Sensory neuron lineage mapping and manipulation in the Drosophila olfactory system

AbstractNervous systems exhibit myriad cell types, but understanding how this diversity arises is hampered by the difficulty to visualize and genetically-interrogate specific lineages, especially at early developmental stages prior to expression of unique molecular markers. Here, we use a genetic immortalization method to analyze the development of sensory neuron lineages in the Drosophila olfactory system, from their origin to terminal differentiation. We apply this approach to first define a fate map of all olfactory lineages and refine the model of temporal patterns of lineage divisions. Taking advantage of a selective marker for the lineage that gives rise to Or67d pheromone-sensing neurons and a genome-wide transcription factor RNAi screen, we identify the spatial and temporal requirements for Pointed, an ETS family member, in this developmental pathway. Transcriptomic analysis of wild-type and Pointed-depleted olfactory tissue reveals a universal requirement for this factor as a switch-like determinant of fates in these sensory lineages.

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F-Box Genes in the Wheat Genome and Expression Profiling in Wheat at Different Developmental Stages

Genes ◽

10.3390/genes11101154 ◽

2020 ◽

Vol 11 (10) ◽

pp. 1154

Author(s):

Min Jeong Hong ◽

Jin-Baek Kim ◽

Yong Weon Seo ◽

Dae Yeon Kim

Keyword(s):

Developmental Stages ◽

Brachypodium Distachyon ◽

Triticum Aestivum L ◽

Wheat Genome ◽

Post Translational Modification ◽

Genome Wide ◽

A Genome ◽

High Sequence Homology ◽

Wide Scale

Genes of the F-box family play specific roles in protein degradation by post-translational modification in several biological processes, including flowering, the regulation of circadian rhythms, photomorphogenesis, seed development, leaf senescence, and hormone signaling. F-box genes have not been previously investigated on a genome-wide scale; however, the establishment of the wheat (Triticum aestivum L.) reference genome sequence enabled a genome-based examination of the F-box genes to be conducted in the present study. In total, 1796 F-box genes were detected in the wheat genome and classified into various subgroups based on their functional C-terminal domain. The F-box genes were distributed among 21 chromosomes and most showed high sequence homology with F-box genes located on the homoeologous chromosomes because of allohexaploidy in the wheat genome. Additionally, a synteny analysis of wheat F-box genes was conducted in rice and Brachypodium distachyon. Transcriptome analysis during various wheat developmental stages and expression analysis by quantitative real-time PCR revealed that some F-box genes were specifically expressed in the vegetative and/or seed developmental stages. A genome-based examination and classification of F-box genes provide an opportunity to elucidate the biological functions of F-box genes in wheat.

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The Antisense Transcriptomes of Human Cells

Science ◽

10.1126/science.1163853 ◽

2008 ◽

Vol 322 (5909) ◽

pp. 1855-1857 ◽

Cited By ~ 345

Author(s):

Yiping He ◽

Bert Vogelstein ◽

Victor E. Velculescu ◽

Nickolas Papadopoulos ◽

Kenneth W. Kinzler

Keyword(s):

Mammalian Cells ◽

Cell Types ◽

Human Cells ◽

Antisense Transcripts ◽

Genome Wide ◽

Human Genes ◽

A Genome ◽

Dna Strand ◽

The Relationship ◽

Rna Transcript

Transcription in mammalian cells can be assessed at a genome-wide level, but it has been difficult to reliably determine whether individual transcripts are derived from the plus or minus strands of chromosomes. This distinction can be critical for understanding the relationship between known transcripts (sense) and the complementary antisense transcripts that may regulate them. Here, we describe a technique that can be used to (i) identify the DNA strand of origin for any particular RNA transcript, and (ii) quantify the number of sense and antisense transcripts from expressed genes at a global level. We examined five different human cell types and in each case found evidence for antisense transcripts in 2900 to 6400 human genes. The distribution of antisense transcripts was distinct from that of sense transcripts, was nonrandom across the genome, and differed among cell types. Antisense transcripts thus appear to be a pervasive feature of human cells, which suggests that they are a fundamental component of gene regulation.

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Integrated computational and experimental identification of p53, KRAS and VHL mutant selection associated with CRISPR-Cas9 editing

10.1101/407767 ◽

2018 ◽

Cited By ~ 2

Author(s):

Sanju Sinha ◽

Karina Barbosa Guerra ◽

Kuoyuan Cheng ◽

Mark DM Leiserson ◽

David M Wilson ◽

...

Keyword(s):

Gene Editing ◽

Mutant Cell ◽

Cell Types ◽

Driver Mutations ◽

Mutant Selection ◽

Cancer Driver ◽

Genome Wide ◽

A Genome ◽

Induced Changes ◽

Different Cell Types

AbstractRecent studies have reported that CRISPR-Cas9 gene editing induces a p53-dependent DNA damage response in primary cells, which may select for cells with oncogenic p53 mutations11,12. It is unclear whether these CRISPR-induced changes are applicable to different cell types, and whether CRISPR gene editing may select for other oncogenic mutations. Addressing these questions, we analyzed genome-wide CRISPR and RNAi screens to systematically chart the mutation selection potential of CRISPR knockouts across the whole exome. Our analysis suggests that CRISPR gene editing can select for mutants of KRAS and VHL, at a level comparable to that reported for p53. These predictions were further validated in a genome-wide manner by analyzing independent CRISPR screens and patients’ tumor data. Finally, we performed a new set of pooled and arrayed CRISPR screens to evaluate the competition between CRISPR-edited isogenic p53 WT and mutant cell lines, which further validated our predictions. In summary, our study systematically charts and points to the potential selection of specific cancer driver mutations during CRISPR-Cas9 gene editing.

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Replication timing maintains the global epigenetic state in human cells

10.1101/2019.12.28.890020 ◽

2019 ◽

Cited By ~ 2

Author(s):

Kyle N. Klein ◽

Peiyao A. Zhao ◽

Xiaowen Lyu ◽

Daniel A. Bartlett ◽

Amar Singh ◽

...

Keyword(s):

Embryonic Stem ◽

Cancer Cell Line ◽

Replication Timing ◽

Cell Types ◽

Control Factor ◽

Epigenetic State ◽

Genome Wide ◽

A Genome ◽

A Cell ◽

Early Replication

AbstractDNA is replicated in a defined temporal order termed the replication timing (RT) program. RT is spatially segregated in the nucleus with early/late replication corresponding to Hi-C A/B chromatin compartments, respectively. Early replication is also associated with active histone modifications and transcriptional permissiveness. However, the mechanistic interplay between RT, chromatin state, and genome compartmentalization is largely unknown. Here we report that RT is central to epigenome maintenance and compartmentalization in both human embryonic stem cells (hESCs) and cancer cell line HCT116. Knockout (KO) of the conserved RT control factor RIF1, rather than causing discrete RT switches as previously suspected, lead to dramatically increased cell to cell heterogeneity of RT genome wide, despite RIF1’s enrichment in late replicating chromatin. RIF1 KO hESCs have a nearly random RT program, unlike all prior RIF1 KO cells, including HCT116, which show localized alterations. Regions that retain RT, which are prevalent in HCT116 but rare in hESCs, consist of large H3K9me3 domains revealing two independent mechanisms of RT regulation that are used to different extents in different cell types. RIF1 KO results in a striking genome wide downregulation of H3K27ac peaks and enrichment of H3K9me3 at large domains that remain late replicating, while H3K27me3 and H3K4me3 are re-distributed genome wide in a cell type specific manner. These histone modification changes coincided with global reorganization of genome compartments, transcription changes and a genome wide strengthening of TAD structures. Inducible degradation of RIF1 revealed that disruption of RT is upstream of genome compartmentalization changes. Our findings demonstrate that disruption of RT leads to widespread epigenetic mis-regulation, supporting previously speculative models in which the timing of chromatin assembly at the replication fork plays a key role in maintaining the global epigenetic state, which in turn drives genome architecture.

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The epigenetic landscape in purified myonuclei from fast and slow muscles

10.1101/2021.02.04.429545 ◽

2021 ◽

Author(s):

Mads Bengtsen ◽

Ivan Myhre Winje ◽

Einar Eftestøl ◽

Johannes Landskron ◽

Chengyi Sun ◽

...

Keyword(s):

Binding Sites ◽

Association Studies ◽

Cell Types ◽

Specific Marker ◽

Fiber Types ◽

Epigenetic Landscape ◽

Super Enhancer ◽

Genome Wide ◽

A Genome ◽

Pericentriolar Material

AbstractMuscle cells have different phenotypes adapted to different usage and can be grossly divided into fast/glycolytic and slow/oxidative types. While most muscles contain a mixture of such fiber types, we aimed at providing a genome-wide analysis of chromatin environment by ChIP-Seq in two muscle extremes, the almost completely fast/glycolytic extensor digitorum longus (EDL) and slow/oxidative soleus muscles. Muscle is a heterogeneous tissue where less than 60% of the nuclei are inside muscle fibers. Since cellular homogeneity is critical in epigenome-wide association studies we devised a new method for purifying skeletal muscle nuclei from whole tissue based on the nuclear envelope protein Pericentriolar material 1 (PCM1) being a specific marker for myonuclei. Using antibody labeling and a magnetic-assisted sorting approach we were able to sort out myonuclei with 95% purity. The sorting eliminated influence from other cell types in the tissue and improved the myo-specific signal. A genome-wide comparison of the epigenetic landscape in EDL and soleus reflected the functional properties of the two muscles each with a distinct regulatory program involving distal enhancers, including a glycolytic super-enhancer in the EDL. The two muscles are also regulated by different sets of transcription factors; e.g. in soleus binding sites for MEF2C, NFATC2 and PPARA were enriched, while in EDL MYOD1 and SOX1 binding sites were found to be overrepresented. In addition, novel factors for muscle regulation such as MAF, ZFX and ZBTB14 were identified.

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Genome-wide survey of the F-box/Kelch (FBK) members and molecular identification of a novel FBK gene TaAFR in wheat

10.1101/2021.04.08.438931 ◽

2021 ◽

Author(s):

Chunru Wei ◽

Weiquan Zhao ◽

Runqiao Fan ◽

Yuyu Meng ◽

Yiming Yang ◽

...

Keyword(s):

Developmental Stages ◽

In Silico Analysis ◽

Digital Pcr ◽

Plant Responses ◽

Plant Signaling ◽

Biotic Stresses ◽

Genome Wide ◽

Cereal Species ◽

A Genome ◽

Genome Wide Survey

F-box proteins play critical roles in plant responses to biotic/abiotic stresses. In the present study, a total of 68 wheat F-box/Kelch ( TaFBK ) gene sequences encoding for 74 proteins were obtained in a genome-wide survey against EnsemblPlants. The 74 TaFBK proteins were divided into 5 categories based on their domain structures. The FBK proteins from wheat, Arabidopsis, and three other cereal species were grouped into 7 clades, and the number of Kelch domains present was their key clustering criterion. Sixty-eight TaFBK genes were unevenly distributed on 21 chromosomes. Most of TaFBKs were predicted to localize in the nucleus and cytoplasm. In silico analysis of a digital PCR revealed that TaFBKs were expressed at multiple developmental stages and tissues, and in response to drought and/or heat stresses. The TaFBK19 gene, a homologous to the Attenuated Far-Red Response ( AFR ) genes in other plant species, and hence named TaAFR , was selected for further analysis. The gene was isolated from the wheat line TcLr15 and its expression evaluated by quantitative real-time PCR. TaAFR transcripts were primarily detected in wheat leaves, and its expression was found to be regulated by various abiotic and biotic stresses as well as plant signaling hormones. Of particular interest, TaAFR expression was differentially regulated in the compatible vs incompatible wheat leaf rust reaction. Subcellular localization studies showed that TaAFR accumulates in the nucleus and cytoplasm. Three TaAFR-interacting proteins were identified experimentally: Skp1/ASK1-like protein (Skp1), ADP-ribosylation factor 2-like isoform X1 (ARL2) and phenylalanine ammonia-lyase (PAL). Further analysis revealed that the Skp1 protein interacted specifically with the F-box domain of TaAFR, while ARL2 and PAL were recognized by the Kelch domain. The data presented herein provides a solid foundation from which the function and metabolic network of TaAFR and other wheat FBKs can be further explored.

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Integrative analysis of epigenetics data identifies gene-specific regulatory elements

10.1101/585125 ◽

2019 ◽

Cited By ~ 2

Author(s):

Florian Schmidt ◽

Alexander Marx ◽

Marie Hebel ◽

Martin Wegner ◽

Nina Baumgarten ◽

...

Keyword(s):

Transcriptional Regulation ◽

Cell Types ◽

Regulatory Elements ◽

Chromatin Accessibility ◽

Genome Wide ◽

A Genome ◽

Gene Level ◽

Regulatory Landscape ◽

Regulatory Sites ◽

Validation Experiments

AbstractUnderstanding the complexity of transcriptional regulation is a major goal of computational biology. Because experimental linkage of regulatory sites to genes is challenging, computational methods considering epigenomics data have been proposed to create tissue-specific regulatory maps. However, we showed that these approaches are not well suited to account for the variations of the regulatory landscape between cell-types. To overcome these drawbacks, we developed a new method called STITCHIT, that identifies and links putative regulatory sites to genes. Within STITCHIT, we consider the chromatin accessibility signal of all samples jointly to identify regions exhibiting a signal variation related to the expression of a distinct gene. STITCHIToutperforms previous approaches in various validation experiments and was used with a genome-wide CRISPR-Cas9 screen to prioritize novel doxorubicin-resistance genes and their associated non-coding regulatory regions. We believe that our work paves the way for a more refined understanding of transcriptional regulation at the gene-level.

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Genome-Wide Analysis of Dynamin Gene Family in cassava (Manihot esculenta Crantz) and Transcriptional Regulation of Family Members ARC5 in Hormonal Treatments

International Journal of Molecular Sciences ◽

10.3390/ijms20205094 ◽

2019 ◽

Vol 20 (20) ◽

pp. 5094

Author(s):

Cao ◽

Liu ◽

Guo ◽

Chen ◽

Li ◽

...

Keyword(s):

Gene Family ◽

Expression Analysis ◽

Defense Mechanism ◽

Developmental Stages ◽

Manihot Esculenta Crantz ◽

Specific Expression ◽

Genome Wide Analysis ◽

Genome Wide ◽

A Genome ◽

Hormonal Treatments

The Dynamin gene family play a significance role in many physiological processes, especially ARC5 (Accumulation and replication of chloroplasts 5) in the process of plastid division. We performed a genome-wide analysis of the cassava Dynamin family based on the published cassava genome sequence and identified ARC5. 23 cassava Dynamins (MeDynamins) were identified and renamed. 23 MeDynamins were further divided into five major groups based on their structural and phylogenetic characteristics. The segmental duplication events have a significant impact on the expansion of MeDynamins. ARC5 expression analysis showed that there were differences between leaves and roots of cassava at different developmental stages. The tissue-specific expression analysis of the MeDynamins showed that most of MeDynamins were expressed in stem apical meristem and embryogenesis, whereas ARC5 was mainly expressed in leaves. The processing of IAA (Indole-3-acetic Acid) and MeJA (Methyl Jasmonate) verified the prediction results of cis-elements, and ACR5 was closely related to plant growth and positively correlated. It also indicated that high concentrations of MeJA treatment caused the cassava defense mechanism to function in advance. In conclusion, these findings provide basic insights for functional validation of the ARC5 genes in exogenous hormonal treatments.

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Genome-wide CRISPR/Cas9-knockout in human induced Pluripotent Stem Cell (iPSC)-derived macrophages

Scientific Reports ◽

10.1038/s41598-021-82137-z ◽

2021 ◽

Vol 11 (1) ◽

Author(s):

Elena Navarro-Guerrero ◽

Chwen Tay ◽

Justin P. Whalley ◽

Sally A. Cowley ◽

Ben Davies ◽

...

Keyword(s):

Cancer Progression ◽

Genome Engineering ◽

Induced Pluripotent Stem Cell ◽

Cell Types ◽

Loss Of Function ◽

Lentiviral Transduction ◽

Genome Wide ◽

A Genome ◽

Human Pathology ◽

Induced Pluripotent

AbstractGenome engineering using CRISPR/Cas9 technology enables simple, efficient and precise genomic modifications in human cells. Conventional immortalized cell lines can be easily edited or screened using genome-wide libraries with lentiviral transduction. However, cell types derived from the differentiation of induced Pluripotent Stem Cells (iPSC), which often represent more relevant, patient-derived models for human pathology, are much more difficult to engineer as CRISPR/Cas9 delivery to these differentiated cells can be inefficient and toxic. Here, we present an efficient, lentiviral transduction protocol for delivery of CRISPR/Cas9 to macrophages derived from human iPSC with efficiencies close to 100%. We demonstrate CRISPR/Cas9 knockouts for three nonessential proof-of-concept genes—HPRT1, PPIB and CDK4. We then scale the protocol and validate for a genome-wide pooled CRISPR/Cas9 loss-of-function screen. This methodology enables, for the first time, systematic exploration of macrophage involvement in immune responses, chronic inflammation, neurodegenerative diseases and cancer progression, using efficient genome editing techniques.

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Identification and characterization of circular RNAs in Ganoderma lucidum

Scientific Reports ◽

10.1038/s41598-019-52932-w ◽

2019 ◽

Vol 9 (1) ◽

Cited By ~ 1

Author(s):

Junjie Shao ◽

Liqiang Wang ◽

Xinyue Liu ◽

Meng Yang ◽

Haimei Chen ◽

...

Keyword(s):

Ganoderma Lucidum ◽

Developmental Stages ◽

Expression Profiles ◽

Circular Rnas ◽

Rna Seq ◽

Polysaccharide Biosynthesis ◽

Genome Wide ◽

A Genome ◽

Positive Correlations

Abstract Circular RNAs (circRNAs) play important roles in animals, plants, and fungi. However, no circRNAs have been reported in Ganoderma lucidum. Here, we carried out a genome-wide identification of the circRNAs in G.lucidum using RNA-Seq data, and analyzed their features. In total, 250 and 2193 circRNAs were identified from strand-specific RNA-seq data generated from the polyA(−) and polyA(−)/RNase R-treated libraries, respectively. Six of 131 (4.58%) predicted circRNAs were experimentally confirmed. Across three developmental stages, 731 exonic circRNAs (back spliced read counts ≥ 5) and their parent genes were further analyzed. CircRNAs were preferred originating from exons with flanking introns, and the lengths of the flanking intron were longer than those of the control introns. A total of 200 circRNAs were differentially expressed across the three developmental stages of G. lucidum. The expression profiles of 119 (16.3%) exonic circRNAs and their parent genes showed significant positive correlations (r ≥ 0.9, q < 0.01), whereas 226 (30.9%) exonic circRNAs and their parent genes exhibited significant negative correlations (r ≤ −0.9, q < 0.01), in which 53 parent genes are potentially involved in the transcriptional regulation, polysaccharide biosynthesis etc. Our results indicated that circRNAs are present in G. lucidum, with potentially important regulatory roles.

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