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Background: Stage IV colorectal cancer patients with liver metastasis represent a special group of CRC patients with poor prognosis. The prognostic factors have not been investigated for stage IV CRC patients undergoing primary cancer resection but not candidates for metastasis resection. Methods: Ninety-nine stage IV CRC patients who underwent primary cancer resection without metastasis resection were retrospectively recruited. Both whole-exome sequencing (WES) and RNA-seq were performed with frozen primary cancer tissues, using para-cancerous normal tissues as the control. Valid data were obtained from 78 patients for WES and 84 patients for RNA-seq. Univariate, multivariate Cox analyses were performed and Nomogram model was established to predict patient prognosis. Results: The correlation between patient prognosis and clinicopathological factors, mutational status, or mRNA level changes was examined. Univariate (p = 0.0007) and subsequent multivariate analyses on clinicopathological factors showed that location (left or right) was the only independent risk factor for patient prognosis (HR = 3.63; 95% CI: 1.56-8.40, p = 0.003), while T, N, M staging, gender, race, location (rectum or colon), and pathological types were not stratifying factors. The mutational status of APC, TP53, KRAS, TTN, SYNE1, SMAD4, PIK3CA, RYR2, and BRAF did not show significant stratification in patient prognosis. RNA-seq showed that genes related to membrane function, ion channels, transporters, or receptors were among those with significant mRNA level alterations. Univariate analysis identified 97 genes with significantly altered mRNA levels, while NEUROD1, FGF18, SFTA2, PLAC1, SAA2, DSCAML1, and OTOP3 were significant in multivariate analysis. A risk model was established to stratify the prognosis of stage IV CRC patients. A Nomogram model was established with these genes to predict individual patient prognosis. Conclusions: A panel of eight genes with significant mRNA level alterations was capable of predicting the prognosis and risk of the specific patient group. Future prospective study is needed to validate the model. [ABSTRACT FROM AUTHOR]

Aflatoxin B1 (AFB1) is a food contaminant metabolized mostly in the liver and leading to hepatic damage. Livestock species are differently susceptible to AFB1, but the underlying mechanisms of toxicity have not yet been fully investigated, especially in ruminants. Thus, the aim of the present study was to better characterize the molecular mechanism by which AFB1 exerts hepatotoxicity in cattle. The bovine fetal hepatocyte cell line (BFH12) was exposed for 48 h to three different AFB1 concentrations (0.9 µM, 1.8 µM and 3.6 µM). Whole-transcriptomic changes were measured by RNA-seq analysis, showing significant differences in the expression of genes mainly involved in inflammatory response, oxidative stress, drug metabolism, apoptosis and cancer. As a confirmatory step, post-translational investigations on genes of interest were implemented. Cell death associated with necrosis rather than apoptosis events was noted. As far as the toxicity mechanism is concerned, a molecular pathway linking inflammatory response and oxidative stress was postulated. Toll-Like Receptor 2 (TLR2) activation, consequent to AFB1 exposure, triggers an intracellular signaling cascade involving a kinase (p38β MAPK), which in turn allows the nuclear translocation of the activator protein-1 (AP-1) and NF-κB, finally leading to the release of pro-inflammatory cytokines. Furthermore, a p38β MAPK negative role in cytoprotective genes regulation was postulated. Overall, our investigations improved the actual knowledge on the molecular effects of this worldwide relevant natural toxin in cattle. [ABSTRACT FROM AUTHOR]

Protein sentezi sürecinde meydana gelen farklılaşmaların metabolik hastalıklar, kanser gibi kompleks hastalıklara neden olduğu farklı çalışmalarda belirtilmiştir. Protein sentezindeki değişimlerin anlaşılması için proteinleri oluşturan genlerin belirlenmesi ve bu genlerin diğer genlerle ilişkilerin ortaya çıkarılması gerekmektedir. Yeni nesil dizileme teknikleriyle hastalıklara neden olan moleküler düzeyde ilişkilerin doğruluklu olarak belirlenmesi kolaylaşmıştır. Gen birlikte ifade edilme (GBİE) ağları düzenleyen-düzenleyici ilişkisi içermeden benzer biyolojik süreçlere katılan genler arasındaki ilişkileri araştırmacılara göstermektedir. Çalışmamızda RNA-Seq verileri kullanılarak prostat kanseriyle ilişkili GBİE ağları elde edilmiştir. RNA- Seq verileri farklı nükleotit uzunluğundaki genlerden ve farklı sayıda okumalar içeren örneklerden oluştuğu için normalizasyon teknikleri moleküler ilişki çıkarımında önem taşımaktadır. Çalışmamızda gen birlikte ifade edilme ağları ham veri ve farklı iki normalizasyon yaklaşımı olan M- Değerinin Kırpılmış Ortalaması (MDKO), Göreceli Log İfadesi (GLİ) hesaplamalarıyla ayrı ayrı oluşturulmuş veriler üzerinde çıkartılarak örtüşme analizi ve topolojik performans değerlendirilmesi yapılmıştır. Örtüşme analizine göre normalize edilmiş RNA- Seq verileri kullanarak elde edilmiş gen birlikte ifade edilme ağlarının ham verilere göre daha fazla literatürde bulunan ilişkileri tahmin ettiği gözlemlenmiştir. İki normalizasyon yöntemiyle elde edilen GBİE'lere ait örtüşme analizi performans metrikleri değerleri ise birbirlerine yakın çıkmıştır. Topolojik değerlendirme sonuçlara göre normalize edilmiş veriler üzerinde elde edilen GBİE ağlarının ölçeksiz ağ tanımına daha yakın olduğu gözlemlenmiştir. Çalışmamızda aynı zamanda ham ve normalize edilmiş veriler üzerinde GBİE ağ çıkarım algoritmaları olan C3NET, ARACNE ve WGCNA yaklaşımlarının performansları da karşılaştırılmıştır.

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Apoptosis of nucleus pulposus (NP) cells has an important role in the process of intervertebral disc degeneration (IDD), and the search for novel compounds to prevent apoptosis from occurring is urgently required. In the present study, syringic acid (SyrA) was found to exhibit no cytotoxicity on NP cells, and was able to reverse the cytotoxicity, as well as the abnormal expression of Bcl‑2 and caspase‑3, that were induced by lipopolysaccharide (LPS). The transcriptomes of each group were then analyzed using RNA‑Seq. A total of 65 differentially expressed genes (DEGs) were identified in LPS‑stimulated groups (LPS group vs. control group), 819 DEGs were identified in the SyrA‑reversed groups (SyrA plus LPS group vs. LPS group), and a further 25 DEGs were identified in the SyrA plus LPS group compared with the control group. Reverse transcription‑quantitative PCR validation indicated that the alterations in expression of uroplakin 3B‑like 1 (UPK3BL1), voltage‑dependent calcium channel subunit α‑2/δ‑1 (CACNA2D1) and polo‑like kinase 4 (PLK4) were consistent with the corresponding results of RNA‑Seq, and that these genes were involved in both LPS‑stimulation and SyrA‑reversion processes. Kyoto Encyclopedia of Genes and Genomes analyses indicated that the DEGs in SyrA‑reversed groups were involved in, amongst other pathways, ‘Autophagy‑other’ and ‘Apoptosis‑multiple species’. In conclusion, the addition of SyrA to the NP cells co‑incubated with LPS appeared to help prevent the abnormal expression of mRNAs and apoptosis that had been identified in NP cells incubated with LPS alone. The potential mechanism underlying the reversion of SyrA might be attributed to the regulation of CACNA2D1 and PLK4. [ABSTRACT FROM AUTHOR]

As a temperate-cold species, S accharina japonica often suffers heat stress when it is transplanted to temperate and subtropical zones. Study the heat stress response and resistance mechanism of S accharina is of great significance for understanding the acclimation to heat stress under domestication as well as for breeding new cultivars with heat stress resistance. In this study, we identified a set of heat stress-responsive miRNAs and analysed their regulation during the heat stress response. CO (control) and heat stress ( HS) sRNA libraries were constructed and sequenced. Forty-nine known miRNAs and 75 novel miRNAs were identified, of which seven known and 25 novel miRNAs were expressed differentially under heat stress. Quantitative PCR of six selected miRNAs confirmed that these loci were responsive to heat stress. Thirty-nine and 712 genes were predicted to be targeted by the seven known miRNAs and 25 novel miRNAs, respectively. Gene function and pathway analyses showed that these genes probably play important roles in S . japonica heat stress tolerance. The miRNAs identified represent the first set of heat-responsive miRNAs identified from S . japonica, and their identification can help elucidate the heat stress response and resistance mechanisms in S . japonica. [ABSTRACT FROM AUTHOR]

Dollar spot, caused by Sclerotinia homoeocarpa, is a prevalent turfgrass disease, and the fungus exhibits widespread fungicide resistance in North America. In a previous study, an ABC- G transporter, ShatrD, was associated with practical field resistance to demethylation inhibitor ( DMI) fungicides. Mining of ABC- G transporters, also known as pleiotropic drug resistance ( PDR) transporters, from RNA- Seq data gave an assortment of transcripts, several with high sequence similarity to functionally characterized transporters from Botrytis cinerea, and others with closest blastx hits from Aspergillus and Monilinia. In addition to ShatrD, another PDR transporter showed significant over-expression in replicated RNA- Seq data, and in a collection of field-resistant isolates, as measured by quantitative polymerase chain reaction. These isolates also showed reduced sensitivity to unrelated fungicide classes. Using a yeast complementation system, we sought to test the hypothesis that this PDR transporter effluxes DMI as well as chemically unrelated fungicides. The transporter ( ShPDR1) was cloned into the Gal1 expression vector and transformed into a yeast PDR transporter deletion mutant, AD12345678. Complementation assays indicated that ShPDR1 complemented the mutant in the presence of propiconazole ( DMI), iprodione (dicarboximide) and boscalid ( SDHI, succinate dehydrogenase inhibitor). Our results indicate that the over-expression of ShPDR1 is correlated with practical field resistance to DMI fungicides and reduced sensitivity to dicarboximide and SDHI fungicides. These findings highlight the potential for the eventual development of a multidrug resistance phenotype in this pathogen. In addition, this study presents a pipeline for the discovery and validation of fungicide resistance genes using de novo next-generation sequencing and molecular biology techniques in an unsequenced plant pathogenic fungus. [ABSTRACT FROM AUTHOR]

Xenotransplantation of porcine organs, tissues, and cells inherits a risk for xenozoonotic infections. Viable tissues and cells intended for transplantation have to be considered as potentially contaminated non-sterile products. The demands on microbial testing, based on the regulatory requirements, are often challenging due to a restricted shelf life or the complexity of the product itself. In Europe, the regulatory framework for xenogeneic cell therapy is based on the advanced therapy medicinal products ( ATMP) regulation (2007), the EMA CHMP Guideline on xenogeneic cell-based medicinal products (2009), as well as the WHO and Council of Europe recommendations. In the USA, FDA guidance for industry (2003) regulates the use of xenotransplants. To comply with the regulations, validated test methods need to be established that reveal the microbial status of a transplant within its given shelf life, complemented by strictly defined action alert limits and supported by breeding in specific pathogen-free ( SPF) facilities. In this review, we focus on assays for the detection of the porcine endogenous retroviruses PERV-A/-B/-C, which exhibit highly polymorphic proviral loci in pig genomes. PERVs are transmitted vertically and cannot be completely eliminated by breeding or gene knock out technology. PERVs entail a public health concern that will persist even if no evidence of PERV infection of xenotransplant recipients in vivo has been revealed yet. Nevertheless, infectious risks must be minimized by full assessment of pigs as donors by combining different molecular screening assays for sensitive and specific detection as well as a functional analysis of the infectivity of PERV including an adequate monitoring of recipients. [ABSTRACT FROM AUTHOR]

Differences in gene expression were studied after Plum pox virus ( PPV, sharka disease) infection in peach GF305 leaves with and without sharka symptoms using RNA-Seq. For each sample, more than 80% of 100-nucleotide paired-end ( PE) Illumina reads were aligned on the peach reference genome. In the symptomatic sample, a significant proportion of reads were mapped to PPV reference genomes (1.04% compared with 0.00002% in non-symptomatic leaves), allowing for the ultra-deep assembly of the complete genome of the PPV isolate used (9775 nucleotides, missing only 11 nucleotides at the 5′ genome end). In addition, significant alternative splicing events were detected in 359 genes and 12 990 single nucleotide polymorphisms ( SNPs) were identified, 425 of which could be annotated. Gene ontology annotation revealed that the high-ranking mRNA target genes associated with the expression of sharka symptoms are mainly related to the response to biotic stimuli, to lipid and carbohydrate metabolism and to the negative regulation of catalytic activity. A greater number of differentially expressed genes were observed in the early asymptomatic phase of PPV infection in comparison with the symptomatic phase. These early infection events were associated with the induction of genes related to pathogen resistance, such as jasmonic acid, chitinases, cytokinin glucosyl transferases and Lys- M proteins. Once the virus had accumulated, the overexpression of Dicer protein 2a genes suggested a gene silencing plant response that was suppressed by the virus HCPro and P1 proteins. These results illustrate the dynamic nature of the peach- PPV interaction at the transcriptome level and confirm that sharka symptom expression is a complex process that can be understood on the basis of changes in plant gene expression. [ABSTRACT FROM AUTHOR]

Frosty pod rot ( FPR) of Theobroma cacao (cacao) is caused by the hemibiotrophic fungus Moniliophthora roreri. Cacao clones tolerant to FPR are being planted throughout Central America. To determine whether M. roreri shows a differential molecular response during successful infections of tolerant clones, we collected field-infected pods at all stages of symptomatology for two highly susceptible clones (Pound-7 and CATIE-1000) and three tolerant clones ( UF-273, CATIE- R7 and CATIE- R4). Metabolite analysis was carried out on clones Pound-7, CATIE-1000, CATIE- R7 and CATIE- R4. As FPR progressed, the concentrations of sugars in pods dropped, whereas the levels of trehalose and mannitol increased. Associations between symptoms and fungal loads and some organic and amino acid concentrations varied depending on the clone. RNA-Seq analysis identified 873 M. roreri genes that were differentially expressed between clones, with the primary difference being whether the clone was susceptible or tolerant. Genes encoding transcription factors, heat shock proteins, transporters, enzymes modifying membranes or cell walls and metabolic enzymes, such as malate synthase and alternative oxidase, were differentially expressed. The differential expression between clones of 43 M. roreri genes was validated by real-time quantitative reverse transcription polymerase chain reaction. The expression profiles of some genes were similar in susceptible and tolerant clones (other than CATIE-R4) and varied with the biotrophic/necrotropic shift. Moniliophthora roreri genes associated with stress metabolism and responses to heat shock and anoxia were induced early in tolerant clones, their expression profiles resembling that of the necrotrophic phase. Moniliophthora roreri stress response genes, induced during the infection of tolerant clones, may benefit the fungus in overcoming cacao defense mechanisms. [ABSTRACT FROM AUTHOR]

An understanding of the tolerance mechanisms of Theobroma cacao used against Moniliophthora roreri, the causal agent of frosty pod rot, is important for the generation of stable disease-tolerant clones. A comparative view was obtained of transcript populations of infected pods from two susceptible and two tolerant clones using RNA sequence ( RNA- Seq) analysis. A total of 3009 transcripts showed differential expression among clones. KEGG (Kyoto Encyclopedia of Genes and Genomes) pathway analysis of differentially expressed genes indicated shifts in 152 different metabolic pathways between the tolerant and susceptible clones. Real-time quantitative reverse transcription polymerase chain reaction (real-time q RT- PCR) analyses of 36 genes verified the differential expression. Regression analysis validated a uniform progression in gene expression in association with infection levels and fungal loads in the susceptible clones. Expression patterns observed in the susceptible clones diverged in tolerant clones, with many genes showing higher expression at a low level of infection and fungal load. Principal coordinate analyses of real-time q RT- PCR data separated the gene expression patterns between susceptible and tolerant clones for pods showing malformation. Although some genes were constitutively differentially expressed between clones, most results suggested that defence responses were induced at low fungal load in the tolerant clones. Several elicitor-responsive genes were highly expressed in tolerant clones, suggesting rapid recognition of the pathogen and induction of defence genes. Expression patterns suggested that the jasmonic acid-ethylene- and/or salicylic acid-mediated defence pathways were activated in the tolerant clones, being enhanced by reduced brassinosteroid ( BR) biosynthesis and catabolic inactivation of both BR and abscisic acids. Finally, several genes associated with hypersensitive response-like cell death were also induced in tolerant clones. [ABSTRACT FROM AUTHOR]

Preclinical Research RNA sequencing (RNA ‐ Seq) is a novel high ‐ throughput technology for comprehensive transcriptome study. It can measure the expression levels of thousands of genes simultaneously and provide insight into functional pathways and regulations in biological processes. In addition, RNA ‐ Seq can provide copious information on alternative splicing, allele ‐ specific expression, unannotated exons, and novel transcripts (gene or noncoding RNAs). This technology has revolutionized the way biologists examine transcriptomes and has been successfully applied in drug discovery and development, being able to identify drug ‐ related genes, microRNAs, and fusion proteins. In this overview, we will review this technology including data analysis, and its recent applications in drug discovery and development. [ABSTRACT FROM AUTHOR]

The development of inexpensive and highly productive biomass sources of biofuel is a priority in global climate change biology. Arundo donax, also known as the giant reed, is recognized as one of the most promising nonfood bioenergy crops in Europe. Despite its relevance, to date no genomic resources are available to support the characterization of the developmental, adaptive and metabolic traits underlying the high productivity of this nonmodel species. We hereby present the first report on the de novo assembly of bud, culm, leaf and root transcriptomes of A. donax, which can be accessed through a customized BLAST server () for mining and exploring the genetic potential of this species. Based on functional annotation and homology comparison to 19 prospective biofuel Poaceae species, we provide the first genomic view of this so far unexplored crop and indicate the model species with highest potential for comparative genomics approaches. The analysis of the transcriptome reveals strong differences in the enrichment of the Gene Ontology categories and the relative expression among different organs, which can guide future efforts for functional genomics or genetic improvement of A. donax. A set of homologs to key genes involved in lignin, cellulose, starch, lipid metabolism and in the domestication of other crops is discussed to provide a platform for possible enhancement of productivity and saccharification efficiency in A. donax. [ABSTRACT FROM AUTHOR]

Background To identify mutant genes with high-frequency-risk-expression between lung adenocarcinoma and normal samples. Methods The ribonucleic acid RNA- Seq data GSE34914 and GSE37765 were downloaded from the Gene Expression Omnibus database, including 12 lung adenocarcinoma samples and six controls. All RNA- Seq reads were processed and the gene-expression level was calculated. Single nucleotide variation ( SNV) was analyzed and the locations of mutant sites were recorded. In addition, the frequency and risk-level of mutant genes were calculated. Gene Ontology ( GO) functional analysis was performed. The reported cancer genes were searched in tumor suppressor genes, Cancer Genes, and the Catalogue of Somatic Mutations in Cancer ( COSMIC) database. Results The SNV annotations of somatic mutation sites showed that 70% of mutation sites in the exon region occurred in the coding sequence ( CDS). Thyroid hormone receptor interactor ( TRIP)12 was identified with the highest frequency. A total of 118 mutant genes with high frequency and high-risk were selected and significantly enriched into several GO terms. No base mutation of cyclin C ( CCNC) or RAB11A was recorded. At fragments per kilobase per million reads (FPKM) ≥ 56.5, reported tumor suppressor genes catenin (cadherin-associated protein), delta ( CTNND)1, dual specificity phosphatase ( DUSP)6, malate dehydrogenase ( MDH)1 and RNA binding motif protein ( RBM)5, were identified. Notably, signal transducer and activator of transcription 2 ( STAT2) was the only transcription factor ( TF) with high-risk mutation and its expression was detected. Conclusion For the mutant genes with high-frequency-risk-expression, CTNND1, DUSP6, MDH1 and RBM5 were identified. TRIP12 might be a potential cancer-related gene, and expression of TF STAT2 with high-risk was detected. These mutant gene candidates might promote the development of lung adenocarcinoma and provide new diagnostic potential targets for treatment. [ABSTRACT FROM AUTHOR]

The pollen beetle ( Meligethes aeneus) is a major pest of oilseed rape ( Brassica napus) and other cruciferous crops in Europe. Pesticide-resistant pollen beetle populations are emerging, increasing the economic impact of this species. We isolated total RNA from the larval and adult stages, the latter either naïve or immunized by injection with bacteria and yeast. High-throughput RNA sequencing ( RNA- Seq) was carried out to establish a comprehensive transcriptome catalogue and to screen for developmental stage-specific and immunity-related transcripts. We assembled the transcriptome de novo by combining sequence tags from all developmental stages and treatments. Gene expression data based on normalized read counts revealed several functional gene categories that were differentially expressed between larvae and adults, particularly genes associated with digestion and detoxification that were induced in larvae, and genes associated with reproduction and environmental signalling that were induced in adults. We also identified many genes associated with microbe recognition, immunity-related signalling and defence effectors, such as antimicrobial peptides ( AMPs) and lysozymes. Digital gene expression analysis revealed significant differences in the profile of AMPs expressed in larvae, naïve adults and immune-challenged adults, providing insight into the steady-state differences between developmental stages and the complex transcriptional remodelling that occurs following the induction of immunity. Our data provide insight into the adaptive mechanisms used by phytophagous insects and could lead to the development of more effective control strategies for insect pests. [ABSTRACT FROM AUTHOR]

Transcriptome studies of insect herbivory are still rare, yet studies in model systems have uncovered patterns of transcript regulation that appear to provide insights into how insect herbivores attain polyphagy, such as a general increase in expression breadth and regulation of ribosomal, digestion- and detoxification-related genes. We investigated the potential generality of these emerging patterns, in the Swedish comma, Polygonia c-album, which is a polyphagous, widely-distributed butterfly. Urtica dioica and Ribes uva-crispa are hosts of P. c-album, but Ribes represents a recent evolutionary shift onto a very divergent host. Utilizing the assembled transcriptome for read mapping, we assessed gene expression finding that caterpillar life-history (i.e. 2nd vs. 4th-instar regulation) had a limited influence on gene expression plasticity. In contrast, differential expression in response to host-plant identified genes encoding serine-type endopeptidases, membrane-associated proteins and transporters. Differential regulation of genes involved in nucleic acid binding was also observed suggesting that polyphagy involves large scale transcriptional changes. Additionally, transcripts coding for structural constituents of the cuticle were differentially expressed in caterpillars in response to their diet indicating that the insect cuticle may be a target for plant defence. Our results state that emerging patterns of transcript regulation from model species appear relevant in species when placed in an evolutionary context. [ABSTRACT FROM AUTHOR]

Heterozyosity is an important feature of many plant genomes, and is related to heterosis. Sweet orange, a highly heterozygous species, is thought to have originated from an inter-species hybrid between pummelo and mandarin. To investigate the heterozygosity of the sweet orange genome and examine how this heterozygosity affects gene expression, we characterized the genome of Valencia orange for single nucleotide variations ( SNVs), small insertions and deletions ( In Dels) and structural variations ( SVs), and determined their functional effects on protein-coding genes and non-coding sequences. Almost half of the genes containing large-effect SNVs and In Dels were expressed in a tissue-specific manner. We identified 3542 large SVs (>50 bp), including deletions, insertions and inversions. Most of the 296 genes located in large-deletion regions showed low expression levels. RNA- Seq reads and DNA sequencing reads revealed that the alleles of 1062 genes were differentially expressed. In addition, we detected approximately 42 Mb of contigs that were not found in the reference genome of a haploid sweet orange by de novo assembly of unmapped reads, and annotated 134 protein-coding genes within these contigs. We discuss how this heterozygosity affects the quality of genome assembly. This study advances our understanding of the genome architecture of sweet orange, and provides a global view of gene expression at heterozygous loci. [ABSTRACT FROM AUTHOR]

Chinese fir ( Cunninghamia lanceolata), a commercially important tree for the timber and pulp industry, is widely distributed in southern China and northern Vietnam, but its large and complex genome has hindered the development of genomic resources. Few efforts have focused on analysis of the modulation of transcriptional networks in vascular cambium during the transition from active growth to dormancy in conifers. , Here, we used Illumina sequencing to analyze the global transcriptome alterations at the different stages of vascular cambium development in Chinese fir., By analyzing dynamic changes in the transcriptome of vascular cambium based on our RNA sequencing ( RNA- Seq) data at the dormant, reactivating and active stages, many potentially interesting genes were identified that encoded putative regulators of cambial activity, cell division, cell expansion and cell wall biosynthesis and modification. In particular, the genes involved in transcriptional regulation and hormone signaling were highlighted to reveal their biological importance in the cambium development and wood formation., Our results reveal the dynamics of transcriptional networks and identify potential key components in the regulation of vascular cambium development in Chinese fir, which will contribute to the in-depth study of cambial differentiation and wood-forming candidate genes in conifers. [ABSTRACT FROM AUTHOR]

To increase knowledge of transcript diversity for the giant kelp, Macrocystis pyrifera, and assess gene expression across naturally occurring depth gradients in light, temperature and nutrients, we sequenced four cDNA libraries created from blades collected at the sea surface and at 18 m depth during the winter and summer., Comparative genomics cluster analyses revealed novel gene families (clusters) in existing brown alga expressed sequence tag data compared with other related algal groups, a pattern also seen with the addition of M. pyrifera sequences., Assembly of 228 Mbp of sequence generated c. 9000 isotigs and c. 12 000 open reading frames. Annotations were assigned using families of hidden Markov models for c. 11% of open reading frames; M. pyrifera had highest similarity to other members of the Phaeophyceae, namely Ectocarpus siliculosus and Laminaria digitata., Quantitative polymerase chain reaction of transcript targets verified depth-related differences in gene expression; stress response and light-harvesting transcripts, especially members of the LI818 (also known as LHCSR) family, showed high expression in the surface compared with 18 m depth, while some nitrogen acquisition transcripts (e.g. nitrite reductase) were upregulated at depth compared with the surface, supporting a conceptual biological model of depth-dependent physiology. [ABSTRACT FROM AUTHOR]

Two common classes of nitrogen-fixing legume root nodules are those that have determinate or indeterminate meristems, as in Phaseolus bean and pea, respectively. In indeterminate nodules, rhizobia terminally differentiate into bacteroids with endoreduplicated genomes, whereas bacteroids from determinate nodules are less differentiated and can regrow. We used RNA sequencing to compare bacteroid gene expression in determinate and indeterminate nodules using two Rhizobium leguminosarum strains whose genomes differ due to replacement of the symbiosis (Sym) plasmid pRP2 (strain Rlp4292) with pRL1 (strain RlvA34), thereby switching symbiosis hosts from Phaseolus bean (determinate nodules) to pea (indeterminate nodules). Both bacteroid types have gene expression patterns typical of a stringent response, a stressful environment and catabolism of dicarboxylates, formate, amino acids and quaternary amines. Gene expression patterns were indicative that bean bacteroids were more limited for phosphate, sulphate and iron than pea bacteroids. Bean bacteroids had higher levels of expression of genes whose products are predicted to be associated with metabolite detoxification or export. Pea bacteroids had increased expression of genes associated with DNA replication, membrane synthesis and the TCA (tricarboxylic acid) cycle. Analysis of bacteroid-specific transporter genes was indicative of distinct differences in sugars and other compounds in the two nodule environments. Cell division genes were down-regulated in pea but not bean bacteroids, while DNA synthesis was increased in pea bacteroids. This is consistent with endoreduplication of pea bacteroids and their failure to regrow once nodules senesce.