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Transcriptome-seq provides insights into sex-preference pattern of gene expression between testis and ovary of the crucifix crab ( Charybdis feriatus)

Zhang Y, Miao G, Fazhan H, Waiho K, Zheng H, Li S, et al.

Physiol Genomics, 2018 05 01;50(5):393-405.
PMID: 29570432 DOI: 10.1152/physiolgenomics.00016.2018

The crucifix crab, Charybdis feriatus, which mainly inhabits Indo-Pacific region, is regarded as one of the most high-potential species for domestication and incorporation into the aquaculture sector. However, the regulatory mechanisms of sex determination and differentiation of this species remain unclear. To identify candidate genes involved in sex determination and differentiation, high throughput sequencing of transcriptome from the testis and ovary of C. feriatus was performed by the Illumina platform. After removing adaptor primers, low-quality sequences and very short (<50 nt) reads, we obtained 80.9 million and 66.2 million clean reads from testis and ovary, respectively. A total of 86,433 unigenes were assembled, and ~43% (37,500 unigenes) were successfully annotated to the NR, NT, Swiss-Prot, KEGG, COG, GO databases. By comparing the testis and ovary libraries, we obtained 27,636 differentially expressed genes. Some candidate genes involved in the sex determination and differentiation of C. feriatus were identified, such as vasa, pgds, vgr, hsp90, dsx-f, fem-1, and gpr. In addition, 88,608 simple sequence repeats were obtained, and 61,929 and 77,473 single nucleotide polymorphisms from testis and ovary were detected, respectively. The transcriptome profiling was validated by quantitative real-time PCR in 30 selected genes, which showed a good consistency. The present study is the first high-throughput transcriptome sequencing of C. feriatus. These findings will be useful for future functional analysis of sex-associated genes and molecular marker-assisted selections in C. feriatus.

Matched MeSH terms: Gene Expression Profiling/methods
Fulltext Comprehensive analysis of the venom gland transcriptome of the spider Dolomedes fimbriatus

Kozlov SA, Lazarev VN, Kostryukova ES, Selezneva OV, Ospanova EA, Alexeev DG, et al.

Sci Data, 2014;1:140023.
PMID: 25977780 DOI: 10.1038/sdata.2014.23

A comprehensive transcriptome analysis of an expressed sequence tag (EST) database of the spider Dolomedes fimbriatus venom glands using single-residue distribution analysis (SRDA) identified 7,169 unique sequences. Mature chains of 163 different toxin-like polypeptides were predicted on the basis of well-established methodology. The number of protein precursors of these polypeptides was appreciably numerous than the number of mature polypeptides. A total of 451 different polypeptide precursors, translated from 795 unique nucleotide sequences, were deduced. A homology search divided the 163 mature polypeptide sequences into 16 superfamilies and 19 singletons. The number of mature toxins in a superfamily ranged from 2 to 49, whereas the diversity of the original nucleotide sequences was greater (2-261 variants). We observed a predominance of inhibitor cysteine knot toxin-like polypeptides among the cysteine-containing structures in the analyzed transcriptome bank. Uncommon spatial folds were also found.

Matched MeSH terms: Gene Expression Profiling/methods
Transcriptional profiling of oral squamous cell carcinoma using formalin-fixed paraffin-embedded samples

Saleh A, Zain RB, Hussaini H, Ng F, Tanavde V, Hamid S, et al.

Oral Oncol, 2010 May;46(5):379-86.
PMID: 20371203 DOI: 10.1016/j.oraloncology.2010.02.022

Despite the advances in cancer treatment, the 5-year survival rate for oral cancer has not changed significantly for the past 40 years and still remains among the worst of all anatomic sites. Gene expression microarrays have been used successfully in the identification of genetic alterations in cancer development, however, these have hitherto been limited by the need for specimens with good quality intact RNA. Here, we demonstrated the use of formalin-fixed paraffin-embedded tissues in microarray experiments to identify genes differentially expressed between cancerous and normal oral tissues. Forty-three tissue samples were macrodissected and gene expression analyses were conducted using the Illumina DASL assay. We report RNA yield of 2.4 and 0.8 microg/mm(3) from tumour and normal tissues, respectively and this correlated directly with the tissue volume used for RNA extraction. Using unsupervised hierarchical clustering, distinct gene expression profiles for tumour and normal samples could be generated, and differentially expressed genes could be identified. The majority of these genes were involved in regulation of apoptosis and cell cycle, metastasis and cell adhesion including BCL2A1, BIRC5, MMP1, MMP9 and ITGB4. Representative genes were further validated in independent samples suggesting that these genes may be directly associated with oral cancer development. The ability to conduct microarrays on formalin-fixed paraffin-embedded specimens represents a significant advancement that could open up avenues for finding genes that could be used as prognostication and predictive tools for cancer.

Matched MeSH terms: Gene Expression Profiling/methods*
Suppression Subtractive Hybridization Versus Next-Generation Sequencing in Plant Genetic Engineering: Challenges and Perspectives

Sahebi M, Hanafi MM, Azizi P, Hakim A, Ashkani S, Abiri R

Mol Biotechnol, 2015 Oct;57(10):880-903.
PMID: 26271955 DOI: 10.1007/s12033-015-9884-z

Suppression subtractive hybridization (SSH) is an effective method to identify different genes with different expression levels involved in a variety of biological processes. This method has often been used to study molecular mechanisms of plants in complex relationships with different pathogens and a variety of biotic stresses. Compared to other techniques used in gene expression profiling, SSH needs relatively smaller amounts of the initial materials, with lower costs, and fewer false positives present within the results. Extraction of total RNA from plant species rich in phenolic compounds, carbohydrates, and polysaccharides that easily bind to nucleic acids through cellular mechanisms is difficult and needs to be considered. Remarkable advancement has been achieved in the next-generation sequencing (NGS) field. As a result of progress within fields related to molecular chemistry and biology as well as specialized engineering, parallelization in the sequencing reaction has exceptionally enhanced the overall read number of generated sequences per run. Currently available sequencing platforms support an earlier unparalleled view directly into complex mixes associated with RNA in addition to DNA samples. NGS technology has demonstrated the ability to sequence DNA with remarkable swiftness, therefore allowing previously unthinkable scientific accomplishments along with novel biological purposes. However, the massive amounts of data generated by NGS impose a substantial challenge with regard to data safe-keeping and analysis. This review examines some simple but vital points involved in preparing the initial material for SSH and introduces this method as well as its associated applications to detect different novel genes from different plant species. This review evaluates general concepts, basic applications, plus the probable results of NGS technology in genomics, with unique mention of feasible potential tools as well as bioinformatics.

Matched MeSH terms: Gene Expression Profiling/methods
Fulltext Global transcriptional analysis of Burkholderia pseudomallei high and low biofilm producers reveals insights into biofilm production and virulence

Chin CY, Hara Y, Ghazali AK, Yap SJ, Kong C, Wong YC, et al.

BMC Genomics, 2015;16:471.
PMID: 26092034 DOI: 10.1186/s12864-015-1692-0

Chronic bacterial infections occur as a result of the infecting pathogen's ability to live within a biofilm, hence escaping the detrimental effects of antibiotics and the immune defense system. Burkholderia pseudomallei, a gram-negative facultative pathogen, is distinctive in its ability to survive within phagocytic and non-phagocytic cells, to persist in vivo for many years and subsequently leading to relapse as well as the development of chronic disease. The capacity to persist has been attributed to the pathogen's ability to form biofilm. However, the underlying biology of B. pseudomallei biofilm development remains unresolved.

Matched MeSH terms: Gene Expression Profiling/methods
Fulltext RNA-seq data of Ganoderma boninense at axenic culture condition and under in planta pathogen-oil palm (Elaeis guineensis Jacq.) interaction

Wong MY, Govender NT, Ong CS

BMC Res Notes, 2019 Sep 24;12(1):631.
PMID: 31551084 DOI: 10.1186/s13104-019-4652-y

OBJECTIVE: Basal stem rot disease causes severe economic losses to oil palm production in South-east Asia and little is known on the pathogenicity of the pathogen, the basidiomyceteous Ganoderma boninense. Our data presented here aims to identify both the house-keeping and pathogenicity genes of G. boninense using Illumina sequencing reads.
DESCRIPTION: The hemibiotroph G. boninense establishes via root contact during early stage of colonization and subsequently kills the host tissue as the disease progresses. Information on the pathogenicity factors/genes that causes BSR remain poorly understood. In addition, the molecular expressions corresponding to G. boninense growth and pathogenicity are not reported. Here, six transcriptome datasets of G. boninense from two contrasting conditions (three biological replicates per condition) are presented. The first datasets, collected from a 7-day-old axenic condition provide an insight onto genes responsible for sustenance, growth and development of G. boninense while datasets of the infecting G. boninense collected from oil palm-G. boninense pathosystem (in planta condition) at 1 month post-inoculation offer a comprehensive avenue to understand G. boninense pathogenesis and infection especially in regard to molecular mechanisms and pathways. Raw sequences deposited in Sequence Read Archive (SRA) are available at NCBI SRA portal with PRJNA514399, bioproject ID.

Matched MeSH terms: Gene Expression Profiling/methods*
Metabolomics and its role in understanding cellular responses in plants

Bhalla R, Narasimhan K, Swarup S

Plant Cell Rep, 2005 Dec;24(10):562-71.
PMID: 16220342

A natural shift is taking place in the approaches being adopted by plant scientists in response to the accessibility of systems-based technology platforms. Metabolomics is one such field, which involves a comprehensive non-biased analysis of metabolites in a given cell at a specific time. This review briefly introduces the emerging field and a range of analytical techniques that are most useful in metabolomics when combined with computational approaches in data analyses. Using cases from Arabidopsis and other selected plant systems, this review highlights how information can be integrated from metabolomics and other functional genomics platforms to obtain a global picture of plant cellular responses. We discuss how metabolomics is enabling large-scale and parallel interrogation of cell states under different stages of development and defined environmental conditions to uncover novel interactions among various pathways. Finally, we discuss selected applications of metabolomics.

Matched MeSH terms: Gene Expression Profiling/methods
Fulltext Comparative transcriptome analysis to identify candidate genes involved in 2-methoxy-1,4-naphthoquinone (MNQ) biosynthesis in Impatiens balsamina L

Foong LC, Chai JY, Ho ASH, Yeo BPH, Lim YM, Tam SM

Sci Rep, 2020 09 30;10(1):16123.
PMID: 32999341 DOI: 10.1038/s41598-020-72997-2

Impatiens balsamina L. is a tropical ornamental and traditional medicinal herb rich in natural compounds, especially 2-methoxy-1,4-naphthoquinone (MNQ) which is a bioactive compound with tested anticancer activities. Characterization of key genes involved in the shikimate and 1,4-dihydroxy-2-naphthoate (DHNA) pathways responsible for MNQ biosynthesis and their expression profiles in I. balsamina will facilitate adoption of genetic/metabolic engineering or synthetic biology approaches to further increase production for pre-commercialization. In this study, HPLC analysis showed that MNQ was present in significantly higher quantities in the capsule pericarps throughout three developmental stages (early-, mature- and postbreaker stages) whilst its immediate precursor, 2-hydroxy-1,4-naphthoquinone (lawsone) was mainly detected in mature leaves. Transcriptomes of I. balsamina derived from leaf, flower, and three capsule developmental stages were generated, totalling 59.643 Gb of raw reads that were assembled into 94,659 unigenes (595,828 transcripts). A total of 73.96% of unigenes were functionally annotated against seven public databases and 50,786 differentially expressed genes (DEGs) were identified. Expression profiles of 20 selected genes from four major secondary metabolism pathways were studied and validated using qRT-PCR method. Majority of the DHNA pathway genes were found to be significantly upregulated in early stage capsule compared to flower and leaf, suggesting tissue-specific synthesis of MNQ. Correlation analysis identified 11 candidate unigenes related to three enzymes (NADH-quinone oxidoreductase, UDP-glycosyltransferases and S-adenosylmethionine-dependent O-methyltransferase) important in the final steps of MNQ biosynthesis based on genes expression profiles consistent with MNQ content. This study provides the first molecular insight into the dynamics of MNQ biosynthesis and accumulation across different tissues of I. balsamina and serves as a valuable resource to facilitate further manipulation to increase production of MNQ.

Matched MeSH terms: Gene Expression Profiling/methods
Fulltext Flexible model-based clustering of mixed binary and continuous data: application to genetic regulation and cancer

Zainul Abidin FN, Westhead DR

Nucleic Acids Res, 2017 04 20;45(7):e53.
PMID: 27994031 DOI: 10.1093/nar/gkw1270

Clustering is used widely in 'omics' studies and is often tackled with standard methods, e.g. hierarchical clustering. However, the increasing need for integration of multiple data sets leads to a requirement for clustering methods applicable to mixed data types, where the straightforward application of standard methods is not necessarily the best approach. A particularly common problem involves clustering entities characterized by a mixture of binary data (e.g. presence/absence of mutations, binding, motifs and epigenetic marks) and continuous data (e.g. gene expression, protein abundance, metabolite levels). Here, we present a generic method based on a probabilistic model for clustering this type of data, and illustrate its application to genetic regulation and the clustering of cancer samples. We show that the resulting clusters lead to useful hypotheses: in the case of genetic regulation these concern regulation of groups of genes by specific sets of transcription factors and in the case of cancer samples combinations of gene mutations are related to patterns of gene expression. The clusters have potential mechanistic significance and in the latter case are significantly linked to survival. The method is available as a stand-alone software package (GNU General Public Licence) from http://github.com/BioToolsLeeds/FlexiCoClusteringPackage.git.

Matched MeSH terms: Gene Expression Profiling/methods*
Biofilm-Related Diseases and Omics: Global Transcriptional Profiling of Enterococcus faecium Reveals Different Gene Expression Patterns in the Biofilm and Planktonic Cells

Lim SY, Teh CSJ, Thong KL

OMICS, 2017 10;21(10):592-602.
PMID: 29049010 DOI: 10.1089/omi.2017.0119

Enterococcus faecium is an opportunistic pathogen with a remarkable ability to acquire resistance toward multiple antibiotics, including those of last-resort drugs such as vancomycin and daptomycin. The occurrence of vancomycin-resistant E. faecium is on the rise and there is a need to understand the virulence of this organism. One of the factors that contributes to the virulence is the ability to form biofilms. Since bacteria in biofilm state are more resistant to antibiotics and host immune response, understanding the molecular mechanism of biofilm development is important to control biofilm-related diseases. The aim of this study was to determine the global gene expression profiles of an E. faecium strain, VREr5, during the early event of sessile growth compared with its planktonic phase through RNA-sequencing approach. The results clearly illustrated distinct expression profiles of the planktonic and biofilm cells. A total of 177 genes were overexpressed in the biofilm cells. Most of them encode for proteins involved in adherence, such as the ebpABCfm locus. Genes associated with plasmid replication, gene exchange, and protein synthesis were also upregulated during the early event of biofilm development. Furthermore, the transcriptome analysis also identified genes such as fsrB, luxS, and spx that might suppress biofilm formation in VREr5. The putative biofilm-related bee locus was found to be downregulated. These new findings could provide caveats for future studies on the regulation and maintenance of biofilm and development of biomarkers for biofilm-related diseases.

Matched MeSH terms: Gene Expression Profiling/methods
Fulltext Reference gene validation for gene expression normalization in canine osteosarcoma: a geNorm algorithm approach

Selvarajah GT, Bonestroo FAS, Timmermans Sprang EPM, Kirpensteijn J, Mol JA

BMC Vet Res, 2017 Nov 25;13(1):354.
PMID: 29178874 DOI: 10.1186/s12917-017-1281-3

BACKGROUND: Quantitative PCR (qPCR) is a common method for quantifying mRNA expression. Given the heterogeneity present in tumor tissues, it is crucial to normalize target mRNA expression data using appropriate reference genes that are stably expressed under a variety of pathological and experimental conditions. No studies have validated specific reference genes in canine osteosarcoma (OS). Previous gene expression studies involving canine OS have used one or two reference genes to normalize gene expression. This study aimed to validate a panel of reference genes commonly used for normalization of canine OS gene expression data using the geNorm algorithm. qPCR analysis of nine canine reference genes was performed on 40 snap-frozen primary OS tumors and seven cell lines.
RESULTS: Tumors with a variety of clinical and pathological characteristics were selected. Gene expression stability and the optimal number of reference genes for gene expression normalization were calculated. RPS5 and HNRNPH were highly stable among OS cell lines, while RPS5 and RPS19 were the best combination for primary tumors. Pairwise variation analysis recommended four and two reference genes for optimal normalization of the expression data of canine OS tumors and cell lines, respectively.
CONCLUSIONS: Appropriate combinations of reference genes are recommended to normalize mRNA levels in canine OS tumors and cell lines to facilitate standardized and reliable quantification of target gene expression, which is essential for investigating key genes involved in canine OS metastasis and for comparative biomarker discovery.

Matched MeSH terms: Gene Expression Profiling/methods
Fulltext Predicting the clinical outcome of oral potentially malignant disorders using transcriptomic-based molecular pathology

Sathasivam HP, Kist R, Sloan P, Thomson P, Nugent M, Alexander J, et al.

Br J Cancer, 2021 Aug;125(3):413-421.
PMID: 33972745 DOI: 10.1038/s41416-021-01411-z

BACKGROUND: This study was undertaken to develop and validate a gene expression signature that characterises oral potentially malignant disorders (OPMD) with a high risk of undergoing malignant transformation.
METHODS: Patients with oral epithelial dysplasia at one hospital were selected as the 'training set' (n = 56) whilst those at another hospital were selected for the 'test set' (n = 66). RNA was extracted from formalin-fixed paraffin-embedded (FFPE) diagnostic biopsies and analysed using the NanoString nCounter platform. A targeted panel of 42 genes selected on their association with oral carcinogenesis was used to develop a prognostic gene signature. Following data normalisation, uni- and multivariable analysis, as well as prognostic modelling, were employed to develop and validate the gene signature.
RESULTS: A prognostic classifier composed of 11 genes was developed using the training set. The multivariable prognostic model was used to predict patient risk scores in the test set. The prognostic gene signature was an independent predictor of malignant transformation when assessed in the test set, with the high-risk group showing worse prognosis [Hazard ratio = 12.65, p = 0.0003].
CONCLUSIONS: This study demonstrates proof of principle that RNA extracted from FFPE diagnostic biopsies of OPMD, when analysed on the NanoString nCounter platform, can be used to generate a molecular classifier that stratifies the risk of malignant transformation with promising clinical utility.

Matched MeSH terms: Gene Expression Profiling/methods*
Fulltext Comparative analysis of nucleus-encoded plastid-targeting proteins in Rafflesia cantleyi against photosynthetic and non-photosynthetic representatives reveals orthologous systems with potentially divergent functions

Ng SM, Lee XW, Mat-Isa MN, Aizat-Juhari MA, Adam JH, Mohamed R, et al.

Sci Rep, 2018 Nov 22;8(1):17258.
PMID: 30467394 DOI: 10.1038/s41598-018-35173-1

Parasitic plants are known to discard photosynthesis thus leading to the deletion or loss of the plastid genes. Despite plastid genome reduction in non-photosynthetic plants, some nucleus-encoded proteins are transported back to the plastid to carry out specific functions. In this work, we study such proteins in Rafflesia cantleyi, a member of the holoparasitic genus well-known for producing the largest single flower in the world. Our analyses of three transcriptome datasets, two holoparasites (R. cantleyi and Phelipanche aegyptiaca) and one photosynthetic plant (Arabidopsis thaliana), suggest that holoparasites, such as R. cantleyi, retain some common plastid associated processes such as biosynthesis of amino acids and lipids, but are missing photosynthesis components that can be extensions of these pathways. The reconstruction of two selected biosynthetic pathways involving plastids correlates the trend of plastid retention to pathway complexity - transcriptome evidence for R. cantleyi suggests alternate mechanisms in regulating the plastidial heme and terpenoid backbone biosynthesis pathways. The evolution to holoparasitism from autotrophy trends towards devolving the plastid genes to the nuclear genome despite the functional sites remaining in the plastid, or maintaining non-photosynthetic processes in the plastid, before the eventual loss of the plastid and any site dependent functions.

Matched MeSH terms: Gene Expression Profiling/methods
Fulltext Long non-coding RNA expression in primary human monocytes

Mirsafian H, Manda SS, Mitchell CJ, Sreenivasamurthy S, Ripen AM, Mohamad SB, et al.

Genomics, 2016 07;108(1):37-45.
PMID: 26778813 DOI: 10.1016/j.ygeno.2016.01.002

Long non-coding RNAs (lncRNAs) have been shown to possess a wide range of functions in both cellular and developmental processes including cancers. Although some of the lncRNAs have been implicated in the regulation of the immune response, the exact function of the large majority of lncRNAs still remains unknown. In this study, we characterized the lncRNAs in human primary monocytes, an essential component of the innate immune system. We performed RNA sequencing of monocytes from four individuals and combined our data with eleven other publicly available datasets. Our analysis led to identification of ~8000 lncRNAs of which >1000 have not been previously reported in monocytes. PCR-based validation of a subset of the identified novel long intergenic noncoding RNAs (lincRNAs) revealed distinct expression patterns. Our study provides a landscape of lncRNAs in monocytes, which could facilitate future experimental studies to characterize the functions of these molecules in the innate immune system.

Matched MeSH terms: Gene Expression Profiling/methods*
Molecular cloning and sequence analysis of a cDNA encoding ornithine decarboxylase cDNA from chilli (Capsicum annuum)

Zainal Z, Sajari R, Ismail I

J. Biochem. Mol. Biol. Biophys., 2002 Dec;6(6):415-9.
PMID: 14972797

Ornithine decarboxylase (ODC) is an enzyme of one of the two pathways of putrescine biosynthesis in plants. The genes encoding ODC have previously been cloned from Datura stramonium and human. Using differential screening, we isolated ODC cDNA clone from a cDNA library of ripening Capsicum annuum fruit. The cDNA clone designated CUKM10 contains an insert of 1523 bp. The longest open reading frame potentially encodes a peptide of 345 amino acids with an estimated molecular mass of 47 kDa and exhibit striking similarity to other ODCs. Expression analysis showed that the capODC hybridised to a single transcript with a size of 1.7 kb. The capODC transcript was first observed in early ripening and increased steadily until it reached fully ripening stage. From the observation it is suggested that capODC is developmentally regulated especially during later stage of ripening.

Matched MeSH terms: Gene Expression Profiling/methods*
Fulltext A blood-based gene expression and signaling pathway analysis to differentiate between high and low grade gliomas

Ponnampalam SN, Kamaluddin NR, Zakaria Z, Matheneswaran V, Ganesan D, Haspani MS, et al.

Oncol Rep, 2017 Jan;37(1):10-22.
PMID: 28004117 DOI: 10.3892/or.2016.5285

The aims of the present study were to undertake gene expression profiling of the blood of glioma patients to determine key genetic components of signaling pathways and to develop a panel of genes that could be used as a potential blood-based biomarker to differentiate between high and low grade gliomas, non-gliomas and control samples. In this study, blood samples were obtained from glioma patients, non-glioma and control subjects. Ten samples each were obtained from patients with high and low grade tumours, respectively, ten samples from non-glioma patients and twenty samples from control subjects. Total RNA was isolated from each sample after which first and second strand synthesis was performed. The resulting cRNA was then hybridized with the Agilent Whole Human Genome (4x44K) microarray chip according to the manufacturer's instructions. Universal Human Reference RNA and samples were labeled with Cy3 CTP and Cy5 CTP, respectively. Microarray data were analyzed by the Agilent Gene Spring 12.1V software using stringent criteria which included at least a 2-fold difference in gene expression between samples. Statistical analysis was performed using the unpaired Student's t-test with a p<0.01. Pathway enrichment was also performed, with key genes selected for validation using droplet digital polymerase chain reaction (ddPCR). The gene expression profiling indicated that were a substantial number of genes that were differentially expressed with more than a 2-fold change (p<0.01) between each of the four different conditions. We selected key genes within significant pathways that were analyzed through pathway enrichment. These key genes included regulators of cell proliferation, transcription factors, cytokines and tumour suppressor genes. In the present study, we showed that key genes involved in significant and well established pathways, could possibly be used as a potential blood-based biomarker to differentiate between high and low grade gliomas, non-gliomas and control samples.

Matched MeSH terms: Gene Expression Profiling/methods
Fulltext Endogenous HIF2A reporter systems for high-throughput functional screening

Zaini MN, Patel SA, Syafruddin SE, Rodrigues P, Vanharanta S

Sci Rep, 2018 08 13;8(1):12063.
PMID: 30104738 DOI: 10.1038/s41598-018-30499-2

Tissue-specific transcriptional programs control most biological phenotypes, including disease states such as cancer. However, the molecular details underlying transcriptional specificity is largely unknown, hindering the development of therapeutic approaches. Here, we describe novel experimental reporter systems that allow interrogation of the endogenous expression of HIF2A, a critical driver of renal oncogenesis. Using a focused CRISPR-Cas9 library targeting chromatin regulators, we provide evidence that these reporter systems are compatible with high-throughput screening. Our data also suggests redundancy in the control of cancer type-specific transcriptional traits. Reporter systems such as those described here could facilitate large-scale mechanistic dissection of transcriptional programmes underlying cancer phenotypes, thus paving the way for novel therapeutic approaches.

Matched MeSH terms: Gene Expression Profiling/methods*
Fulltext Transcriptome Profiling of Haloxylon persicum (Bunge ex Boiss and Buhse) an Endangered Plant Species under PEG-Induced Drought Stress

Thayale Purayil F, Rajashekar B, S Kurup S, Cheruth AJ, Subramaniam S, Hassan Tawfik N, et al.

Genes (Basel), 2020 06 10;11(6).
PMID: 32531994 DOI: 10.3390/genes11060640

Haloxylon persicum is an endangered western Asiatic desert plant species, which survives under extreme environmental conditions. In this study, we focused on transcriptome analysis of H. persicum to understand the molecular mechanisms associated with drought tolerance. Two different periods of polyethylene glycol (PEG)-induced drought stress (48 h and 72 h) were imposed on H. persicum under in vitro conditions, which resulted in 18 million reads, subsequently assembled by de novo method with more than 8000 transcripts in each treatment. The N50 values were 1437, 1467, and 1524 for the control sample, 48 h samples, and 72 h samples, respectively. The gene ontology (GO) and Kyoto encyclopedia of genes and genomes (KEGG) pathway analysis resulted in enrichment of mitogen-activated protein kinase (MAPK) and plant hormone signal transduction pathways under PEG-induced drought conditions. The differential gene expression analysis (DGEs) revealed significant changes in the expression pattern between the control and the treated samples. The KEGG analysis resulted in mapping transcripts with 138 different pathways reported in plants. The differential expression of drought-responsive transcription factors depicts the possible signaling cascades involved in drought tolerance. The present study provides greater insight into the fundamental transcriptome reprogramming of desert plants under drought.

Matched MeSH terms: Gene Expression Profiling/methods
Fulltext A comparison of physiological and transcriptome responses to water deprivation and salt loading in the rat supraoptic nucleus

Greenwood MP, Mecawi AS, Hoe SZ, Mustafa MR, Johnson KR, Al-Mahmoud GA, et al.

Am J Physiol Regul Integr Comp Physiol, 2015 Apr 01;308(7):R559-68.
PMID: 25632023 DOI: 10.1152/ajpregu.00444.2014

Salt loading (SL) and water deprivation (WD) are experimental challenges that are often used to study the osmotic circuitry of the brain. Central to this circuit is the supraoptic nucleus (SON) of the hypothalamus, which is responsible for the biosynthesis of the hormones, arginine vasopressin (AVP) and oxytocin (OXT), and their transport to terminals that reside in the posterior lobe of the pituitary. On osmotic challenge evoked by a change in blood volume or osmolality, the SON undergoes a function-related plasticity that creates an environment that allows for an appropriate hormone response. Here, we have described the impact of SL and WD compared with euhydrated (EU) controls in terms of drinking and eating behavior, body weight, and recorded physiological data including circulating hormone data and plasma and urine osmolality. We have also used microarrays to profile the transcriptome of the SON following SL and remined data from the SON that describes the transcriptome response to WD. From a list of 2,783 commonly regulated transcripts, we selected 20 genes for validation by qPCR. All of the 9 genes that have already been described as expressed or regulated in the SON by osmotic stimuli were confirmed in our models. Of the 11 novel genes, 5 were successfully validated while 6 were false discoveries.

Matched MeSH terms: Gene Expression Profiling/methods
Transcriptome-wide identification and characterization of the Rab GTPase family in mango

Lawson T, Lycett GW, Mayes S, Ho WK, Chin CF

Mol Biol Rep, 2020 Jun;47(6):4183-4197.
PMID: 32444976 DOI: 10.1007/s11033-020-05519-y

The Rab GTPase family plays a vital role in several plant physiological processes including fruit ripening. Fruit softening during ripening involves trafficking of cell wall polymers and enzymes between cellular compartments. Mango, an economically important fruit crop, is known for its delicious taste, exotic flavour and nutritional value. So far, there is a paucity of information on the mango Rab GTPase family. In this study, 23 genes encoding Rab proteins were identified in mango by a comprehensive in silico approach. Sequence alignment and similarity tree analysis with the model plant Arabidopsis as a reference enabled the bona fide assignment of the deduced mango proteins to classify into eight subfamilies. Expression analysis by RNA-Sequencing (RNA-Seq) showed that the Rab genes were differentially expressed in ripe and unripe mangoes suggesting the involvement of vesicle trafficking during ripening. Interaction analysis showed that the proteins involved in vesicle trafficking and cell wall softening were interconnected providing further evidence of the involvement of the Rab GTPases in fruit softening. Correlation analyses showed a significant relationship between the expression level of the RabA3 and RabA4 genes and fruit firmness at the unripe stage of the mango varieties suggesting that the differences in gene expression level might be associated with the contrasting firmness of these varieties. This study will not only provide new insights into the complexity of the ripening-regulated molecular mechanism but also facilitate the identification of potential Rab GTPases to address excessive fruit softening.

Matched MeSH terms: Gene Expression Profiling/methods

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