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Article ; Online: A role for the stringent response in ciprofloxacin resistance in Pseudomonas aeruginosa.

García-Villada, Libertad / Degtyareva, Natalya P / Brooks, Ashley M / Goldberg, Joanna B / Doetsch, Paul W

2024 Volume 14, Issue 1, Page(s) 8598

Abstract: Pseudomonas aeruginosa is a major cause of nosocomial infections and the leading cause of chronic lung infections in cystic fibrosis and chronic obstructive pulmonary disease patients. Antibiotic treatment remains challenging because P. aeruginosa is ... ...

Abstract	Pseudomonas aeruginosa is a major cause of nosocomial infections and the leading cause of chronic lung infections in cystic fibrosis and chronic obstructive pulmonary disease patients. Antibiotic treatment remains challenging because P. aeruginosa is resistant to high concentrations of antibiotics and has a remarkable ability to acquire mutations conferring resistance to multiple groups of antimicrobial agents. Here we report that when P. aeruginosa is plated on ciprofloxacin (cipro) plates, the majority of cipro-resistant (ciproR) colonies observed at and after 48 h of incubation carry mutations in genes related to the Stringent Response (SR). Mutations in one of the major SR components, spoT, were present in approximately 40% of the ciproR isolates. Compared to the wild-type strain, most of these isolates had decreased growth rate, longer lag phase and altered intracellular ppGpp content. Also, 75% of all sequenced mutations were insertions and deletions, with short deletions being the most frequently occurring mutation type. We present evidence that most of the observed mutations are induced on the selective plates in a subpopulation of cells that are not instantly killed by cipro. Our results suggests that the SR may be an important contributor to antibiotic resistance acquisition in P. aeruginosa.
MeSH term(s)	Humans ; Ciprofloxacin/pharmacology ; Pseudomonas aeruginosa/genetics ; Pseudomonas Infections/drug therapy ; Anti-Bacterial Agents/pharmacology ; Bone Plates
Chemical Substances	Ciprofloxacin (5E8K9I0O4U) ; Anti-Bacterial Agents
Language	English
Publishing date	2024-04-13
Publishing country	England
Document type	Journal Article
ZDB-ID	2615211-3
ISSN	2045-2322 ; 2045-2322
ISSN (online)	2045-2322
ISSN	2045-2322
DOI	10.1038/s41598-024-59188-z
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Article ; Online: Insights into Repeated Renal Injury Using RNA-Seq with Two New RPTEC Cell Lines.

Merrick, B Alex / Martin, Negin P / Brooks, Ashley M / Foley, Julie F / Dunlap, Paul E / Ramaiahgari, Sreenivasa / Fannin, Rick D / Gerrish, Kevin E

International journal of molecular sciences

2023 Volume 24, Issue 18

Abstract: Renal proximal tubule epithelial cells (RPTECs) are a primary site for kidney injury. We created two RPTEC lines from CD-1 mice immortalized with hTERT (human telomerase reverse transcriptase) or SV40 LgT antigen (Simian Virus 40 Large T antigen). Our ... ...

Abstract	Renal proximal tubule epithelial cells (RPTECs) are a primary site for kidney injury. We created two RPTEC lines from CD-1 mice immortalized with hTERT (human telomerase reverse transcriptase) or SV40 LgT antigen (Simian Virus 40 Large T antigen). Our hypothesis was that low-level, repeated exposure to subcytotoxic levels of 0.25-2.5 μM cisplatin (CisPt) or 12.5-100 μM aflatoxin B1 (AFB1) would activate distinctive genes and pathways in these two differently immortalized cell lines. RNA-seq showed only LgT cells responded to AFB1 with 1139 differentially expressed genes (DEGs) at 72 h. The data suggested that AFB1 had direct nephrotoxic properties on the LgT cells. However, both the cell lines responded to 2.5 μM CisPt from 3 to 96 h expressing 2000-5000 total DEGs. For CisPt, the findings indicated a coordinated transcriptional program of injury signals and repair from the expression of immune receptors with cytokine and chemokine secretion for leukocyte recruitment; robust expression of synaptic and substrate adhesion molecules (SAMs) facilitating the expression of neural and hormonal receptors, ion channels/transporters, and trophic factors; and the expression of nephrogenesis transcription factors. Pathway analysis supported the concept of a renal repair transcriptome. In summary, these cell lines provide in vitro models for the improved understanding of repeated renal injury and repair mechanisms. High-throughput screening against toxicant libraries should provide a wider perspective of their capabilities in nephrotoxicity.
MeSH term(s)	Humans ; Mice ; Animals ; RNA-Seq ; Cell Line ; Kidney Tubules, Proximal/metabolism ; Epithelial Cells ; Cisplatin/metabolism
Chemical Substances	Cisplatin (Q20Q21Q62J)
Language	English
Publishing date	2023-09-18
Publishing country	Switzerland
Document type	Journal Article
ZDB-ID	2019364-6
ISSN	1422-0067 ; 1422-0067 ; 1661-6596
ISSN (online)	1422-0067
ISSN	1422-0067 ; 1661-6596
DOI	10.3390/ijms241814228
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Article ; Online: Genetic profiling of rat gliomas and cardiac schwannomas from life-time radiofrequency radiation exposure study using a targeted next-generation sequencing gene panel.

Brooks, Ashley M / Vornoli, Andrea / Kovi, Ramesh C / Ton, Thai Vu T / Xu, Miaofei / Mashal, Ahmed / Tibaldi, Eva / Gnudi, Federica / Li, Jian-Liang / Sills, Robert C / Bucher, John R / Mandrioli, Daniele / Belpoggi, Fiorella / Pandiri, Arun R

PloS one

2024 Volume 19, Issue 1, Page(s) e0296699

Abstract: The cancer hazard associated with lifetime exposure to radiofrequency radiation (RFR) was examined in Sprague Dawley (SD) rats at the Ramazzini Institute (RI), Italy. There were increased incidences of gliomas and cardiac schwannomas. The translational ... ...

Abstract	The cancer hazard associated with lifetime exposure to radiofrequency radiation (RFR) was examined in Sprague Dawley (SD) rats at the Ramazzini Institute (RI), Italy. There were increased incidences of gliomas and cardiac schwannomas. The translational relevance of these rare rat tumors for human disease is poorly understood. We examined the genetic alterations in RFR-derived rat tumors through molecular characterization of important cancer genes relevant for human gliomagenesis. A targeted next-generation sequencing (NGS) panel was designed for rats based on the top 23 orthologous human glioma-related genes. Single-nucleotide variants (SNVs) and small insertion and deletions (indels) were characterized in the rat gliomas and cardiac schwannomas. Translational relevance of these genetic alterations in rat tumors to human disease was determined through comparison with the Catalogue of Somatic Mutations in Cancer (COSMIC) database. These data suggest that rat gliomas resulting from life-time exposure to RFR histologically resemble low grade human gliomas but surprisingly no mutations were detected in rat gliomas that had homology to the human IDH1 p.R132 or IDH2 p.R172 suggesting that rat gliomas are primarily wild-type for IDH hotspot mutations implicated in human gliomas. The rat gliomas appear to share some genetic alterations with IDH1 wildtype human gliomas and rat cardiac schwannomas also harbor mutations in some of the queried cancer genes. These data demonstrate that targeted NGS panels based on tumor specific orthologous human cancer driver genes are an important tool to examine the translational relevance of rodent tumors resulting from chronic/life-time rodent bioassays.
MeSH term(s)	Humans ; Rats ; Animals ; Rats, Sprague-Dawley ; Glioma/genetics ; Glioma/pathology ; Mutation ; Neurilemmoma/genetics ; High-Throughput Nucleotide Sequencing/methods ; Isocitrate Dehydrogenase/genetics ; Radiation Exposure ; Brain Neoplasms/pathology
Chemical Substances	Isocitrate Dehydrogenase (EC 1.1.1.41)
Language	English
Publishing date	2024-01-17
Publishing country	United States
Document type	Journal Article
ZDB-ID	2267670-3
ISSN	1932-6203 ; 1932-6203
ISSN (online)	1932-6203
ISSN	1932-6203
DOI	10.1371/journal.pone.0296699
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Article: Mitochondrial membrane potential regulates nuclear DNA methylation and gene expression through phospholipid remodeling.

Mori, Mateus Prates / Lozoya, Oswaldo / Brooks, Ashley M / Grenet, Dagoberto / Nadalutti, Cristina A / Ryback, Birgitta / Huang, Kai Ting / Hasan, Prottoy / Hajnóczky, Gyӧrgy / Santos, Janine H

bioRxiv : the preprint server for biology

2024

Abstract: Maintenance of the mitochondrial inner membrane potential (ΔΨM) is critical for many aspects of mitochondrial function, including mitochondrial protein import and ion homeostasis. While ΔΨM loss and its consequences are well studied, little is known ... ...

Abstract	Maintenance of the mitochondrial inner membrane potential (ΔΨM) is critical for many aspects of mitochondrial function, including mitochondrial protein import and ion homeostasis. While ΔΨM loss and its consequences are well studied, little is known about the effects of increased ΔΨM. In this study, we used cells deleted of
Language	English
Publishing date	2024-01-13
Publishing country	United States
Document type	Preprint
DOI	10.1101/2024.01.12.575075
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Article ; Online: Mitochondrial dysfunction and DNA damage accompany enhanced levels of formaldehyde in cultured primary human fibroblasts.

Nadalutti, Cristina A / Stefanick, Donna F / Zhao, Ming-Lang / Horton, Julie K / Prasad, Rajendra / Brooks, Ashley M / Griffith, Jack D / Wilson, Samuel H

Scientific reports

2020 Volume 10, Issue 1, Page(s) 5575

Abstract: Formaldehyde (FA) is a simple biological aldehyde that is produced inside cells by several processes such as demethylation of DNA and proteins, amino acid metabolism, lipid peroxidation and one carbon metabolism (1-C). Although accumulation of excess FA ... ...

Abstract	Formaldehyde (FA) is a simple biological aldehyde that is produced inside cells by several processes such as demethylation of DNA and proteins, amino acid metabolism, lipid peroxidation and one carbon metabolism (1-C). Although accumulation of excess FA in cells is known to be cytotoxic, it is unknown if an increase in FA level might be associated with mitochondrial dysfunction. We choose to use primary human fibroblasts cells in culture (foreskin, FSK) as a physiological model to gain insight into whether an increase in the level of FA might affect cellular physiology, especially with regard to the mitochondrial compartment. FSK cells were exposed to increasing concentrations of FA, and different cellular parameters were studied. Elevation in intracellular FA level was achieved and was found to be cytotoxic by virtue of both apoptosis and necrosis and was accompanied by both G2/M arrest and reduction in the time spent in S phase. A gene expression assessment by microarray analysis revealed FA affected FSK cells by altering expression of many genes including genes involved in mitochondrial function and electron transport. We were surprised to observe increased DNA double-strand breaks (DSBs) in mitochondria after exposure to FA, as revealed by accumulation of γH2A.X and 53BP1 at mitochondrial DNA foci. This was associated with mitochondrial structural rearrangements, loss of mitochondrial membrane potential and activation of mitophagy. Collectively, these results indicate that an increase in the cellular level of FA can trigger mitochondrial DNA double-strand breaks and dysfunction.
MeSH term(s)	Apoptosis/genetics ; Cell Cycle/genetics ; Cell Line ; DNA Breaks, Double-Stranded ; DNA Damage/genetics ; DNA Repair/genetics ; DNA, Mitochondrial/genetics ; Fibroblasts/metabolism ; Formaldehyde/metabolism ; Humans ; Membrane Potential, Mitochondrial/genetics ; Mitochondria/genetics ; Mitochondria/metabolism ; Reactive Oxygen Species/metabolism
Chemical Substances	DNA, Mitochondrial ; Reactive Oxygen Species ; Formaldehyde (1HG84L3525)
Language	English
Publishing date	2020-03-27
Publishing country	England
Document type	Journal Article ; Research Support, N.I.H., Intramural
ZDB-ID	2615211-3
ISSN	2045-2322 ; 2045-2322
ISSN (online)	2045-2322
ISSN	2045-2322
DOI	10.1038/s41598-020-61477-2
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Article ; Online: Arabidopsis DNA Replication Initiates in Intergenic, AT-Rich Open Chromatin.

Wheeler, Emily / Brooks, Ashley M / Concia, Lorenzo / Vera, Daniel L / Wear, Emily E / LeBlanc, Chantal / Ramu, Umamaheswari / Vaughn, Matthew W / Bass, Hank W / Martienssen, Robert A / Thompson, William F / Hanley-Bowdoin, Linda

Plant physiology

2020 Volume 183, Issue 1, Page(s) 206–220

Abstract: The selection and firing of DNA replication origins play key roles in ensuring that eukaryotes accurately replicate their genomes. This process is not well documented in plants due in large measure to difficulties in working with plant systems. We ... ...

Abstract	The selection and firing of DNA replication origins play key roles in ensuring that eukaryotes accurately replicate their genomes. This process is not well documented in plants due in large measure to difficulties in working with plant systems. We developed a new functional assay to label and map very early replicating loci that must, by definition, include at least a subset of replication origins. Arabidopsis (
MeSH term(s)	Arabidopsis/genetics ; Arabidopsis/metabolism ; Arabidopsis Proteins/genetics ; Arabidopsis Proteins/metabolism ; Chromatin/metabolism ; DNA Replication/genetics ; DNA Replication/physiology ; DNA, Plant/metabolism ; DNA, Plant/physiology ; Replication Origin/genetics ; Replication Origin/physiology
Chemical Substances	Arabidopsis Proteins ; Chromatin ; DNA, Plant
Language	English
Publishing date	2020-03-23
Publishing country	United States
Document type	Journal Article ; Research Support, N.I.H., Extramural ; Research Support, U.S. Gov't, Non-P.H.S.
ZDB-ID	208914-2
ISSN	1532-2548 ; 0032-0889
ISSN (online)	1532-2548
ISSN	0032-0889
DOI	10.1104/pp.19.01520
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Article ; Online: Isolation of Plant Nuclei at Defined Cell Cycle Stages Using EdU Labeling and Flow Cytometry.

Wear, Emily E / Concia, Lorenzo / Brooks, Ashley M / Markham, Emily A / Lee, Tae-Jin / Allen, George C / Thompson, William F / Hanley-Bowdoin, Linda

Methods in molecular biology (Clifton, N.J.)

2016 Volume 1370, Page(s) 69–86

Abstract: 5-Ethynyl-2'-deoxyuridine (EdU) is a nucleoside analog of thymidine that can be rapidly incorporated into replicating DNA in vivo and, subsequently, detected by using "click" chemistry to couple its terminal alkyne group to fluorescent azides such as ... ...

Abstract	5-Ethynyl-2'-deoxyuridine (EdU) is a nucleoside analog of thymidine that can be rapidly incorporated into replicating DNA in vivo and, subsequently, detected by using "click" chemistry to couple its terminal alkyne group to fluorescent azides such as Alexa Fluor 488. Recently, EdU incorporation followed by coupling with a fluorophore has been used to visualize newly synthesized DNA in a wide range of plant species. One particularly useful application is in flow cytometry, where two-parameter sorting can be employed to analyze different phases of the cell cycle, as defined both by total DNA content and the amount of EdU pulse-labeled DNA. This approach allows analysis of the cell cycle without the need for synchronous cell populations, which can be difficult to obtain in many plant systems. The approach presented here, which was developed for fixed, EdU-labeled nuclei, can be used to prepare analytical profiles as well as to make highly purified preparations of G1, S, or G2/M phase nuclei for molecular or biochemical analysis. We present protocols for EdU pulse labeling, tissue fixation and harvesting, nuclei preparation, and flow sorting. Although developed for Arabidopsis suspension cells and maize root tips, these protocols should be modifiable to many other plant systems.
MeSH term(s)	Arabidopsis/cytology ; Arabidopsis/genetics ; Cell Cycle ; Cell Fractionation/methods ; Cell Nucleus/genetics ; Click Chemistry/methods ; DNA Replication ; DNA, Plant/analysis ; DNA, Plant/genetics ; Deoxyuridine/analogs & derivatives ; Deoxyuridine/analysis ; Flow Cytometry/methods ; Fluorescent Dyes/analysis ; Zea mays/cytology ; Zea mays/genetics
Chemical Substances	DNA, Plant ; Fluorescent Dyes ; 5-ethynyl-2'-deoxyuridine (G373S00W2J) ; Deoxyuridine (W78I7AY22C)
Language	English
Publishing date	2016
Publishing country	United States
Document type	Journal Article ; Research Support, U.S. Gov't, Non-P.H.S.
ISSN	1940-6029
ISSN (online)	1940-6029
DOI	10.1007/978-1-4939-3142-2_6
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Article ; Online: Genome-Wide Analysis of the Arabidopsis Replication Timing Program.

Concia, Lorenzo / Brooks, Ashley M / Wheeler, Emily / Zynda, Gregory J / Wear, Emily E / LeBlanc, Chantal / Song, Jawon / Lee, Tae-Jin / Pascuzzi, Pete E / Martienssen, Robert A / Vaughn, Matthew W / Thompson, William F / Hanley-Bowdoin, Linda

Plant physiology

2018 Volume 176, Issue 3, Page(s) 2166–2185

Abstract: Eukaryotes use a temporally regulated process, known as the replication timing program, to ensure that their genomes are fully and accurately duplicated during S phase. Replication timing programs are predictive of genomic features and activity and are ... ...

Abstract	Eukaryotes use a temporally regulated process, known as the replication timing program, to ensure that their genomes are fully and accurately duplicated during S phase. Replication timing programs are predictive of genomic features and activity and are considered to be functional readouts of chromatin organization. Although replication timing programs have been described for yeast and animal systems, much less is known about the temporal regulation of plant DNA replication or its relationship to genome sequence and chromatin structure. We used the thymidine analog, 5-ethynyl-2'-deoxyuridine, in combination with flow sorting and Repli-Seq to describe, at high-resolution, the genome-wide replication timing program for Arabidopsis (
MeSH term(s)	Arabidopsis/genetics ; Chromatin/genetics ; Chromatin/metabolism ; Chromosomes, Plant ; DNA Replication Timing ; DNA Transposable Elements ; Flow Cytometry ; Genome, Plant ; Genome-Wide Association Study ; S Phase/genetics ; Sequence Analysis, DNA/methods
Chemical Substances	Chromatin ; DNA Transposable Elements
Language	English
Publishing date	2018-01-04
Publishing country	United States
Document type	Journal Article ; Research Support, U.S. Gov't, Non-P.H.S.
ZDB-ID	208914-2
ISSN	1532-2548 ; 0032-0889
ISSN (online)	1532-2548
ISSN	0032-0889
DOI	10.1104/pp.17.01537
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Article: Estimation of heritability of benzoxazinoid production in rye (Secale cereale) using gas chromatographic analysis

Brooks, Ashley M / Danehower, David A / Murphy, J. Paul / Reberg‐Horton, S. Chris / Burton, James D

Plant breeding. 2012 Feb., v. 131, no. 1

2012

Abstract: With 4 tables ABSTRACT: The 2,4‐dihydroxy‐1,4‐benzoxazin‐3‐one (DIBOA) content of Secale cereale is strongly associated with allelopathy. This has led to interest in developing allelopathic cultivars with increased DIBOA to improve weed control in this ... ...

Abstract	With 4 tables ABSTRACT: The 2,4‐dihydroxy‐1,4‐benzoxazin‐3‐one (DIBOA) content of Secale cereale is strongly associated with allelopathy. This has led to interest in developing allelopathic cultivars with increased DIBOA to improve weed control in this important cover crop. Objectives of this study were to determine heritability estimates for DIBOA in rye and determine the utility of gas chromatography (GC) as a screening tool in a rye allelopathy breeding programme. A synthetic population of half‐sib families varying in production of DIBOA was analysed. DIBOA concentrations ranged from 0.52 to 1.15 mg/g dwt tissue (mean = 0.70 mg/g dwt). Analysis of variance indicated significant variability for DIBOA content in rye harvested at the flag leaf stage. Year × location × genotype and block (year × location) interactions were also significant. Several genotypes were consistently ‘high’ or ‘low’ DIBOA producers across all locations and years. Narrow sense heritability estimates were 0.18 ± 0.04 SE on a per plot basis and 0.57 ± 0.07 SE on an entry mean basis. GC analysis was determined to be a good system for moderate throughput screening of lines.
Keywords	Secale cereale ; allelopathy ; analysis of variance ; breeding ; crop production ; cultivars ; gas chromatography ; genotype ; heritability ; leaves ; rye ; screening ; synthetic populations ; weed control
Language	English
Dates of publication	2012-02
Size	p. 104-109.
Publishing place	Blackwell Publishing Ltd
Document type	Article
ZDB-ID	232853-7
ISSN	0179-9541
ISSN	0179-9541
DOI	10.1111/j.1439-0523.2011.01885.x
Database	NAL-Catalogue (AGRICOLA)

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Article: Systematic assessment of long-read RNA-seq methods for transcript identification and quantification.

Pardo-Palacios, Francisco J / Wang, Dingjie / Reese, Fairlie / Diekhans, Mark / Carbonell-Sala, Sílvia / Williams, Brian / Loveland, Jane E / De María, Maite / Adams, Matthew S / Balderrama-Gutierrez, Gabriela / Behera, Amit K / Gonzalez, Jose M / Hunt, Toby / Lagarde, Julien / Liang, Cindy E / Li, Haoran / Jerryd Meade, Marcus / Moraga Amador, David A / Prjibelski, Andrey D /

Birol, Inanc / Bostan, Hamed / Brooks, Ashley M / Hasan Çelik, Muhammed / Chen, Ying / Du, Mei R M / Felton, Colette / Göke, Jonathan / Hafezqorani, Saber / Herwig, Ralf / Kawaji, Hideya / Lee, Joseph / Liang Li, Jian / Lienhard, Matthias / Mikheenko, Alla / Mulligan, Dennis / Ming Nip, Ka / Pertea, Mihaela / Ritchie, Matthew E / Sim, Andre D / Tang, Alison D / Kei Wan, Yuk / Wang, Changqing / Wong, Brandon Y / Yang, Chen / Barnes, If / Berry, Andrew / Capella, Salvador / Dhillon, Namrita / Fernandez-Gonzalez, Jose M / Ferrández-Peral, Luis / Garcia-Reyero, Natàlia / Goetz, Stefan / Hernández-Ferrer, Carles / Kondratova, Liudmyla / Liu, Tianyuan / Martinez-Martin, Alessandra / Menor, Carlos / Mestre-Tomás, Jorge / Mudge, Jonathan M / Panayotova, Nedka G / Paniagua, Alejandro / Repchevsky, Dmitry / Rouchka, Eric / Saint-John, Brandon / Sapena, Enrique / Sheynkman, Leon / Laird Smith, Melissa / Suner, Marie-Marthe / Takahashi, Hazuki / Youngworth, Ingrid Ashley / Carninci, Piero / Denslow, Nancy D / Guigó, Roderic / Hunter, Margaret E / Tilgner, Hagen U / Wold, Barbara J / Vollmers, Christopher / Frankish, Adam / Fai Au, Kin / Sheynkman, Gloria M / Mortazavi, Ali / Conesa, Ana / Brooks, Angela N

bioRxiv : the preprint server for biology

2023

Abstract: The Long-read RNA-Seq Genome Annotation Assessment Project (LRGASP) Consortium was formed to evaluate the effectiveness of long-read approaches for transcriptome analysis. The consortium generated over 427 million long-read sequences from cDNA and direct ...

Abstract	The Long-read RNA-Seq Genome Annotation Assessment Project (LRGASP) Consortium was formed to evaluate the effectiveness of long-read approaches for transcriptome analysis. The consortium generated over 427 million long-read sequences from cDNA and direct RNA datasets, encompassing human, mouse, and manatee species, using different protocols and sequencing platforms. These data were utilized by developers to address challenges in transcript isoform detection and quantification, as well as
Language	English
Publishing date	2023-07-27
Publishing country	United States
Document type	Preprint
DOI	10.1101/2023.07.25.550582
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