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Transposon mutagenesis

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https://www.readbyqxmd.com/read/28829764/rna-helicase-dhx15-acts-as-a-tumour-suppressor-in-glioma
#1
Shingo Ito, Hideto Koso, Kazuhiro Sakamoto, Sumiko Watanabe
BACKGROUND: Glioblastoma is the most common form of malignant brain cancer and has a poor prognosis in adults. We identified Dhx15 as a candidate tumour suppressor gene in glioma by transposon-based mutagenesis. Dhx15 is an adenosine triphosphate (ATP)-dependent RNA helicase belonging to the DEAH-box (DHX) helicase family, but its role in cancer remains elusive. METHODS: DHX15 expression levels were examined in glioma cell lines. DHX15 functions were examined by gain- and loss-of-function analyses...
August 22, 2017: British Journal of Cancer
https://www.readbyqxmd.com/read/28790987/comparative-analysis-of-the-flavobacterium-columnare-genomovar-i-and-ii-genomes
#2
Salih Kumru, Hasan C Tekedar, Nagihan Gulsoy, Geoffrey C Waldbieser, Mark L Lawrence, Attila Karsi
Columnaris disease caused by Gram-negative rod Flavobacterium columnare is one of the most common diseases of catfish. F. columnare is also a common problem in other cultured fish species worldwide. F. columnare has three major genomovars; we have sequenced a representative strain from genomovar I (ATCC 49512, which is avirulent in catfish) and genomovar II (94-081, which is highly pathogenic in catfish). Here, we present a comparative analysis of the two genomes. Interestingly, F. columnare ATCC 49512 and 94-081 meet criteria to be considered different species based on the Average Nucleotide Identity (90...
2017: Frontiers in Microbiology
https://www.readbyqxmd.com/read/28782559/trap-seq-an-rna-sequencing-based-pipeline-for-the-identification-of-gene-trap-insertions-in-mammalian-cells
#3
Cristina Mayor-Ruiz, Orlando Dominguez, Oscar Fernandez-Capetillo
The development of haploid mammalian cell lines, coupled to next-generation sequencing, has recently facilitated forward genetic screenings in mammals. For mutagenesis, retrovirus- or transposon-based gene traps are frequently used. Current methods to map gene-trap insertions are based on inverse or splinkerette PCR, which despite their efficacy are prone to artifacts and do not provide information on expression of the targeted gene. Here, we describe a new RNA sequencing-based method (Trap(Seq)) to map gene-trap insertions...
August 4, 2017: Journal of Molecular Biology
https://www.readbyqxmd.com/read/28765224/genome-wide-sensitivity-analysis-of-the-microsymbiont-sinorhizobium-meliloti-to-symbiotically-important-defensin-like-host-peptides
#4
Markus F F Arnold, Mohammed Shabab, Jon Penterman, Kevin L Boehme, Joel S Griffitts, Graham C Walker
The model legume species Medicago truncatula expresses more than 700 nodule-specific cysteine-rich (NCR) signaling peptides that mediate the differentiation of Sinorhizobium meliloti bacteria into nitrogen-fixing bacteroids. NCR peptides are essential for a successful symbiosis in legume plants of the inverted-repeat-lacking clade (IRLC) and show similarity to mammalian defensins. In addition to signaling functions, many NCR peptides exhibit antimicrobial activity in vitro and in vivo Bacterial resistance to these antimicrobial activities is likely to be important for symbiosis...
August 1, 2017: MBio
https://www.readbyqxmd.com/read/28724549/identification-of-new-tumor-suppressor-genes-in-triple-negative-breast-cancer
#5
Roberto Rangel, Liliana Guzman-Rojas, Takahiro Kodama, Michiko Kodama, Justin Y Newberg, Neal G Copeland, Nancy A Jenkins
Although genomic sequencing has provided a better understating of the genetic landmarks in triple-negative breast cancer (TNBC), functional validation of candidate cancer genes (CCG) remains unsolved. In this study, we used a transposon mutagenesis strategy based on a two-step Sleeping Beauty (SB) forward genetic screen to identify and validate new tumor suppressors (TS) in this disease. We generated 120 siRNAs targeting 40 SB-identified candidate breast cancer TS genes and used them to downregulate expression of these genes in four human TNBC cell lines...
July 19, 2017: Cancer Research
https://www.readbyqxmd.com/read/28698237/maize-yabby-genes-drooping-leaf1-and-drooping-leaf2-regulate-plant-architecture
#6
Josh Strable, Jason G Wallace, Erica Unger-Wallace, Sarah Briggs, Peter J Bradbury, Edward S Buckler, Erik Vollbrecht
Leaf architecture directly influences canopy structure, consequentially affecting yield. We discovered a maize (Zea mays) mutant with aberrant leaf architecture, which we named drooping leaf1 (drl1). Pleiotropic mutations in drl1 affect leaf length and width, leaf angle, and internode length and diameter. These phenotypes are enhanced by natural variation at the drl2 enhancer locus, including reduced expression of the drl2-Mo17 allele in the Mo17 inbred. A second drl2 allele, produced by transposon mutagenesis, interacted synergistically with drl1 mutants and reduced drl2 transcript levels...
July 2017: Plant Cell
https://www.readbyqxmd.com/read/28683522/role-of-eptc-in-biofilm-formation-by-campylobacter-jejuni-nctc11168-on-polystyrene-and-glass-surfaces
#7
Eun Seob Lim, Joo-Sung Kim
The complex roles of cell surface modification in the biofilm formation of Campylobacter jejuni, a major cause of worldwide foodborne diarrheal disease, are poorly understood. In a screen of mutants from random transposon mutagenesis, an insertional mutation in eptC gene (cj0256) resulted in a significant decrease in C. jejuni NCTC11168 biofilm formation (<20%) on major food contact surfaces, such as polystyrene and borosilicate glass, compared with wild-type cells (p < 0.05). In C. jejuni strain 81-176, the protein encoded by eptC modifies cell surface structures, such as lipid A, the inner core of lipooligosaccharide, and the flagellar rod protein (FlgG), by attaching phosphoethanolamine...
July 7, 2017: Journal of Microbiology and Biotechnology
https://www.readbyqxmd.com/read/28680437/knots-untie-molecular-determinants-involved-in-knot-formation-induced-by-pseudomonas-savastanoi-in-woody-hosts
#8
REVIEW
Eloy Caballo-Ponce, Jesús Murillo, Marta Martínez-Gil, Alba Moreno-Pérez, Adrián Pintado, Cayo Ramos
The study of the molecular basis of tree diseases is lately receiving a renewed attention, especially with the emerging perception that pathogens require specific pathogenicity and virulence factors to successfully colonize woody hosts. Pathosystems involving woody plants are notoriously difficult to study, although the use of model bacterial strains together with genetically homogeneous micropropagated plant material is providing a significant impetus to our understanding of the molecular determinants leading to disease...
2017: Frontiers in Plant Science
https://www.readbyqxmd.com/read/28650484/insertional-mutagenesis-identifies-drivers-of-a-novel-oncogenic-pathway-in-invasive-lobular-breast-carcinoma
#9
Sjors M Kas, Julian R de Ruiter, Koen Schipper, Stefano Annunziato, Eva Schut, Sjoerd Klarenbeek, Anne Paulien Drenth, Eline van der Burg, Christiaan Klijn, Jelle J Ten Hoeve, David J Adams, Marco J Koudijs, Jelle Wesseling, Micha Nethe, Lodewyk F A Wessels, Jos Jonkers
Invasive lobular carcinoma (ILC) is the second most common breast cancer subtype and accounts for 8-14% of all cases. Although the majority of human ILCs are characterized by the functional loss of E-cadherin (encoded by CDH1), inactivation of Cdh1 does not predispose mice to develop mammary tumors, implying that mutations in additional genes are required for ILC formation in mice. To identify these genes, we performed an insertional mutagenesis screen using the Sleeping Beauty transposon system in mice with mammary-specific inactivation of Cdh1...
August 2017: Nature Genetics
https://www.readbyqxmd.com/read/28614737/glucose-triggers-the-cytotoxicity-of-citrobacter-sp-r1-against-microcystis-aeruginosa
#10
Pengfei Sun, Sofia Esquivel-Elizondo, Yuhua Zhao, Yonghong Wu
Algicidal bacteria offer a promising option for killing Microcystis aeruginosa, one notorious cyanobacteria causing harmful algal blooms. In this study, Citrobacter sp. R1 presented high algicidal activity (81.6±2.2%, 72h) against M. aeruginosa when cultured using glucose, while it showed no algicidal activity (0±3.4%) when cultured using wheat bran, suggesting that appropriate carbon source is crucial for algicidal bacteria in killing M. aeruginosa. The underlying algicidal mechanism of strain R1 was explored by studying the effect of different carbon sources (glucose and wheat bran) on its key algicidal gene expression and total protein translation...
June 11, 2017: Science of the Total Environment
https://www.readbyqxmd.com/read/28614382/genome-wide-transposon-mutagenesis-of-proteus-mirabilis-essential-genes-fitness-factors-for-catheter-associated-urinary-tract-infection-and-the-impact-of-polymicrobial-infection-on-fitness-requirements
#11
Chelsie E Armbruster, Valerie Forsyth-DeOrnellas, Alexandra O Johnson, Sara N Smith, Lili Zhao, Weisheng Wu, Harry L T Mobley
The Gram-negative bacterium Proteus mirabilis is a leading cause of catheter-associated urinary tract infections (CAUTIs), which are often polymicrobial. Numerous prior studies have uncovered virulence factors for P. mirabilis pathogenicity in a murine model of ascending UTI, but little is known concerning pathogenesis during CAUTI or polymicrobial infection. In this study, we utilized five pools of 10,000 transposon mutants each and transposon insertion-site sequencing (Tn-Seq) to identify the full arsenal of P...
June 2017: PLoS Pathogens
https://www.readbyqxmd.com/read/28584132/transposon-mutagenesis-identifies-chromatin-modifiers-cooperating-with-ras-in-thyroid-tumorigenesis-and-detects-atxn7-as-a-cancer-gene
#12
Cristina Montero-Conde, Luis J Leandro-Garcia, Xu Chen, Gisele Oler, Sergio Ruiz-Llorente, Mabel Ryder, Iñigo Landa, Francisco Sanchez-Vega, Konnor La, Ronald A Ghossein, Dean F Bajorin, Jeffrey A Knauf, Jesse D Riordan, Adam J Dupuy, James A Fagin
Oncogenic RAS mutations are present in 15-30% of thyroid carcinomas. Endogenous expression of mutant Ras is insufficient to initiate thyroid tumorigenesis in murine models, indicating that additional genetic alterations are required. We used Sleeping Beauty (SB) transposon mutagenesis to identify events that cooperate with Hras(G12V) in thyroid tumor development. Random genomic integration of SB transposons primarily generated loss-of-function events that significantly increased thyroid tumor penetrance in Tpo-Cre/homozygous FR-Hras(G12V) mice...
June 20, 2017: Proceedings of the National Academy of Sciences of the United States of America
https://www.readbyqxmd.com/read/28575524/identifying-transposon-insertions-and-their-effects-from-rna-sequencing-data
#13
Julian R de Ruiter, Sjors M Kas, Eva Schut, David J Adams, Marco J Koudijs, Lodewyk F A Wessels, Jos Jonkers
Insertional mutagenesis using engineered transposons is a potent forward genetic screening technique used to identify cancer genes in mouse model systems. In the analysis of these screens, transposon insertion sites are typically identified by targeted DNA-sequencing and subsequently assigned to predicted target genes using heuristics. As such, these approaches provide no direct evidence that insertions actually affect their predicted targets or how transcripts of these genes are affected. To address this, we developed IM-Fusion, an approach that identifies insertion sites from gene-transposon fusions in standard single- and paired-end RNA-sequencing data...
June 1, 2017: Nucleic Acids Research
https://www.readbyqxmd.com/read/28557013/unravelling-the-complexity-and-redundancy-of-carbon-catabolic-repression-in-pseudomonas-fluorescens-sbw25
#14
Yunhao Liu, Chaitanya S Gokhale, Paul B Rainey, Xue-Xian Zhang
The two-component system CbrAB is the principal regulator for cellular metabolic balance in Pseudomonas fluorescens SBW25 and is necessary for growth on many substrates including xylose. To understand the regulatory linkage between CbrAB and genes for xylose utilization (xut), we performed transposon mutagenesis of ΔcbrB to select for Xut(+) suppressors. This led to identification of crc and hfq. Subsequent genetic and biochemical analysis showed that Crc and Hfq are key mediators of succinate-provoked carbon catabolite repression (CCR)...
May 29, 2017: Molecular Microbiology
https://www.readbyqxmd.com/read/28550060/surface-sensing-for-paenibacillus-sp-naist15-1-flagellar-gene-expression-on-solid-medium
#15
Kazuo Kobayashi, Yu Kanesaki, Hirofumi Yoshikawa
A rhizosphere Gram-positive bacterial isolate, Paenibacillus sp. NAIST15-1, exhibits intriguing motility behavior on hard agar medium. Paenibacillus sp. shows increased transcription of flagellar genes and hyperflagellation when transferred from liquid to solid medium. Hyperflagellated cells form wandering colonies that are capable of moving around on the surface of medium containing ≥1.5% agar. Transposon mutagenesis was used to identify genes critical for motility. In addition to flagellar genes, this mutagenesis identified five nonflagellar structural genes that were important for motility...
August 1, 2017: Applied and Environmental Microbiology
https://www.readbyqxmd.com/read/28515302/transposon-mutagenesis-of-the-zika-virus-genome-highlights-regions-essential-for-rna-replication-and-restricted-for-immune-evasion
#16
Benjamin O Fulton, David Sachs, Megan C Schwarz, Peter Palese, Matthew J Evans
The molecular constraints affecting Zika virus (ZIKV) evolution are not well understood. To investigate ZIKV genetic flexibility, we used transposon mutagenesis to add 15-nucleotide insertions throughout the ZIKV MR766 genome and subsequently deep sequenced the viable mutants. Few ZIKV insertion mutants replicated, which likely reflects a high degree of functional constraints on the genome. The NS1 gene exhibited distinct mutational tolerances at different stages of the screen. This result may define regions of the NS1 protein that are required for the different stages of the viral life cycle...
August 1, 2017: Journal of Virology
https://www.readbyqxmd.com/read/28512090/the-vibrio-cholerae-vexgh-rnd-efflux-system-maintains-cellular-homeostasis-by-effluxing-vibriobactin
#17
Dillon E Kunkle, X Renee Bina, James E Bina
Resistance-nodulation-division (RND) superfamily efflux systems have been widely studied for their role in antibiotic resistance, but their native biological functions remain poorly understood. We previously showed that loss of RND-mediated efflux in Vibrio cholerae resulted in activation of the Cpx two-component regulatory system, which mediates adaptation to stress resulting from misfolded membrane proteins. Here, we investigated the mechanism linking RND-mediated efflux to the Cpx response. We performed transposon mutagenesis screening of RND-deficient V...
May 16, 2017: MBio
https://www.readbyqxmd.com/read/28509393/nmea-a-novel-efflux-transporter-specific-for-nucleobases-and-nucleosides-contributes-to-metal-resistance-in-aspergillus-nidulans
#18
Sofia Balaska, Vassilios Myrianthopoulos, Martha Tselika, Dimitris G Hatzinikolaou, Emmanuel Mikros, George Diallinas
Through Minos transposon mutagenesis we obtained A. nidulans mutants resistant to 5-fluorouracil due to insertions into the upstream region of the uncharacterized gene nmeA, encoding a Major Facilitator Superfamily (MFS) transporter. Minos transpositions increased nmeA transcription, which is otherwise extremely low under all conditions tested. To dissect the function of NmeA we used strains overexpressing or genetically lacking the nmeA gene. Strains overexpressing NmeA are resistant to toxic purine analogues, but also, to cadmium, zinc and borate, whereas an isogenic nmeAΔ null mutant exhibits increased sensitivity to these compounds...
May 16, 2017: Molecular Microbiology
https://www.readbyqxmd.com/read/28507068/genome-wide-survey-of-pseudomonas-aeruginosa-pa14-reveals-a-role-for-the-glyoxylate-pathway-and-extracellular-proteases-in-the-utilization-of-mucin
#19
Jeffrey M Flynn, Chi Phan, Ryan C Hunter
Chronic airway infections by the opportunistic pathogen Pseudomonas aeruginosa are a major cause of mortality in cystic fibrosis (CF) patients. Although this bacterium has been extensively studied for its virulence determinants, biofilm growth, and immune evasion mechanisms, comparatively little is known about the nutrient sources that sustain its growth in vivo Respiratory mucins represent a potentially abundant bioavailable nutrient source, although we have recently shown that canonical pathogens inefficiently use these host glycoproteins as a growth substrate...
August 2017: Infection and Immunity
https://www.readbyqxmd.com/read/28500044/rpon-%C3%AF-54-is-required-for-floc-formation-but-not-for-extracellular-polysaccharide-biosynthesis-in-a-floc-forming-aquincola-tertiaricarbonis-strain
#20
Dianzhen Yu, Ming Xia, Liping Zhang, Yulong Song, You Duan, Tong Yuan, Minjie Yao, Liyou Wu, Chunyuan Tian, Zhenbin Wu, Xiangzhen Li, Jizhong Zhou, Dongru Qiu
Some bacteria are capable of forming flocs, in which bacterial cells become self-flocculated by secreted extracellular polysaccharides and other biopolymers. The floc-forming bacteria play a central role in activated sludge, which has been widely utilized for the treatment of municipal sewage and industrial wastewater. Here, we use a floc-forming bacterium, Aquincolatertiaricarbonis RN12, as a model to explore the biosynthesis of extracellular polysaccharides and the regulation of floc formation. A large gene cluster for exopolysaccharide biosynthesis and a gene encoding the alternative sigma factor RpoN1, one of the four paralogues, have been identified in floc formation-deficient mutants generated by transposon mutagenesis, and the gene functions have been further confirmed by genetic complementation analyses...
July 15, 2017: Applied and Environmental Microbiology
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