68 results on '"Sturgill D"'
Search Results
2. 80S translation initiation complex with ac4c(-1) mRNA and Harringtonine
- Author
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Yang, R., primary, Arango, D., additional, Sturgill, D., additional, and Oberdoerffer, S., additional
- Published
- 2022
- Full Text
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3. Identifying Centromeric RNAs Involved in Histone Dynamics In Vivo
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Quénet, D., primary, Sturgill, D., additional, and Dalal, Y., additional
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- 2016
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4. Genetic interactions between Drosophila melanogaster menin and Jun/Fos
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Cerrato A, Parisi M, Santa Anna S, Missirlis F, Guru S, Agarwal S, Sturgill D, Talbot T, Spiegel A, Collins F, Chandrasekharappa S, Marx S, and Oliver B.
- Subjects
endocrine system ,endocrine system diseases - Abstract
Menin is a tumor suppressor required to prevent multiple endocrine neoplasia in humans. Mammalian menin protein is associated with chromatin modifying complexes and has been shown to bind a number of nuclear proteins, including the transcription factor JunD. Menin shows bidirectional effects acting positively on c-Jun and negatively on JunD. We have produced protein null alleles of Drosophila menin (mnn1) and have over expressed the Mnn1 protein. Flies homozygous for protein-null mnn1 alleles are viable and fertile. Localized over-expression of Mnn1 causes defects in thoracic closure, a phenotype that sometimes results from insufficient Jun activity. We observed complex genetic interactions between mnn1 and jun in different developmental settings. Our data support the idea that one function of menin is to modulate Jun activity in a manner dependent on the cellular context.
- Published
- 2006
5. A Waste-Heat Recovery Boiler on a Glass-Melting Furnace
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Nycz, J. A., primary and Sturgill, D. T., additional
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6. Drosophila mojoless, a Retroposed GSK-3, Has Functionally Diverged to Acquire an Essential Role in Male Fertility
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Kalamegham, R., primary, Sturgill, D., additional, Siegfried, E., additional, and Oliver, B., additional
- Published
- 2006
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7. GenoMosaic: on-demand multiple genome comparison and comparative annotation.
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Gibas, C., Sturgill, D., and Weller, J.
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- 2003
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8. Macronutrient Content of a Hypoenergy Diet Affects Nitrogen Retention and Muscle Function in Weight Lifters.
- Author
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Walberg, J. L., Leidy, M. K., Sturgill, D. J., Hinkle, D. E., Ritchey, S. J., and Sebolt, D. R.
- Published
- 1988
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9. Predicting negative priming: effects of schizotypy, cognitive failures, and anxiety
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Sturgill, D. S. and Ferraro, F. R.
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- 1997
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10. 416
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Walberg, J. L., primary, Leidy, M. K., additional, Sturgill, D. J., additional, Hinkle, D. E., additional, Ritchey, S. J., additional, and Sebolt, D. R., additional
- Published
- 1987
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11. GenoMosaic: on-demand multiple genome comparison and comparative annotation
- Author
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Gibas, C., primary, Sturgill, D., additional, and Weller, J., additional
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12. Germline-dependent gene expression in distant non-gonadal somatic tissues of Drosophila
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Zhang Yu, Malone John H, Jallon Jean-Marc, Warren James T, Sturgill David, Gupta Vaijayanti, Parisi Michael J, Gilbert Lawrence I, and Oliver Brian
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Biotechnology ,TP248.13-248.65 ,Genetics ,QH426-470 - Abstract
Abstract Background Drosophila females commit tremendous resources to egg production and males produce some of the longest sperm in the animal kingdom. We know little about the coordinated regulation of gene expression patterns in distant somatic tissues that support the developmental cost of gamete production. Results We determined the non-gonadal gene expression patterns of Drosophila females and males with or without a germline. Our results show that germline-dependent expression in the non-gonadal soma is extensive. Interestingly, gene expression patterns and hormone titers are consistent with a hormone axis between the gonads and non-gonadal soma. Conclusions The germline has a long-range influence on gene expression in the Drosophila sexes. We suggest that this is the result of a germline/soma hormonal axis.
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- 2010
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13. Molecular targets for rapid identification of Brucella spp
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Halling Shirley M, Sriranganathan Nammalwar, Lathigra Raju, He Yongqun, Reichow Sherry A, Ramamoorthy Sheela, Sturgill David M, Ratushna Vladyslava G, Boyle Stephen M, and Gibas Cynthia J
- Subjects
Microbiology ,QR1-502 - Abstract
Abstract Background Brucella is an intracellular pathogen capable of infecting animals and humans. There are six recognized species of Brucella that differ in their host preference. The genomes of the three Brucella species have been recently sequenced. Comparison of the three revealed over 98% sequence similarity at the protein level and enabled computational identification of common and differentiating genes. We validated these computational predictions and examined the expression patterns of the putative unique and differentiating genes, using genomic and reverse transcription PCR. We then screened a set of differentiating genes against classical Brucella biovars and showed the applicability of these regions in the design of diagnostic tests. Results We have identified and tested set of molecular targets that are associated in unique patterns with each of the sequenced Brucella spp. A comprehensive comparison was made among the published genome sequences of B. abortus, B. melitensis and B. suis. The comparison confirmed published differences between the three Brucella genomes, and identified subsets of features that were predicted to be of interest in a functional comparison of B. melitensis and B. suis to B. abortus. Differentiating sequence regions from B. abortus, B. melitensis and B. suis were used to develop PCR primers to test for the existence and in vitro transcription of these genes in these species. Only B. suis is found to have a significant number of unique genes, but combinations of genes and regions that exist in only two out of three genomes and are therefore useful for diagnostics were identified and confirmed. Conclusion Although not all of the differentiating genes identified were transcribed under steady state conditions, a group of genes sufficient to discriminate unambiguously between B. suis, B. melitensis, and B. abortus was identified. We present an overview of these genomic differences and the use of these features to discriminate among a number of Brucella biovars.
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- 2006
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14. Global analysis of X-chromosome dosage compensation
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Malley James D, Dudko Olga K, Doctolero Michael, Nuttall Rachel, Sturgill David, Parisi Michael, Gupta Vaijayanti, Eastman P Scott, and Oliver Brian
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Biology (General) ,QH301-705.5 - Abstract
Abstract Background Drosophila melanogaster females have two X chromosomes and two autosome sets (XX;AA), while males have a single X chromosome and two autosome sets (X;AA). Drosophila male somatic cells compensate for a single copy of the X chromosome by deploying male-specific-lethal (MSL) complexes that increase transcription from the X chromosome. Male germ cells lack MSL complexes, indicating that either germline X-chromosome dosage compensation is MSL-independent, or that germ cells do not carry out dosage compensation. Results To investigate whether dosage compensation occurs in germ cells, we directly assayed X-chromosome transcripts using DNA microarrays and show equivalent expression in XX;AA and X;AA germline tissues. In X;AA germ cells, expression from the single X chromosome is about twice that of a single autosome. This mechanism ensures balanced X-chromosome expression between the sexes and, more importantly, it ensures balanced expression between the single X chromosome and the autosome set. Oddly, the inactivation of an X chromosome in mammalian females reduces the effective X-chromosome dose and means that females face the same X-chromosome transcript deficiency as males. Contrary to most current dosage-compensation models, we also show increased X-chromosome expression in X;AA and XX;AA somatic cells of Caenorhabditis elegans and mice. Conclusion Drosophila germ cells compensate for X-chromosome dose. This occurs by equilibrating X-chromosome and autosome expression in X;AA cells. Increased expression of the X chromosome in X;AA individuals appears to be phylogenetically conserved.
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- 2006
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15. Are the Clinical Outcomes and Cost-Effectiveness of Robot-Assisted Pedicle Screw Placement in Lumbar Fusion Surgery Superior to CT-Navigated and Freehand Fluoroscopy-Guided Techniques? A Systematic Review and Network Meta-Analysis.
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Sturgill D, How J, Blajda T, Davis Z, Ali M, O'Malley G Jr, Patel NV, Khan MF, and Goldstein I
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Introduction: Spinal fusion surgery is known to be an expensive intervention. Although innovative technologies in the field aim at improving operative efficiency and outcomes, total costs must be considered. The authors hope to elucidate any differences between robot-assisted (RA) and CT-navigated (CT-nav) or freehand fluoroscopy-guided (FFG) pedicle screw placement in relation to patient outcomes and cost-effectiveness in lumbar fusion surgery (LFS)., Methods: Following the PRISMA guidelines, the authors performed a systematic review to identify studies comparing clinical outcomes between CT-nav or RA versus FFG in LFS patients. All included studies utilized bilateral pedicle screws. Statistical analysis was performed using R., Results: Of the 1162 identified studies, five were included in the analysis. Direct evidence showed that RA decreased hospital length of stay (LOS) when compared to FFG (MD: -2.67 days; 95% CI: -4.25 to -1.08; p<0.01). Indirect evidence showed that RA decreased operative time when compared to CT-nav (MD: -65.57 mins; 95% CI: -127.7 to -3.44 ; p<0.05). For estimated blood loss (EBL), direct evidence showed that RA was superior to FFG (MD: -120.62 mL; 95% CI: -206.39 to -34.86; p<0.01). However, no significant difference was found between RA and CT-nav for EBL (MD: 14.88 mL; 95% CI: -105.54 to 135.3; p>0.05). There were no other significant differences in ODI, VAS, or complication or re-operation rates between RA and FFG or CT-nav., Conclusion: This study shows that RA pedicle screw placement in LFS provides similar patient outcomes to CT-nav and FFG. Robot-assisted operations were found to give rise to cost savings via decreased LOS when compared to both CT-nav and FFG techniques. Cost-savings of $4,086-$4,865/patient and $7,317-$9,654/patient could be achieved when utilizing RA over CT-nav and FFG, respectively. However, extra upfront and maintenance costs may impact full adoption of RA in LFS., (Copyright © 2024 Elsevier Inc. All rights reserved.)
- Published
- 2024
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16. Detection of ac4C in human mRNA is preserved upon data reassessment.
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Beiki H, Sturgill D, Arango D, Relier S, Schiffers S, and Oberdoerffer S
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- Humans, Reproducibility of Results, RNA, Messenger genetics, Cytidine analogs & derivatives, Nucleotides
- Abstract
We recently reported the distribution of N4-acetylcytidine (ac4C) in HeLa mRNA at base resolution through chemical reduction and the induction of C:T mismatches in sequencing (RedaC:T-seq). Our results contradicted an earlier report from Schwartz and colleagues utilizing a similar method termed ac4C-seq. Here, we revisit both datasets and reaffirm our findings. Through RedaC:T-seq reanalysis, we establish a low basal error rate at unmodified nucleotides that is not skewed to any specific mismatch type and a prominent increase in C:T substitutions as the dominant mismatch type in both treated wild-type replicates, with a high degree of reproducibility across replicates. In contrast, through ac4C-seq reanalysis, we uncover significant data quality issues including insufficient depth, with one wild-type replicate yielding 2.7 million reads, inconsistencies in reduction efficiencies between replicates, and an overall increase in mismatches involving thymine that could obscure ac4C detection. These analyses bolster the detection of ac4C in HeLa mRNA through RedaC:T-seq., Competing Interests: Declaration of interests The authors declare no competing interests., (Published by Elsevier Inc.)
- Published
- 2024
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17. PancrESS - a meta-analysis resource for understanding cell-type specific expression in the human pancreas.
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Sturgill D, Wang L, and Arda HE
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- Humans, Single-Cell Analysis methods, Transcriptome, Pancreas, Gene Expression Profiling methods, Sequence Analysis, RNA methods, Software, Pancreatic Diseases
- Abstract
Background: The human pancreas is composed of specialized cell types producing hormones and enzymes critical to human health. These specialized functions are the result of cell type-specific transcriptional programs which manifest in cell-specific gene expression. Understanding these programs is essential to developing therapies for pancreatic disorders. Transcription in the human pancreas has been widely studied by single-cell RNA technologies, however the diversity of protocols and analysis methods hinders their interpretability in the aggregate., Results: In this work, we perform a meta-analysis of pancreatic single-cell RNA sequencing data. We present a database for reference transcriptome abundances and cell-type specificity metrics. This database facilitates the identification and definition of marker genes within the pancreas. Additionally, we introduce a versatile tool which is freely available as an R package, and should permit integration into existing workflows. Our tool accepts count data files generated by widely-used single-cell gene expression platforms in their original format, eliminating an additional pre-formatting step. Although we designed it to calculate expression specificity of pancreas cell types, our tool is agnostic to the biological source of count data, extending its applicability to other biological systems., Conclusions: Our findings enhance the current understanding of expression specificity within the pancreas, surpassing previous work in terms of scope and detail. Furthermore, our database and tool enable researchers to perform similar calculations in diverse biological systems, expanding the applicability of marker gene identification and facilitating comparative analyses., (© 2024. This is a U.S. Government work and not under copyright protection in the US; foreign copyright protection may apply.)
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- 2024
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18. Post-acute Withdrawal Syndrome.
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Grover C, Sturgill D, and Goldman L
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- Male, Humans, Adult, Narcotic Antagonists therapeutic use, Acute Disease, Analgesics, Opioid therapeutic use, Buprenorphine therapeutic use, Substance Withdrawal Syndrome diagnosis, Substance Withdrawal Syndrome drug therapy, Opioid-Related Disorders drug therapy
- Abstract
Post-acute withdrawal syndrome is an underresearched topic that affects many patients with substance use disorders after the cessation of substance use. Post-acute withdrawal syndrome is exemplified by the occurrence of substance-specific withdrawal signs and symptoms lasting well past the known timeframe for acute withdrawal of a used substance. Post-acute withdrawal syndrome may also include signs and symptoms that are not substance specific that persist, evolve, or appear well past the expected acute withdrawal timeframe. Unfortunately, there is very little scientific literature on post-acute withdrawal syndrome. As a result of this, there are no diagnostic criteria for post-acute withdrawal syndrome, and there is no consensus on the proper name for the condition. We present a case of a 38-year-old man who developed post-acute withdrawal syndrome after treatment with buprenorphine for opioid use disorder and review the limited existing literature on post-acute withdrawal syndrome., Competing Interests: The authors report no conflicts of interest., (Copyright © 2022 American Society of Addiction Medicine.)
- Published
- 2023
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19. Protocol for base resolution mapping of ac4C using RedaC:T-seq.
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Sturgill D, Arango D, and Oberdoerffer S
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- DNA, Complementary, Thymidine, Cytidine, High-Throughput Nucleotide Sequencing
- Abstract
N4-acetylcytidine (ac4C) is an mRNA modification catalyzed by the enzyme N-acetyltransferase 10 (NAT10), with position-dependent effects on mRNA translation. This protocol details a procedure to map ac4C at base resolution using NaBH
4 -induced reduction of ac4C and conversion to thymidine followed by sequencing (RedaC:T-seq). Total RNA is ribodepleted and then treated with NaBH4 to reduce ac4C to tetrahydro-ac4C, which specifically alters base pairing during cDNA synthesis, allowing the detection of ac4C at positions called as thymidine following Illumina sequencing. For complete details on the use and execution of this protocol, please refer to Arango et al. (2022).1 ., Competing Interests: Declaration of interests The authors declare no competing interests., (Published by Elsevier Inc.)- Published
- 2022
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20. Direct epitranscriptomic regulation of mammalian translation initiation through N4-acetylcytidine.
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Arango D, Sturgill D, Yang R, Kanai T, Bauer P, Roy J, Wang Z, Hosogane M, Schiffers S, and Oberdoerffer S
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- 5' Untranslated Regions, Animals, Codon, Initiator, Mammals metabolism, Peptide Chain Initiation, Translational, RNA, Messenger genetics, RNA, Messenger metabolism, Cytidine analogs & derivatives, Cytidine genetics, Protein Biosynthesis
- Abstract
mRNA function is influenced by modifications that modulate canonical nucleobase behavior. We show that a single modification mediates distinct impacts on mRNA translation in a position-dependent manner. Although cytidine acetylation (ac4C) within protein-coding sequences stimulates translation, ac4C within 5' UTRs impacts protein synthesis at the level of initiation. 5' UTR acetylation promotes initiation at upstream sequences, competitively inhibiting annotated start codons. Acetylation further directly impedes initiation at optimal AUG contexts: ac4C within AUG-flanking Kozak sequences reduced initiation in base-resolved transcriptome-wide HeLa results and in vitro utilizing substrates with site-specific ac4C incorporation. Cryo-EM of mammalian 80S initiation complexes revealed that ac4C in the -1 position adjacent to an AUG start codon disrupts an interaction between C and hypermodified t6A at nucleotide 37 of the initiator tRNA. These findings demonstrate the impact of RNA modifications on nucleobase function at a molecular level and introduce mRNA acetylation as a factor regulating translation in a location-specific manner., Competing Interests: Declaration of interests The authors declare no competing interests., (Published by Elsevier Inc.)
- Published
- 2022
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21. Oxalate Nephropathy in a Renal Transplant Recipient After Receiving High Dose Ascorbic Acid.
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Maike A, Sturgill D, and Gallan A
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- Ascorbic Acid adverse effects, Humans, Kidney, Oxalates, Hyperoxaluria chemically induced, Kidney Transplantation adverse effects
- Abstract
Competing Interests: Declaration of Competing Interest The authors disclose no conflicts of interest
- Published
- 2022
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22. Oncogenic lncRNAs alter epigenetic memory at a fragile chromosomal site in human cancer cells.
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Arunkumar G, Baek S, Sturgill D, Bui M, and Dalal Y
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- Carcinogenesis genetics, Centromere metabolism, Centromere Protein A genetics, Centromere Protein A metabolism, Chromatin genetics, Epigenesis, Genetic, Humans, Neoplasms genetics, Neoplasms metabolism, RNA, Long Noncoding genetics
- Abstract
Chromosome instability is a critical event in cancer progression. Histone H3 variant CENP-A plays a fundamental role in defining centromere identity, structure, and function but is innately overexpressed in several types of solid cancers. In the cancer background, excess CENP-A is deposited ectopically on chromosome arms, including 8q24/ cMYC locus, by invading transcription-coupled H3.3 chaperone pathways. Up-regulation of lncRNAs in many cancers correlates with poor prognosis and recurrence in patients. We report that transcription of 8q24-derived oncogenic lncRNAs plays an unanticipated role in altering the 8q24 chromatin landscape by H3.3 chaperone-mediated deposition of CENP-A-associated complexes. Furthermore, a transgene cassette carrying specific 8q24-derived lncRNA integrated into a naïve chromosome locus recruits CENP-A to the new location in a cis-acting manner. These data provide a plausible mechanistic link between locus-specific oncogenic lncRNAs, aberrant local chromatin structure, and the generation of new epigenetic memory at a fragile site in human cancer cells.
- Published
- 2022
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23. Activation of Crtc2/Creb1 in skeletal muscle enhances weight loss during intermittent fasting.
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Bruno NE, Nwachukwu JC, Hughes DC, Srinivasan S, Hawkins R, Sturgill D, Hager GL, Hurst S, Sheu SS, Bodine SC, Conkright MD, and Nettles KW
- Subjects
- Animals, Male, Mice, Mice, Transgenic, Cyclic AMP Response Element-Binding Protein physiology, Energy Metabolism, Fasting, Insulin Resistance, Muscle, Skeletal physiology, Transcription Factors physiology, Weight Loss physiology
- Abstract
The Creb-Regulated Transcriptional Coactivator (Crtc) family of transcriptional coregulators drive Creb1-mediated transcription effects on metabolism in many tissues, but the in vivo effects of Crtc2/Creb1 transcription on skeletal muscle metabolism are not known. Skeletal muscle-specific overexpression of Crtc2 (Crtc2 mice) induced greater mitochondrial activity, metabolic flux capacity for both carbohydrates and fats, improved glucose tolerance and insulin sensitivity, and increased oxidative capacity, supported by upregulation of key metabolic genes. Crtc2 overexpression led to greater weight loss during alternate day fasting (ADF), selective loss of fat rather than lean mass, maintenance of higher energy expenditure during the fast and reduced binge-eating during the feeding period. ADF downregulated most of the mitochondrial electron transport genes, and other regulators of mitochondrial function, that were substantially reversed by Crtc2-driven transcription. Glucocorticoids acted with AMPK to drive atrophy and mitophagy, which was reversed by Crtc2/Creb1 signaling. Crtc2/Creb1-mediated signaling coordinates metabolic adaptations in skeletal muscle that explain how Crtc2/Creb1 regulates metabolism and weight loss., (© 2021 The Authors. The FASEB Journal published by Wiley Periodicals LLC on behalf of Federation of American Societies for Experimental Biology.)
- Published
- 2021
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24. The X chromosome from telomere to telomere: key achievements and future opportunities.
- Author
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Heard E, Johnson AD, Korbel JO, Lee C, Snyder MP, and Sturgill D
- Abstract
While the human genome represents the most accurate vertebrate reference assembly to date, it still contains numerous gaps, including centromeric and other large repeat-containing regions - often termed the "dark side" of the genome - many of which are of fundamental biological importance. Miga et al .
1,2 present the first gapless assembly of the human X chromosome, with the help of ultra-long-read nanopore reads generated for the haploid complete hydatidiform mole (CHM13) genome. They reconstruct the ~3.1 megabase centromeric satellite DNA array and map DNA methylation patterns across complex tandem repeats and satellite arrays. This Telomere-to-Telomere assembly provides a superior human X chromosome reference enabling future sex-determination and X-linked disease research, and provides a path towards finishing the entire human genome sequence., Competing Interests: The authors declare that they have no competing interests., (Copyright: © 2021 Faculty Opinions Ltd.)- Published
- 2021
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25. Point-of-care venous Doppler ultrasound in the management of heart failure and hyponatremia.
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Koratala A and Sturgill D
- Published
- 2021
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26. AKI in Hospitalized Patients with COVID-19 and Seasonal Influenza: A Comparative Analysis.
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Bhasin B, Veitla V, Dawson AZ, Garacci Z, Sturgill D, Ozieh MN, and Regner KR
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- Hospital Mortality, Humans, Retrospective Studies, SARS-CoV-2, Seasons, Acute Kidney Injury epidemiology, COVID-19 epidemiology, Influenza, Human complications
- Abstract
Background: Coronavirus disease 2019 (COVID-19) is often compared with seasonal influenza and the two diseases have similarities, including the risk of systemic manifestations such as AKI. The aim of this study was to perform a comparative analysis of the prevalence, risk factors, and outcomes of AKI in patients who were hospitalized with COVID-19 and influenza., Methods: Retrospective cohort study of patients who were hospitalized with COVID-19 ( n =325) or seasonal influenza ( n =433). AKI was defined by Kidney Disease: Improving Global Outcomes (KDIGO) criteria. Baseline characteristics and hospitalization data were collected, and multivariable analysis was performed to determine the independent predictors for AKI., Results: AKI occurred in 33% of COVID-19 hospitalizations (COV-AKI) and 33% of influenza hospitalizations (FLU-AKI). After adjusting for age, sex, and comorbidity count, the risk of stage 3 AKI was significantly higher in COV-AKI (OR, 3.46; 95% CI, 1.63 to 7.37). Pre-existing CKD was associated with a six- to seven-fold increased likelihood for FLU-AKI and COV-AKI. Mechanical ventilation was associated with a higher likelihood of developing AKI in the COVID-19 cohort (OR, 5.85; 95% CI, 2.30 to 15.63). Black race, after adjustment for comorbidities, was an independent risk for COV-AKI., Conclusions: Pre-existing CKD was a major risk factor for AKI in both cohorts. Black race (independent of comorbidities) and mechanical ventilation were associated with a higher risk of developing COV-AKI, which is characterized by a higher burden of stage 3 AKI and overall poorer prognosis., Competing Interests: M. Ozieh is supported by the National Center for Advancing Translational Sciences, National Institutes of Health award KL2TR001438. K.R. Regner reports having consultancy agreements with Mallinckrodt Pharma and reports receiving honoraria from Mallinckrodt. All remaining authors have nothing to disclose., (Copyright © 2021 by the American Society of Nephrology.)
- Published
- 2021
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27. Acute Kidney Injury Incidence in Hospitalized Patients and Implications for Nutrition Support.
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Meyer D, Mohan A, Subev E, Sarav M, and Sturgill D
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- Adult, Humans, Incidence, Nutritional Support, Risk Factors, Acute Kidney Injury, Hospitalization
- Abstract
Hospitalized adult patients suffer from high rates of acute kidney injury (AKI), which puts them at risk for multiple nutrition problems. The etiology and management strategy of AKI has major implications for the management of the hospitalized patient. The purpose of this review is to understand the incidence and management of AKI in hospitalized adult patients and review the challenges to providing adequate nutrition support., (© 2020 American Society for Parenteral and Enteral Nutrition.)
- Published
- 2020
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28. Hemoglobin Cast Nephropathy.
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Mahmud S, Dernell C, Bal N, Gallan AJ, Blumenthal S, Koratala A, and Sturgill D
- Published
- 2020
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29. Immunoprecipitation and Sequencing of Acetylated RNA.
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Arango D, Sturgill D, and Oberdoerffer S
- Abstract
Generation of the epitranscriptome through chemical modifications of protein-coding messenger RNAs (mRNAs) has emerged as a new mechanism of post-transcriptional gene regulation. While most mRNA modifications are methylation events, a single acetylated ribonucleoside has been described in eukaryotes, occurring at the N4-position of cytidine (N4-acetylcytidine or ac4C). Using a combination of antibody-based enrichment of acetylated regions and deep sequencing, we recently reported ac4C as a novel mRNA modification that is catalyzed by the N-acetyltransferase enzyme NAT10. In this protocol, we describe in detail the procedures to identify acetylated mRNA regions transcriptome-wide using acetylated RNA immunoprecipitation and sequencing (acRIP-seq)., Competing Interests: Competing interestsThe authors declare no competing interests., (Copyright © 2019 The Authors; exclusive licensee Bio-protocol LLC.)
- Published
- 2019
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30. Membraneless nuclear organelles and the search for phases within phases.
- Author
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Sawyer IA, Sturgill D, and Dundr M
- Subjects
- Animals, Cell Nucleus chemistry, Humans, Organelles chemistry, Proteome metabolism, RNA metabolism, Cell Nucleus metabolism, Organelles metabolism
- Abstract
Cells are segregated into two distinct compartment groups to optimize cellular function. The first is characterized by lipid membranes that encapsulate specific regions and regulate macromolecular flux. The second, known collectively as membraneless organelles (MLOs), lacks defining lipid membranes and exhibits self-organizing properties. MLOs are enriched with specific RNAs and proteins that catalyze essential cellular processes. A prominent sub-class of MLOs are known as nuclear bodies, which includes nucleoli, paraspeckles, and other droplets. These microenvironments contain specific RNAs, exhibit archetypal liquid-liquid phase separation characteristics, and harbor intrinsically disordered, multivalent hub proteins. We present an analysis of nuclear body protein disorder that suggests MLO proteomes are significantly more disordered than structured cellular features. We also outline common MLO ultrastructural features, exemplified by the three sub-compartments present inside the nucleolus. A core-shell configuration, or phase within a phase, is displayed by several nuclear bodies and may be functionally important. Finally, we summarize evidence indicating extensive RNA and protein sharing between distinct nuclear bodies, suggesting functional cooperation and similar nucleation principles. Considering the substantial accumulation of specific coding and noncoding RNA classes inside MLOs, evidence that RNA buffers specific phase transition events, and the absence of a clear correlation between total intrinsic protein disorder and MLO accumulation, we conclude that RNA biogenesis may play a key role in MLO formation, internal organization, and function. This article is categorized under: RNA Export and Localization > RNA Localization RNA Interactions with Proteins and Other Molecules > Protein-RNA Interactions: Functional Implications., (© 2018 Wiley Periodicals, Inc.)
- Published
- 2019
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31. Unique palliative care needs of patients with advanced chronic kidney disease - the scope of the problem and several solutions.
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Sturgill D and Bear A
- Subjects
- Humans, Health Services Needs and Demand, Palliative Care methods, Renal Insufficiency, Chronic therapy
- Abstract
Patients with advanced chronic kidney disease (CKD), including end-stage renal disease (ESRD), have a life-threatening illness complicated by high morbidity and mortality and, therefore, should be suitable candidates for early intervention by palliative care specialists. However, the average patient with CKD does not have an advanced care plan, has multiple debilitating symptoms, and does not utilise hospice care at the end of life. In this review, we outline the scope of the problem of unmet palliative care needs for patients with advanced CKD and ESRD, barriers to improving palliative care for patients with renal failure, and possible future directions for palliative nephrology., (© Royal College of Physicians 2019. All rights reserved.)
- Published
- 2019
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32. Acetylation of Cytidine in mRNA Promotes Translation Efficiency.
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Arango D, Sturgill D, Alhusaini N, Dillman AA, Sweet TJ, Hanson G, Hosogane M, Sinclair WR, Nanan KK, Mandler MD, Fox SD, Zengeya TT, Andresson T, Meier JL, Coller J, and Oberdoerffer S
- Subjects
- Acetylation, Cytidine genetics, Cytidine metabolism, HeLa Cells, Humans, N-Terminal Acetyltransferase E genetics, N-Terminal Acetyltransferases, RNA, Messenger genetics, Cytidine analogs & derivatives, N-Terminal Acetyltransferase E metabolism, Protein Biosynthesis, RNA, Messenger metabolism
- Abstract
Generation of the "epitranscriptome" through post-transcriptional ribonucleoside modification embeds a layer of regulatory complexity into RNA structure and function. Here, we describe N4-acetylcytidine (ac4C) as an mRNA modification that is catalyzed by the acetyltransferase NAT10. Transcriptome-wide mapping of ac4C revealed discretely acetylated regions that were enriched within coding sequences. Ablation of NAT10 reduced ac4C detection at the mapped mRNA sites and was globally associated with target mRNA downregulation. Analysis of mRNA half-lives revealed a NAT10-dependent increase in stability in the cohort of acetylated mRNAs. mRNA acetylation was further demonstrated to enhance substrate translation in vitro and in vivo. Codon content analysis within ac4C peaks uncovered a biased representation of cytidine within wobble sites that was empirically determined to influence mRNA decoding efficiency. These findings expand the repertoire of mRNA modifications to include an acetylated residue and establish a role for ac4C in the regulation of mRNA translation., (Published by Elsevier Inc.)
- Published
- 2018
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33. HJURP antagonizes CENP-A mislocalization driven by the H3.3 chaperones HIRA and DAXX.
- Author
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Nye J, Sturgill D, Athwal R, and Dalal Y
- Subjects
- Cell Line, Tumor, Chromosomes, Human, Pair 8 genetics, Co-Repressor Proteins, DNA Damage, Gene Amplification, Humans, Kinetochores metabolism, Mitosis, Molecular Chaperones, Protein Binding, Protein Transport, Proto-Oncogene Proteins c-myc metabolism, Adaptor Proteins, Signal Transducing metabolism, Cell Cycle Proteins metabolism, Centromere Protein A metabolism, DNA-Binding Proteins metabolism, Histone Chaperones metabolism, Histones metabolism, Nuclear Proteins metabolism, Transcription Factors metabolism
- Abstract
The centromere specific histone H3 variant CENP-A/CENH3 specifies where the kinetochore is formed in most eukaryotes. Despite tight regulation of CENP-A levels in normal cells, overexpression of CENP-A is a feature shared by various types of solid tumors and results in its mislocalization to non-centromeric DNA. How CENP-A is assembled ectopically and the consequences of this mislocalization remain topics of high interest. Here, we report that in human colon cancer cells, the H3.3 chaperones HIRA and DAXX promote ectopic CENP-A deposition. Moreover, the correct balance between levels of the centromeric chaperone HJURP and CENP-A is essential to preclude ectopic assembly by H3.3 chaperones. In addition, we find that ectopic localization can recruit kinetochore components, and correlates with mitotic defects and DNA damage in G1 phase. Finally, CENP-A occupancy at the 8q24 locus is also correlated with amplification and overexpression of the MYC gene within that locus. Overall, these data provide insights into the causes and consequences of histone variant mislocalization in human cancer cells., Competing Interests: The authors have declared that no competing interests exist.
- Published
- 2018
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34. Replication Stress Shapes a Protective Chromatin Environment across Fragile Genomic Regions.
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Kim J, Sturgill D, Sebastian R, Khurana S, Tran AD, Edwards GB, Kruswick A, Burkett S, Hosogane EK, Hannon WW, Weyemi U, Bonner WM, Luger K, and Oberdoerffer P
- Subjects
- BRCA1 Protein metabolism, Cell Division genetics, Cells, Cultured, Cellular Senescence genetics, Genomic Instability physiology, Humans, Signal Transduction genetics, Carcinogenesis genetics, Chromatin genetics, DNA Damage genetics, DNA Repair genetics, DNA Replication genetics, Histones genetics, Homologous Recombination genetics
- Abstract
Recent integrative epigenome analyses highlight the importance of functionally distinct chromatin states for accurate cell function. How these states are established and maintained is a matter of intense investigation. Here, we present evidence for DNA damage as an unexpected means to shape a protective chromatin environment at regions of recurrent replication stress (RS). Upon aberrant fork stalling, DNA damage signaling and concomitant H2AX phosphorylation coordinate the FACT-dependent deposition of macroH2A1.2, a histone variant that promotes DNA repair by homologous recombination (HR). MacroH2A1.2, in turn, facilitates the accumulation of the tumor suppressor and HR effector BRCA1 at replication forks to protect from RS-induced DNA damage. Consequently, replicating primary cells steadily accrue macroH2A1.2 at fragile regions, whereas macroH2A1.2 loss in these cells triggers DNA damage signaling-dependent senescence, a hallmark of RS. Altogether, our findings demonstrate that recurrent DNA damage contributes to the chromatin landscape to ensure the epigenomic integrity of dividing cells., (Published by Elsevier Inc.)
- Published
- 2018
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35. Independence between pre-mRNA splicing and DNA methylation in an isogenic minigene resource.
- Author
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Nanan KK, Ocheltree C, Sturgill D, Mandler MD, Prigge M, Varma G, and Oberdoerffer S
- Subjects
- Animals, Chromatin genetics, Chromatin metabolism, DNA (Cytosine-5-)-Methyltransferases metabolism, Gene Expression Regulation, HEK293 Cells, Histones metabolism, Humans, Lysine metabolism, Methylation, Mice, Models, Genetic, DNA Methyltransferase 3B, DNA Methylation, Exons genetics, RNA Precursors genetics, RNA Splicing
- Abstract
Actively transcribed genes adopt a unique chromatin environment with characteristic patterns of enrichment. Within gene bodies, H3K36me3 and cytosine DNA methylation are elevated at exons of spliced genes and have been implicated in the regulation of pre-mRNA splicing. H3K36me3 is further responsive to splicing, wherein splicing inhibition led to a redistribution and general reduction over gene bodies. In contrast, little is known of the mechanisms supporting elevated DNA methylation at actively spliced genic locations. Recent evidence associating the de novo DNA methyltransferase Dnmt3b with H3K36me3-rich chromatin raises the possibility that genic DNA methylation is influenced by splicing-associated H3K36me3. Here, we report the generation of an isogenic resource to test the direct impact of splicing on chromatin. A panel of minigenes of varying splicing potential were integrated into a single FRT site for inducible expression. Profiling of H3K36me3 confirmed the established relationship to splicing, wherein levels were directly correlated with splicing efficiency. In contrast, DNA methylation was equivalently detected across the minigene panel, irrespective of splicing and H3K36me3 status. In addition to revealing a degree of independence between genic H3K36me3 and DNA methylation, these findings highlight the generated minigene panel as a flexible platform for the query of splicing-dependent chromatin modifications., (Published by Oxford University Press on behalf of Nucleic Acids Research 2017.)
- Published
- 2017
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36. Cajal body function in genome organization and transcriptome diversity.
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Sawyer IA, Sturgill D, Sung MH, Hager GL, and Dundr M
- Subjects
- Coiled Bodies metabolism, Humans, Coiled Bodies genetics, Genome, Human, Spliceosomes, Transcriptome
- Abstract
Nuclear bodies contribute to non-random organization of the human genome and nuclear function. Using a major prototypical nuclear body, the Cajal body, as an example, we suggest that these structures assemble at specific gene loci located across the genome as a result of high transcriptional activity. Subsequently, target genes are physically clustered in close proximity in Cajal body-containing cells. However, Cajal bodies are observed in only a limited number of human cell types, including neuronal and cancer cells. Ultimately, Cajal body depletion perturbs splicing kinetics by reducing target small nuclear RNA (snRNA) transcription and limiting the levels of spliceosomal snRNPs, including their modification and turnover following each round of RNA splicing. As such, Cajal bodies are capable of shaping the chromatin interaction landscape and the transcriptome by influencing spliceosome kinetics. Future studies should concentrate on characterizing the direct influence of Cajal bodies upon snRNA gene transcriptional dynamics. Also see the video abstract here., Competing Interests: The authors report no conflict of interest., (© 2016 WILEY Periodicals, Inc.)
- Published
- 2016
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37. Linking Genes and Brain Development of Honeybee Workers: A Whole-Transcriptome Approach.
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Vleurinck C, Raub S, Sturgill D, Oliver B, and Beye M
- Subjects
- Animals, Bees growth & development, Bees metabolism, Brain growth & development, Female, Gene Expression Profiling, Gene Expression Regulation, Developmental, Genes, Insect, Insect Proteins genetics, Male, RNA chemistry, RNA genetics, RNA metabolism, RNA Splicing, Sequence Analysis, RNA, Bees genetics, Brain metabolism, Genetic Linkage, Transcriptome
- Abstract
Honeybees live in complex societies whose capabilities far exceed those of the sum of their single members. This social synergism is achieved mainly by the worker bees, which form a female caste. The worker bees display diverse collaborative behaviors and engage in different behavioral tasks, which are controlled by the central nervous system (CNS). The development of the worker brain is determined by the female sex and the worker caste determination signal. Here, we report on genes that are controlled by sex or by caste during differentiation of the worker's pupal brain. We sequenced and compared transcriptomes from the pupal brains of honeybee workers, queens and drones. We detected 333 genes that are differently expressed and 519 genes that are differentially spliced between the sexes, and 1760 genes that are differentially expressed and 692 genes that are differentially spliced between castes. We further found that 403 genes are differentially regulated by both the sex and caste signals, providing evidence of the integration of both signals through differential gene regulation. In this gene set, we found that the molecular processes of restructuring the cell shape and cell-to-cell signaling are overrepresented. Our approach identified candidate genes that may be involved in brain differentiation that ensures the various social worker behaviors.
- Published
- 2016
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38. Controlled DNA double-strand break induction in mice reveals post-damage transcriptome stability.
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Kim J, Sturgill D, Tran AD, Sinclair DA, and Oberdoerffer P
- Subjects
- Animals, Cells, Cultured, Endodeoxyribonucleases, Genetic Loci, Mice, Mice, Transgenic, Signal Transduction, DNA Breaks, Double-Stranded, Transcriptome
- Abstract
DNA double-strand breaks (DSBs) and their repair can cause extensive epigenetic changes. As a result, DSBs have been proposed to promote transcriptional and, ultimately, physiological dysfunction via both cell-intrinsic and cell-non-autonomous pathways. Studying the consequences of DSBs in higher organisms has, however, been hindered by a scarcity of tools for controlled DSB induction. Here, we describe a mouse model that allows for both tissue-specific and temporally controlled DSB formation at ∼140 defined genomic loci. Using this model, we show that DSBs promote a DNA damage signaling-dependent decrease in gene expression in primary cells specifically at break-bearing genes, which is reversed upon DSB repair. Importantly, we demonstrate that restoration of gene expression can occur independently of cell cycle progression, underlining its relevance for normal tissue maintenance. Consistent with this, we observe no evidence for persistent transcriptional repression in response to a multi-day course of continuous DSB formation and repair in mouse lymphocytes in vivo Together, our findings reveal an unexpected capacity of primary cells to maintain transcriptome integrity in response to DSBs, pointing to a limited role for DNA damage as a mediator of cell-autonomous epigenetic dysfunction., (Published by Oxford University Press on behalf of Nucleic Acids Research 2015. This work is written by (a) US Government employee(s) and is in the public domain in the US.)
- Published
- 2016
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39. Cajal bodies are linked to genome conformation.
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Wang Q, Sawyer IA, Sung MH, Sturgill D, Shevtsov SP, Pegoraro G, Hakim O, Baek S, Hager GL, and Dundr M
- Subjects
- Chromosomes, Human genetics, Epigenesis, Genetic, Genetic Loci, HeLa Cells, Histones genetics, Humans, In Situ Hybridization, Fluorescence, RNA Polymerase II chemistry, RNA Polymerase II metabolism, RNA Splicing genetics, RNA, Small Nuclear genetics, RNA, Small Nucleolar genetics, Reproducibility of Results, Sequence Deletion, Spliceosomes metabolism, Transcription, Genetic, Coiled Bodies genetics, Genome, Human, Nucleic Acid Conformation
- Abstract
The mechanisms underlying nuclear body (NB) formation and their contribution to genome function are unknown. Here we examined the non-random positioning of Cajal bodies (CBs), major NBs involved in spliceosomal snRNP assembly and their role in genome organization. CBs are predominantly located at the periphery of chromosome territories at a multi-chromosome interface. Genome-wide chromosome conformation capture analysis (4C-seq) using CB-interacting loci revealed that CB-associated regions are enriched with highly expressed histone genes and U small nuclear or nucleolar RNA (sn/snoRNA) loci that form intra- and inter-chromosomal clusters. In particular, we observed a number of CB-dependent gene-positioning events on chromosome 1. RNAi-mediated disassembly of CBs disrupts the CB-targeting gene clusters and suppresses the expression of U sn/snoRNA and histone genes. This loss of spliceosomal snRNP production results in increased splicing noise, even in CB-distal regions. Therefore, we conclude that CBs contribute to genome organization with global effects on gene expression and RNA splicing fidelity.
- Published
- 2016
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40. TET-catalyzed oxidation of intragenic 5-methylcytosine regulates CTCF-dependent alternative splicing.
- Author
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Marina RJ, Sturgill D, Bailly MA, Thenoz M, Varma G, Prigge MF, Nanan KK, Shukla S, Haque N, and Oberdoerffer S
- Subjects
- CCCTC-Binding Factor, Cell Line, Dioxygenases, Humans, Mixed Function Oxygenases, Oxidation-Reduction, 5-Methylcytosine metabolism, Alternative Splicing, DNA-Binding Proteins metabolism, Proto-Oncogene Proteins metabolism, Repressor Proteins metabolism
- Abstract
Intragenic 5-methylcytosine and CTCF mediate opposing effects on pre-mRNA splicing: CTCF promotes inclusion of weak upstream exons through RNA polymerase II pausing, whereas 5-methylcytosine evicts CTCF, leading to exon exclusion. However, the mechanisms governing dynamic DNA methylation at CTCF-binding sites were unclear. Here, we reveal the methylcytosine dioxygenases TET1 and TET2 as active regulators of CTCF-mediated alternative splicing through conversion of 5-methylcytosine to its oxidation derivatives. 5-hydroxymethylcytosine and 5-carboxylcytosine are enriched at an intragenic CTCF-binding sites in the CD45 model gene and are associated with alternative exon inclusion. Reduced TET levels culminate in increased 5-methylcytosine, resulting in CTCF eviction and exon exclusion. In vitro analyses establish the oxidation derivatives are not sufficient to stimulate splicing, but efficiently promote CTCF association. We further show genomewide that reciprocal exchange of 5-hydroxymethylcytosine and 5-methylcytosine at downstream CTCF-binding sites is a general feature of alternative splicing in naïve and activated CD4(+) T cells. These findings significantly expand our current concept of the pre-mRNA "splicing code" to include dynamic intragenic DNA methylation catalyzed by the TET proteins., (Published 2015. This article is a U.S. Government work and is in the public domain in the USA.)
- Published
- 2016
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41. Identifying Centromeric RNAs Involved in Histone Dynamics In Vivo.
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Quénet D, Sturgill D, and Dalal Y
- Subjects
- Animals, Autoantigens analysis, Autoantigens metabolism, Cell Culture Techniques methods, Centromere chemistry, Centromere Protein A, Chromatin chemistry, Chromatin metabolism, Chromosomal Proteins, Non-Histone analysis, Chromosomal Proteins, Non-Histone metabolism, Fluorescent Antibody Technique methods, HeLa Cells, Histones analysis, Humans, Microscopy, Fluorescence methods, RNA, Untranslated analysis, RNA, Untranslated genetics, Sequence Analysis, RNA methods, Centromere metabolism, Histones metabolism, RNA, Untranslated metabolism
- Abstract
Over the last decade, the long accepted dogma that heterochromatin is silent has been challenged by increasing evidence of active transcription in these apocryphally annotated quiescent regions of the genome. The recent discovery of noncoding RNAs (ncRNAs) originating from, or localizing to, centromeres, pericentromeres, and telomeres (ie, constitutive heterochromatin) suggest a potential role for ncRNAs in genome integrity. This new paradigm suggests that ncRNAs may recruit chromatin-binding factors, stabilize the higher order folded state of the chromatin fiber, and participate in regulation of processes such as transcription-mediated nucleosome assembly. Thus, identifying, purifying, and elucidating the function of ncRNAs has the potential to provide key insights into genome organization and is currently a topic of intense experimental investigation., (© 2016 Elsevier Inc. All rights reserved.)
- Published
- 2016
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42. Sxl-Dependent, tra/tra2-Independent Alternative Splicing of the Drosophila melanogaster X-Linked Gene found in neurons.
- Author
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Sun X, Yang H, Sturgill D, Oliver B, Rabinow L, and Samson ML
- Subjects
- Alleles, Animals, Female, Gene Expression Profiling, Gene Expression Regulation, High-Throughput Nucleotide Sequencing, Male, Mutation, Transcriptome, Alternative Splicing, Drosophila Proteins genetics, Drosophila Proteins metabolism, Drosophila melanogaster genetics, Drosophila melanogaster metabolism, Genes, X-Linked, Neurons metabolism, Nuclear Proteins genetics, RNA-Binding Proteins metabolism, Ribonucleoproteins genetics
- Abstract
Somatic sexual determination and behavior in Drosophila melanogaster are under the control of a genetic cascade initiated by Sex lethal (Sxl). In the female soma, SXL RNA-binding protein regulates the splicing of transformer (tra) transcripts into a female-specific form. The RNA-binding protein TRA and its cofactor TRA2 function in concert in females, whereas SXL, TRA, and TRA2 are thought to not function in males. To better understand sex-specific regulation of gene expression, we analyzed male and female head transcriptome datasets for expression levels and splicing, quantifying sex-biased gene expression via RNA-Seq and qPCR. Our data uncouple the effects of Sxl and tra/tra2 in females in the-sex-biased alternative splicing of head transcripts from the X-linked locus found in neurons (fne), encoding a pan-neuronal RNA-binding protein of the ELAV family. We show that FNE protein levels are downregulated by Sxl in female heads, also independently of tra/tra2. We argue that this regulation may have important sexually dimorphic consequences for the regulation of nervous system development or function., (Copyright © 2015 Sun et al.)
- Published
- 2015
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43. Sex- and tissue-specific functions of Drosophila doublesex transcription factor target genes.
- Author
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Clough E, Jimenez E, Kim YA, Whitworth C, Neville MC, Hempel LU, Pavlou HJ, Chen ZX, Sturgill D, Dale RK, Smith HE, Przytycka TM, Goodwin SF, Van Doren M, and Oliver B
- Subjects
- Animals, Animals, Genetically Modified, Binding Sites, Female, Gene Expression Profiling, Gene Expression Regulation, Developmental, Genome, Genome-Wide Association Study, Male, Mice, Phenotype, RNA Interference, Sequence Analysis, DNA, Sex Factors, Transcription Factors metabolism, DNA-Binding Proteins metabolism, Drosophila Proteins metabolism, Drosophila melanogaster genetics, Gene Expression Regulation
- Abstract
Primary sex-determination "switches" evolve rapidly, but Doublesex (DSX)-related transcription factors (DMRTs) act downstream of these switches to control sexual development in most animal species. Drosophila dsx encodes female- and male-specific isoforms (DSX(F) and DSX(M)), but little is known about how dsx controls sexual development, whether DSX(F) and DSX(M) bind different targets, or how DSX proteins direct different outcomes in diverse tissues. We undertook genome-wide analyses to identify DSX targets using in vivo occupancy, binding site prediction, and evolutionary conservation. We find that DSX(F) and DSX(M) bind thousands of the same targets in multiple tissues in both sexes, yet these targets have sex- and tissue-specific functions. Interestingly, DSX targets show considerable overlap with targets identified for mouse DMRT1. DSX targets include transcription factors and signaling pathway components providing for direct and indirect regulation of sex-biased expression., (Copyright © 2014 Elsevier Inc. All rights reserved.)
- Published
- 2014
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44. Diversity and dynamics of the Drosophila transcriptome.
- Author
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Brown JB, Boley N, Eisman R, May GE, Stoiber MH, Duff MO, Booth BW, Wen J, Park S, Suzuki AM, Wan KH, Yu C, Zhang D, Carlson JW, Cherbas L, Eads BD, Miller D, Mockaitis K, Roberts J, Davis CA, Frise E, Hammonds AS, Olson S, Shenker S, Sturgill D, Samsonova AA, Weiszmann R, Robinson G, Hernandez J, Andrews J, Bickel PJ, Carninci P, Cherbas P, Gingeras TR, Hoskins RA, Kaufman TC, Lai EC, Oliver B, Perrimon N, Graveley BR, and Celniker SE
- Subjects
- Alternative Splicing genetics, Animals, Drosophila melanogaster anatomy & histology, Drosophila melanogaster cytology, Female, Male, Molecular Sequence Annotation, Nerve Tissue metabolism, Organ Specificity, Poly A genetics, Polyadenylation, Promoter Regions, Genetic genetics, RNA, Long Noncoding genetics, RNA, Messenger genetics, RNA, Messenger metabolism, Sex Characteristics, Stress, Physiological genetics, Drosophila melanogaster genetics, Gene Expression Profiling, Transcriptome genetics
- Abstract
Animal transcriptomes are dynamic, with each cell type, tissue and organ system expressing an ensemble of transcript isoforms that give rise to substantial diversity. Here we have identified new genes, transcripts and proteins using poly(A)+ RNA sequencing from Drosophila melanogaster in cultured cell lines, dissected organ systems and under environmental perturbations. We found that a small set of mostly neural-specific genes has the potential to encode thousands of transcripts each through extensive alternative promoter usage and RNA splicing. The magnitudes of splicing changes are larger between tissues than between developmental stages, and most sex-specific splicing is gonad-specific. Gonads express hundreds of previously unknown coding and long non-coding RNAs (lncRNAs), some of which are antisense to protein-coding genes and produce short regulatory RNAs. Furthermore, previously identified pervasive intergenic transcription occurs primarily within newly identified introns. The fly transcriptome is substantially more complex than previously recognized, with this complexity arising from combinatorial usage of promoters, splice sites and polyadenylation sites.
- Published
- 2014
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45. Comparative validation of the D. melanogaster modENCODE transcriptome annotation.
- Author
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Chen ZX, Sturgill D, Qu J, Jiang H, Park S, Boley N, Suzuki AM, Fletcher AR, Plachetzki DC, FitzGerald PC, Artieri CG, Atallah J, Barmina O, Brown JB, Blankenburg KP, Clough E, Dasgupta A, Gubbala S, Han Y, Jayaseelan JC, Kalra D, Kim YA, Kovar CL, Lee SL, Li M, Malley JD, Malone JH, Mathew T, Mattiuzzo NR, Munidasa M, Muzny DM, Ongeri F, Perales L, Przytycka TM, Pu LL, Robinson G, Thornton RL, Saada N, Scherer SE, Smith HE, Vinson C, Warner CB, Worley KC, Wu YQ, Zou X, Cherbas P, Kellis M, Eisen MB, Piano F, Kionte K, Fitch DH, Sternberg PW, Cutter AD, Duff MO, Hoskins RA, Graveley BR, Gibbs RA, Bickel PJ, Kopp A, Carninci P, Celniker SE, Oliver B, and Richards S
- Subjects
- Animals, Cluster Analysis, Drosophila melanogaster classification, Evolution, Molecular, Exons, Female, Genome, Insect, Humans, Male, Nucleotide Motifs, Phylogeny, Position-Specific Scoring Matrices, Promoter Regions, Genetic, RNA Editing, RNA Splice Sites, RNA Splicing, Reproducibility of Results, Transcription Initiation Site, Computational Biology methods, Drosophila melanogaster genetics, Gene Expression Profiling, Molecular Sequence Annotation, Transcriptome
- Abstract
Accurate gene model annotation of reference genomes is critical for making them useful. The modENCODE project has improved the D. melanogaster genome annotation by using deep and diverse high-throughput data. Since transcriptional activity that has been evolutionarily conserved is likely to have an advantageous function, we have performed large-scale interspecific comparisons to increase confidence in predicted annotations. To support comparative genomics, we filled in divergence gaps in the Drosophila phylogeny by generating draft genomes for eight new species. For comparative transcriptome analysis, we generated mRNA expression profiles on 81 samples from multiple tissues and developmental stages of 15 Drosophila species, and we performed cap analysis of gene expression in D. melanogaster and D. pseudoobscura. We also describe conservation of four distinct core promoter structures composed of combinations of elements at three positions. Overall, each type of genomic feature shows a characteristic divergence rate relative to neutral models, highlighting the value of multispecies alignment in annotating a target genome that should prove useful in the annotation of other high priority genomes, especially human and other mammalian genomes that are rich in noncoding sequences. We report that the vast majority of elements in the annotation are evolutionarily conserved, indicating that the annotation will be an important springboard for functional genetic testing by the Drosophila community., (© 2014 Chen et al.; Published by Cold Spring Harbor Laboratory Press.)
- Published
- 2014
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46. Design of RNA splicing analysis null models for post hoc filtering of Drosophila head RNA-Seq data with the splicing analysis kit (Spanki).
- Author
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Sturgill D, Malone JH, Sun X, Smith HE, Rabinow L, Samson ML, and Oliver B
- Subjects
- Animals, Base Sequence, Computational Biology, Computer Simulation, Female, Gene Expression Profiling methods, Male, Molecular Sequence Data, Protein Isoforms genetics, RNA, Messenger analysis, RNA, Messenger genetics, Alternative Splicing genetics, Drosophila genetics, Models, Genetic, Sequence Analysis, RNA methods, Software
- Abstract
Background: The production of multiple transcript isoforms from one gene is a major source of transcriptome complexity. RNA-Seq experiments, in which transcripts are converted to cDNA and sequenced, allow the resolution and quantification of alternative transcript isoforms. However, methods to analyze splicing are underdeveloped and errors resulting in incorrect splicing calls occur in every experiment., Results: We used RNA-Seq data to develop sequencing and aligner error models. By applying these error models to known input from simulations, we found that errors result from false alignment to minor splice motifs and antisense stands, shifted junction positions, paralog joining, and repeat induced gaps. By using a series of quantitative and qualitative filters, we eliminated diagnosed errors in the simulation, and applied this to RNA-Seq data from Drosophila melanogaster heads. We used high-confidence junction detections to specifically interrogate local splicing differences between transcripts. This method out-performed commonly used RNA-seq methods to identify known alternative splicing events in the Drosophila sex determination pathway. We describe a flexible software package to perform these tasks called Splicing Analysis Kit (Spanki), available at http://www.cbcb.umd.edu/software/spanki., Conclusions: Splice-junction centric analysis of RNA-Seq data provides advantages in specificity for detection of alternative splicing. Our software provides tools to better understand error profiles in RNA-Seq data and improve inference from this new technology. The splice-junction centric approach that this software enables will provide more accurate estimates of differentially regulated splicing than current tools.
- Published
- 2013
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47. Evidence for compensatory upregulation of expressed X-linked genes in mammals, Caenorhabditis elegans and Drosophila melanogaster.
- Author
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Deng X, Hiatt JB, Nguyen DK, Ercan S, Sturgill D, Hillier LW, Schlesinger F, Davis CA, Reinke VJ, Gingeras TR, Shendure J, Waterston RH, Oliver B, Lieb JD, and Disteche CM
- Subjects
- Animals, Cell Line, Dosage Compensation, Genetic, Drosophila Proteins genetics, Drosophila Proteins metabolism, Female, Gene Expression Profiling, Humans, Male, Mice, Oligonucleotide Array Sequence Analysis, Organ Specificity, Ovary metabolism, RNA Polymerase II metabolism, Testis metabolism, Transcription, Genetic, Up-Regulation, X Chromosome genetics, Caenorhabditis elegans genetics, Drosophila melanogaster genetics, Gene Expression Regulation, Genes, X-Linked
- Abstract
Many animal species use a chromosome-based mechanism of sex determination, which has led to the coordinate evolution of dosage-compensation systems. Dosage compensation not only corrects the imbalance in the number of X chromosomes between the sexes but also is hypothesized to correct dosage imbalance within cells that is due to monoallelic X-linked expression and biallelic autosomal expression, by upregulating X-linked genes twofold (termed 'Ohno's hypothesis'). Although this hypothesis is well supported by expression analyses of individual X-linked genes and by microarray-based transcriptome analyses, it was challenged by a recent study using RNA sequencing and proteomics. We obtained new, independent RNA-seq data, measured RNA polymerase distribution and reanalyzed published expression data in mammals, C. elegans and Drosophila. Our analyses, which take into account the skewed gene content of the X chromosome, support the hypothesis of upregulation of expressed X-linked genes to balance expression of the genome.
- Published
- 2011
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48. The developmental transcriptome of Drosophila melanogaster.
- Author
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Graveley BR, Brooks AN, Carlson JW, Duff MO, Landolin JM, Yang L, Artieri CG, van Baren MJ, Boley N, Booth BW, Brown JB, Cherbas L, Davis CA, Dobin A, Li R, Lin W, Malone JH, Mattiuzzo NR, Miller D, Sturgill D, Tuch BB, Zaleski C, Zhang D, Blanchette M, Dudoit S, Eads B, Green RE, Hammonds A, Jiang L, Kapranov P, Langton L, Perrimon N, Sandler JE, Wan KH, Willingham A, Zhang Y, Zou Y, Andrews J, Bickel PJ, Brenner SE, Brent MR, Cherbas P, Gingeras TR, Hoskins RA, Kaufman TC, Oliver B, and Celniker SE
- Subjects
- Alternative Splicing genetics, Animals, Base Sequence, Drosophila Proteins genetics, Drosophila melanogaster embryology, Exons genetics, Female, Genes, Insect genetics, Genome, Insect genetics, Male, MicroRNAs genetics, Oligonucleotide Array Sequence Analysis, Protein Isoforms genetics, RNA Editing genetics, RNA, Messenger analysis, RNA, Messenger genetics, RNA, Small Untranslated analysis, RNA, Small Untranslated genetics, Sequence Analysis, Sex Characteristics, Drosophila melanogaster genetics, Drosophila melanogaster growth & development, Gene Expression Profiling, Gene Expression Regulation, Developmental genetics, Transcription, Genetic genetics
- Abstract
Drosophila melanogaster is one of the most well studied genetic model organisms; nonetheless, its genome still contains unannotated coding and non-coding genes, transcripts, exons and RNA editing sites. Full discovery and annotation are pre-requisites for understanding how the regulation of transcription, splicing and RNA editing directs the development of this complex organism. Here we used RNA-Seq, tiling microarrays and cDNA sequencing to explore the transcriptome in 30 distinct developmental stages. We identified 111,195 new elements, including thousands of genes, coding and non-coding transcripts, exons, splicing and editing events, and inferred protein isoforms that previously eluded discovery using established experimental, prediction and conservation-based approaches. These data substantially expand the number of known transcribed elements in the Drosophila genome and provide a high-resolution view of transcriptome dynamics throughout development.
- Published
- 2011
- Full Text
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49. Identification of functional elements and regulatory circuits by Drosophila modENCODE.
- Author
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Roy S, Ernst J, Kharchenko PV, Kheradpour P, Negre N, Eaton ML, Landolin JM, Bristow CA, Ma L, Lin MF, Washietl S, Arshinoff BI, Ay F, Meyer PE, Robine N, Washington NL, Di Stefano L, Berezikov E, Brown CD, Candeias R, Carlson JW, Carr A, Jungreis I, Marbach D, Sealfon R, Tolstorukov MY, Will S, Alekseyenko AA, Artieri C, Booth BW, Brooks AN, Dai Q, Davis CA, Duff MO, Feng X, Gorchakov AA, Gu T, Henikoff JG, Kapranov P, Li R, MacAlpine HK, Malone J, Minoda A, Nordman J, Okamura K, Perry M, Powell SK, Riddle NC, Sakai A, Samsonova A, Sandler JE, Schwartz YB, Sher N, Spokony R, Sturgill D, van Baren M, Wan KH, Yang L, Yu C, Feingold E, Good P, Guyer M, Lowdon R, Ahmad K, Andrews J, Berger B, Brenner SE, Brent MR, Cherbas L, Elgin SC, Gingeras TR, Grossman R, Hoskins RA, Kaufman TC, Kent W, Kuroda MI, Orr-Weaver T, Perrimon N, Pirrotta V, Posakony JW, Ren B, Russell S, Cherbas P, Graveley BR, Lewis S, Micklem G, Oliver B, Park PJ, Celniker SE, Henikoff S, Karpen GH, Lai EC, MacAlpine DM, Stein LD, White KP, and Kellis M
- Subjects
- Animals, Binding Sites, Computational Biology methods, Drosophila Proteins genetics, Drosophila Proteins metabolism, Drosophila melanogaster growth & development, Drosophila melanogaster metabolism, Epigenesis, Genetic, Gene Expression Regulation, Genes, Insect, Genomics methods, Histones metabolism, Nucleosomes genetics, Nucleosomes metabolism, Promoter Regions, Genetic, RNA, Small Untranslated genetics, RNA, Small Untranslated metabolism, Transcription Factors metabolism, Transcription, Genetic, Chromatin genetics, Chromatin metabolism, Drosophila melanogaster genetics, Gene Regulatory Networks, Genome, Insect, Molecular Sequence Annotation
- Abstract
To gain insight into how genomic information is translated into cellular and developmental programs, the Drosophila model organism Encyclopedia of DNA Elements (modENCODE) project is comprehensively mapping transcripts, histone modifications, chromosomal proteins, transcription factors, replication proteins and intermediates, and nucleosome properties across a developmental time course and in multiple cell lines. We have generated more than 700 data sets and discovered protein-coding, noncoding, RNA regulatory, replication, and chromatin elements, more than tripling the annotated portion of the Drosophila genome. Correlated activity patterns of these elements reveal a functional regulatory network, which predicts putative new functions for genes, reveals stage- and tissue-specific regulators, and enables gene-expression prediction. Our results provide a foundation for directed experimental and computational studies in Drosophila and related species and also a model for systematic data integration toward comprehensive genomic and functional annotation.
- Published
- 2010
- Full Text
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50. Germline-dependent gene expression in distant non-gonadal somatic tissues of Drosophila.
- Author
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Parisi MJ, Gupta V, Sturgill D, Warren JT, Jallon JM, Malone JH, Zhang Y, Gilbert LI, and Oliver B
- Subjects
- Analysis of Variance, Animals, Drosophila melanogaster metabolism, Feedback, Physiological, Female, Genotype, Gonads, Hormones metabolism, Male, Oligonucleotide Array Sequence Analysis, Sex Characteristics, Sexual Behavior, Animal, Drosophila melanogaster cytology, Drosophila melanogaster genetics, Gene Expression Profiling, Germ Cells metabolism
- Abstract
Background: Drosophila females commit tremendous resources to egg production and males produce some of the longest sperm in the animal kingdom. We know little about the coordinated regulation of gene expression patterns in distant somatic tissues that support the developmental cost of gamete production., Results: We determined the non-gonadal gene expression patterns of Drosophila females and males with or without a germline. Our results show that germline-dependent expression in the non-gonadal soma is extensive. Interestingly, gene expression patterns and hormone titers are consistent with a hormone axis between the gonads and non-gonadal soma., Conclusions: The germline has a long-range influence on gene expression in the Drosophila sexes. We suggest that this is the result of a germline/soma hormonal axis.
- Published
- 2010
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