Exploration of wheat and pathogen transcriptomes during tan spot infection

Objectives: The fungus Pyrenophora tritici-repentis is the causal agent of tan spot, a major disease of wheat (Triticum aestivum). Here, we used RNA sequencing to generate transcriptional datasets for both the host and pathogen during infection and during in vitro pathogen growth stages. Data description: To capture gene expression during wheat infection with the P. tritici-repentis isolate M4, RNA datasets were generated for wheat inoculated with P. tritici-repentis (infection) and a mock (control) at 3 and 4 days post-infection, when scorable leaf disease symptoms manifest. The P. tritici-repentis isolate M4 was also RNA sequenced to capture gene expression in vitro at two different growth stages: 7-day old vegetative mycelia and 9-day old sporulating mycelia, to coincide with a latent growth stage and early sporulation respectively. In total, 6 RNA datasets are available to aid in the validation of predicted genes of P. tritici-repentis and wheat. The datasets generated offer an insight into the transcriptomic profile of the host-pathogen interaction and can be used to investigate the expression of a subset of transcripts or targeted genes prior to designing cost-intensive RNA sequencing experiments, that would be best further explored with replication and a time series analysis

A gene expression predictor of response to EGFR-targeted therapy stratifies progression-free survival to cetuximab in KRAS wild-type metastatic colorectal cancer

<p>Abstract</p> <p>Background</p> <p>The anti-EGFR monoclonal antibody cetuximab is used in metastatic colorectal cancer (CRC), and predicting responsive patients garners great interest, due to the high cost of therapy. Mutations in the KRAS gene occur in ~40% of CRC and are a negative predictor of response to cetuximab. However, many KRAS-wildtype patients do not benefit from cetuximab. We previously published a gene expression predictor of sensitivity to erlotinib, an EGFR inhibitor. The purpose of this study was to determine if this predictor could identify KRAS-wildtype CRC patients who will benefit from cetuximab therapy.</p> <p>Methods</p> <p>Microarray data from 80 metastatic CRC patients subsequently treated with cetuximab were extracted from the study by Khambata-Ford et al. The study included KRAS status, response, and PFS for each patient. The gene expression data were scaled and analyzed using our predictive model. An improved predictive model of response was identified by removing features in the 180-gene predictor that introduced noise.</p> <p>Results</p> <p>Forty-three of eighty patients were identified as harboring wildtype-KRAS. When the model was applied to these patients, the predicted-sensitive group had significantly longer PFS than the predicted-resistant group (median 88 days vs. 56 days; mean 117 days vs. 63 days, respectively, p = 0.008). Kaplan-Meier curves were also significantly improved in the predicted-sensitive group (p = 0.0059, HR = 0.4109. The model was simplified to 26 of the original 180 genes and this further improved stratification of PFS (median 147 days vs. 56.5 days in the predicted sensitive and resistant groups, respectively, p < 0.0001). However, the simplified model will require further external validation, as features were selected based on their correlation to PFS in this dataset.</p> <p>Conclusion</p> <p>Our model of sensitivity to EGFR inhibition stratified PFS following cetuximab in KRAS-wildtype CRC patients. This study represents the first true external validation of a molecular predictor of response to cetuximab in KRAS-WT metastatic CRC. Our model may hold clinical utility for identifying patients responsive to cetuximab and may therefore minimize toxicity and cost while maximizing benefit.</p

Minimal change in structural, functional and inflammatory markers of lung disease in newborn screened infants with cystic fibrosis at one year

BACKGROUND: With the widespread introduction of newborn screening for cystic fibrosis (CF), there has been considerable emphasis on the need to develop objective markers of lung health that can be used during infancy. We hypothesised that in a newborn screened (NBS) UK cohort, evidence of airway inflammation and infection at one year would be associated with adverse structural and functional outcomes at the same age. METHODS: Infants underwent lung function testing, chest CT scan and bronchoscopy with bronchoalveolar lavage (BAL) at 1 year of age when clinically well. Microbiology cultures were also available from routine cough swabs. RESULTS: 65 infants had lung function, CT and BAL. Mean (SD) lung clearance index and forced expiratory volume in 0.5 s z-scores were 0.9(1.2) and -0.6(1.1) respectively; median Brody II CF-CT air trapping score on chest CT =0 (interquartile range 0-1, maximum possible score 27). Infants isolating any significant pathogen by 1 yr of age had higher LCI z-score (mean difference 0.9; 95%CI:0.4-1.4; p = 0.001) and a trend towards higher air trapping scores on CT (p = 0.06). BAL neutrophil elastase was detectable in 23% (10/43) infants in whom BAL supernatant was available. This did not relate to air trapping score on CT. CONCLUSIONS: In this UK NBS cohort at one year of age, lung and airway damage is much milder and associations between inflammation, abnormal physiology and structural changes were at best weak, contrary to our hypothesis and previously published reports. Continued follow-up will clarify longer term implications of these very mild structural, functional and inflammatory changes

Hsp-27 expression at diagnosis predicts poor clinical outcome in prostate cancer independent of ETS-gene rearrangement

BACKGROUND: This study was performed to test the hypothesis that expression of small heat shock protein Hsp-27 is, at diagnosis, a reliable predictive biomarker of clinically aggressive prostate cancer. METHODS: A panel of tissue microarrays constructed from a well-characterised cohort of 553 men with conservatively managed prostate cancer was stained immunohistochemically to detect Hsp-27 protein. Hsp-27 expression was compared with a series of pathological and clinical parameters, including outcome. RESULTS: Hsp-27 staining was indicative of higher Gleason score (P7, the presence of Hsp-27 retained its power to independently predict poor clinical outcome (P<0.002). Higher levels of Hsp-27 staining were almost entirely restricted to cancers lacking ERG rearrangements (chi2 trend=31.4, P<0.001), although this distribution did not have prognostic significance. INTERPRETATION: This study has confirmed that, in prostate cancers managed conservatively over a period of more than 15 years, expression of Hsp-27 is an accurate and independent predictive biomarker of aggressive disease with poor clinical outcome (P<0.001). These findings suggest that apoptotic and cell-migration pathways modulated by Hsp-27 may contain targets susceptible to the development of biologically appropriate chemotherapeutic agents that are likely to prove effective in treating aggressive prostate cancers

Benzoxazinoids in Root Exudates of Maize Attract Pseudomonas putida to the Rhizosphere

Benzoxazinoids, such as 2,4-dihydroxy-7-methoxy-2H-1,4-benzoxazin-3(4H)-one (DIMBOA), are secondary metabolites in grasses. In addition to their function in plant defence against pests and diseases above-ground, benzoxazinoids (BXs) have also been implicated in defence below-ground, where they can exert allelochemical or antimicrobial activities. We have studied the impact of BXs on the interaction between maize and Pseudomonas putida KT2440, a competitive coloniser of the maize rhizosphere with plant-beneficial traits. Chromatographic analyses revealed that DIMBOA is the main BX compound in root exudates of maize. In vitro analysis of DIMBOA stability indicated that KT2440 tolerance of DIMBOA is based on metabolism-dependent breakdown of this BX compound. Transcriptome analysis of DIMBOA-exposed P. putida identified increased transcription of genes controlling benzoate catabolism and chemotaxis. Chemotaxis assays confirmed motility of P. putida towards DIMBOA. Moreover, colonisation essays in soil with Green Fluorescent Protein (GFP)-expressing P. putida showed that DIMBOA-producing roots of wild-type maize attract significantly higher numbers of P. putida cells than roots of the DIMBOA-deficient bx1 mutant. Our results demonstrate a central role for DIMBOA as a below-ground semiochemical for recruitment of plant-beneficial rhizobacteria during the relatively young and vulnerable growth stages of maize

Transcriptional Profiling of Non-Small Cell Lung Cancer Cells with Activating EGFR Somatic Mutations

Activating somatic mutations in epidermal growth factor receptor (EGFR) confer unique biologic features to non-small cell lung cancer (NSCLC) cells, but the transcriptional mediators of EGFR in this subgroup of NSCLC have not been fully elucidated.Here we used genetic and pharmacologic approaches to elucidate the transcriptomes of NSCLC cell lines. We transcriptionally profiled a panel of EGFR-mutant and -wild-type NSCLC cell lines cultured in the presence or absence of an EGFR tyrosine kinase inhibitor. Hierarchical analysis revealed that the cell lines segregated on the basis of EGFR mutational status (mutant versus wild-type), and expression signatures were identified by supervised analysis that distinguished the cell lines based on mutational status (wild-type versus mutant) and type of mutation (L858R versus Delta746-750). Using an EGFR mutation-specific expression signature as a probe, we mined the gene expression profiles of two independent cohorts of NSCLC patients and found the signature in a subset. EGFR tyrosine kinase inhibitor treatment regulated the expression of multiple genes, and pharmacologic inhibition of the protein products of two of them (PTGS2 and EphA2) inhibited anchorage-independent growth in EGFR-mutant NSCLC cells.We have elucidated genes not previously associated with EGFR-mutant NSCLC, two of which enhanced the clonogenicity of these cells, distinguishing these mediators from others previously shown to maintain cell survival. These findings have potential clinical relevance given the availability of pharmacologic tools to inhibit the protein products of these genes

Association of BANK1 and TNFSF4 with systemic lupus erythematosus in Hong Kong Chinese

Systemic lupus erythematosus (SLE) is a prototypic autoimmune disease with complex genetic inheritance. Recently, single nucleotide polymorphisms (SNPs) in BANK1 and TNFSF4 have been shown to be associated with SLE in Caucasian populations, but it is not known whether they are also involved in the disease in other ethnic groups. Recent data from our genome-wide association study (GWAS) for 314 SLE cases and 920 controls collected in Hong Kong identified SNPs in and around BANK1 and TNFSF4 to be associated with SLE risk. On the basis of the results of the reported studies and our GWAS, SNPs were selected for further genotyping in 949 SLE patients (overlapping with the 314 cases in our GWAS) and non-overlapping 1042 healthy controls. We confirmed the associations of BANK1 and TNFSF4 with SLE in Chinese (BANK1, rs3733197, odds ratio (OR)=0.84, P=0.021; BANK1, rs17266594, OR=0.61, P=4.67 × 10−9; TNFSF4, rs844648, OR=1.22, P=2.47 × 10−3; TNFSF4, rs2205960, OR=1.30, P=2.41 × 10−4). Another SNP located in intron 1 of BANK1, rs4522865, was separately replicated by Sequenom in 360 cases and 360 controls and was also confirmed to be associated with SLE (OR=0.725, P=2.93 × 10−3). Logistic regression analysis showed that rs3733197 (A383T in ankyrin domain) and rs17266594 (a branch point-site SNP) from BANK1 had independent contributions towards the disease association (P=0.037 and 6.63 × 10−8, respectively). In TNFSF4, rs2205960 was associated with SLE independently from the effect of rs844648 (P=6.26 × 10−3), but not vice versa (P=0.55). These findings suggest that multiple independent genetic variants may be present within the gene locus, which exert their effects on SLE pathogenesis through different mechanisms

Profiling Critical Cancer Gene Mutations in Clinical Tumor Samples

BACKGROUND: Detection of critical cancer gene mutations in clinical tumor specimens may predict patient outcomes and inform treatment options; however, high-throughput mutation profiling remains underdeveloped as a diagnostic approach. We report the implementation of a genotyping and validation algorithm that enables robust tumor mutation profiling in the clinical setting. METHODOLOGY: We developed and implemented an optimized mutation profiling platform ("OncoMap") to interrogate approximately 400 mutations in 33 known oncogenes and tumor suppressors, many of which are known to predict response or resistance to targeted therapies. The performance of OncoMap was analyzed using DNA derived from both frozen and FFPE clinical material in a diverse set of cancer types. A subsequent in-depth analysis was conducted on histologically and clinically annotated pediatric gliomas. The sensitivity and specificity of OncoMap were 93.8% and 100% in fresh frozen tissue; and 89.3% and 99.4% in FFPE-derived DNA. We detected known mutations at the expected frequencies in common cancers, as well as novel mutations in adult and pediatric cancers that are likely to predict heightened response or resistance to existing or developmental cancer therapies. OncoMap profiles also support a new molecular stratification of pediatric low-grade gliomas based on BRAF mutations that may have immediate clinical impact. CONCLUSIONS: Our results demonstrate the clinical feasibility of high-throughput mutation profiling to query a large panel of "actionable" cancer gene mutations. In the future, this type of approach may be incorporated into both cancer epidemiologic studies and clinical decision making to specify the use of many targeted anticancer agents

Integration of Evolutionary Features for the Identification of Functionally Important Residues in Major Facilitator Superfamily Transporters

The identification of functionally important residues is an important challenge for understanding the molecular mechanisms of proteins. Membrane protein transporters operate two-state allosteric conformational changes using functionally important cooperative residues that mediate long-range communication from the substrate binding site to the translocation pathway. In this study, we identified functionally important cooperative residues of membrane protein transporters by integrating sequence conservation and co-evolutionary information. A newly derived evolutionary feature, the co-evolutionary coupling number, was introduced to measure the connectivity of co-evolving residue pairs and was integrated with the sequence conservation score. We tested this method on three Major Facilitator Superfamily (MFS) transporters, LacY, GlpT, and EmrD. MFS transporters are an important family of membrane protein transporters, which utilize diverse substrates, catalyze different modes of transport using unique combinations of functional residues, and have enough characterized functional residues to validate the performance of our method. We found that the conserved cores of evolutionarily coupled residues are involved in specific substrate recognition and translocation of MFS transporters. Furthermore, a subset of the residues forms an interaction network connecting functional sites in the protein structure. We also confirmed that our method is effective on other membrane protein transporters. Our results provide insight into the location of functional residues important for the molecular mechanisms of membrane protein transporters