.7% T. Two start codons and two stop codons are found in the protein-coding genes. The 22 tRNA genes ranged from 67 to 75 nucleotides. The tRNA-Ser2 lost the DHU arm and could not be folded to the typical cloverleaf secondary structure. The origin of L-strand replication (OL) sequence was identified between tRNA-Asn and tRNA-Cys genes. The high A+T content of control region is due to a lot of poly A and poly T.

Genomic insights into the serine protease gene family and expression profile analysis in the planthopper, Nilaparvata lugens.

BackgroundThe brown planthopper (Nilaparvata lugens) is one of the most destructive rice plant pests in Asia. N. lugens causes extensive damage to rice by sucking rice phloem sap, which results in hopper burn (complete death of the rice plants). Despite its importance, little is known about the digestion, development and defense mechanisms of this hemimetabolous insect pest. In this study, we aim to identify the serine protease (SP) and serine protease homolog (SPH) genes, which form a large family in eukaryotes, due to the potential for multiple physiological roles. Having a fully sequenced genome for N. lugens allows us to perform in-depth analysis of the gene structures, reveal the evolutionary relationships and predict the physiological functions of SP genes.ResultsThe genome- and transcriptome-wide analysis identified 90 putative SP (65) and SPH (25) genes in N. lugens. Detailed gene information regarding the exon-intron organization, size, distribution and transcription orientation in the genome revealed that many SP/SPH loci are closely situated on the same scaffold, indicating the frequent occurrence of gene duplications in this large gene family. The gene expression profiles revealed new findings with regard to how SPs/SPHs respond to bacterial infections as well as their tissue-, development- and sex-specific expressions.ConclusionsOur findings provide comprehensive gene sequence resources and expression profiles of the N. lugens SP and SPH genes, which give insights into clarifying the potentially functional roles of these genes in the biological processes including development, digestion, reproduction and immunity.Electronic supplementary materialThe online version of this article (doi:10.1186/1471-2164-15-507) contains supplementary material, which is available to authorized users.

Complete mitochondrial genome of Sinilabeo decorus tungting (Cypriniformes, Cyprinidae)

The complete mitochondrial genome of Sinilabeo decorus tungting was determined in this study. The gene composition, arrangement and transcriptional orientation in S. decorus tungting mitogenome were identical to most vertebrates. Two start codon patterns (ATG and GTG) and three stop codon patterns (TAG, TAA and T) were found in protein-coding genes. Only the tRNA-Ser2 could not fold into a typical clover-leaf secondary structure for lacking the dihydrouridine arm. Sequences alignment results suggest that the complete mitogenome of S. decorus tungting is an efficient tool to check species identification by comparing different gene sequences.

A comprehensive phylogenetic and structural analysis of the carcinoembryonic antigen (CEA) gene family.

The carcinoembryonic antigen (CEA) gene family belongs to the immunoglobulin (Ig) superfamily and codes for a vast number of glycoproteins that differ greatly both in amino acid composition and function. The CEA family is divided into two groups, the carcinoembryonic antigen-related cell adhesion molecules (CEACAMs) and the pregnancy-specific glycoproteins. The CEA family members are implicated in pleiotropic (patho)physiological functions including cell–cell adhesion, pregnancy, immunity, neovascularization, regulation of insulin homeostasis, and carcinogenesis. In general, the CEA-encoded proteins are composed of an extracellular region with Ig variable and constant-like domains and a cytoplasmic region containing signaling motifs. Of particular interest, the well-studied human and mouse CEA genes are arranged in clusters in a single chromosome. Taking into account this characteristic, we made an effort to reconstruct the evolutionary history of the CEA gene family. Toward this end, the publicly available genomes were searched extensively for CEA homologs. The domain organization of the retrieved protein sequences was analyzed, and, subsequently, comprehensive phylogenetic analyses of the entire length CEA homologous proteins were performed. A series of evolutionarily conserved amino acid residues, functionally important, were identified. The relative positioning of these residues on the modeled tertiary structure of novel CEA protein domains revealed that they are, also, spatially conserved. Furthermore, the chromosomal arrangement of CEA genes was examined, and it was found that the CEA genes are preserved in terms of position, transcriptional orientation, and number in all species under investigation.

Complete mitochondrial genome of milk sole fish Aseraggodes kobensis (Pleuronectiformes: Soleidae)

The complete mitochondrial genome of the milk sole fish Aseraggodes kobensis was determined in this study. It is 16,944 bp in length, contains 13 protein-coding genes, 22 tRNA genes, 2 rRNA genes and one putative control region with the typical gene order and transcriptional orientation in vertebrates. The nucleotide base content of A. kobensis mitochondrial genome is: 31.7% A, 27.3% C, 15.3% G and 25.8% T. There are 24 bp overlaps and 48 bp short intergenic spaces located in 6 and 12 gene junctions, respectively. Two start codons (ATG and GTG) and two stop codons (TAG and TAA/T) were found in protein-coding genes. The origin of L-strand replication sequence was found between the tRNAAsn and tRNACys genes, and could be folded into a hairpin structure. The control region is 1239 bp in length with the rich A+T (66.4%) and poor G (13.1%) content.

Genomic characteristics of the T cell receptor (TRB) locus in the rabbit (Oryctolagus cuniculus) revealed by comparative and phylogenetic analyses

The present study identifies the genomic structure and the gene content of the T cell receptor beta (TRB) locus in the Oryctolagus cuniculus whole genome assembly. The rabbit locus spans less than 600Kb and the general genomic organization is highly conserved with respect to other mammalian species. A pool of 74 TRB variable (TRBV) genes distributed in 24 subgroups are located upstream of two in tandem-aligned D-J-C gene clusters, each composed of one TRBD, six TRBJ genes, and one TRBC gene, followed by a single TRBV gene with an inverted transcriptional orientation. All TRB genes (functional, ORF, pseudogenes) of this paper have been approved by the IMGT/WHO-IUIS nomenclature committee. Additionally, five potentially functional protease serine (PRSS) trypsinogen or trypsinogen-like genes were identified: two in tandem PRSS-like genes, followed by two PRSS genes with unique traits, lie downstream of the TRBV1 gene and one PRSS gene is located about 400Kb away downstream of the TRBV genes. Comparative and phylogenetic analyses revealed that multiple duplication events within a few subgroups have generated the germline repertoire of the rabbit TRBV genes, which is substantially larger than those described in humans, mice, and dogs, suggesting that a strong evolutionary pressure has selected the development of a species-specific TRBV repertoire. Hence, the genomic organization of the TRB locus in the genomes appears to be the result of a balance between the maintenance of a core-number of genes essential for the immunological performances and the requirement of newly arisen genes.