ALT-FISH quantifies alternative lengthening of telomeres activity by imaging of single-stranded repeats DOI Creative Commons
Lukas Frank, Anne Rademacher, Norbert Mücke

et al.

Nucleic Acids Research, Journal Year: 2022, Volume and Issue: 50(11), P. e61 - e61

Published: Feb. 5, 2022

Alternative lengthening of telomeres (ALT) occurs in ∼10% cancer entities. However, little is known about the heterogeneity ALT activity since robust detection assays with high-throughput situ readouts are lacking. Here, we introduce ALT-FISH, a method to quantitate single cells from accumulation single-stranded telomeric DNA and RNA. It involves one-step fluorescent hybridization approach followed by fluorescence microscopy imaging. Our reliably identified cell lines different tumor entities was validated three established models induction suppression. Furthermore, successfully applied ALT-FISH spatially resolve primary tissue sections leiomyosarcoma neuroblastoma tumors. Thus, our assay provides insights into tumors suited for applications, which will facilitate screening ALT-specific drugs.

Language: Английский

Spatial organization of translation and translational repression in two phases of germ granules DOI Creative Commons
Anne Ramat,

Ali Haidar,

Céline Garret

et al.

Nature Communications, Journal Year: 2024, Volume and Issue: 15(1)

Published: Sept. 13, 2024

Language: Английский

Citations

7
miR-7 controls glutamatergic transmission and neuronal connectivity in a Cdr1as-dependent manner DOI Creative Commons
C Cerda, Seung Joon Kim,

Gwendolin Thomas

et al.

EMBO Reports, Journal Year: 2024, Volume and Issue: 25(7), P. 3008 - 3039

Published: June 3, 2024

Abstract The circular RNA (circRNA) Cdr1as is conserved across mammals and highly expressed in neurons, where it directly interacts with microRNA miR-7. However, the biological function of this interaction unknown. Here, using primary cortical murine we demonstrate that stimulating neurons by sustained depolarization rapidly induces two-fold transcriptional upregulation strong post-transcriptional stabilization loss causes doubling glutamate release from stimulated synapses increased frequency duration local neuronal bursts. Moreover, periodicity networks increases, synchronicity impaired. Strikingly, these effects are reverted expression miR-7, which also clears molecules projections. Consistently, without Cdr1as, transcriptomic changes caused miR-7 overexpression stronger (including miR-7-targets downregulation) enriched secretion/synaptic plasticity pathways. Altogether, our results suggest buffers activity to control glutamatergic excitatory transmission connectivity important for long-lasting synaptic adaptations.

Language: Английский

Citations

6
Wnt Signaling Coordinates the Expression of Limb Patterning Genes During Axolotl Forelimb Development and Regeneration DOI Creative Commons

Alexander M. Lovely,

Timothy J. Duerr,

Qingchao Qiu

et al.

Frontiers in Cell and Developmental Biology, Journal Year: 2022, Volume and Issue: 10

Published: April 21, 2022

After amputation, axolotl salamanders can regenerate their limbs, but the degree to which limb regeneration recapitulates development remains unclear. One limitation in answering this question is our lack of knowledge about salamander development. Here, we address by studying expression patterns genes important for patterning during and regeneration. We focus on Wnt signaling pathway because it regulates multiple functions tetrapod development, including bud initiation, outgrowth, patterning, skeletal differentiation. use fluorescence situ hybridization show ligands, receptors, developing regenerating limbs. Inhibition ligand secretion permanently blocks outgrowth when treated early Inhibiting decreases critical genes, Fgf10, Fgf8, Shh, leading reduced limb. Patterns gene are similar between impacted similarly. Overall, findings suggest that utilize It also provides new insights into interaction with other pathways

Language: Английский

Citations

26
Circadian plasticity evolves through regulatory changes in a neuropeptide gene DOI Creative Commons
Michael P. Shahandeh,

Liliane Abuin,

Lou Lescuyer De Decker

et al.

Nature, Journal Year: 2024, Volume and Issue: 635(8040), P. 951 - 959

Published: Oct. 16, 2024

Many organisms, including cosmopolitan drosophilids, show circadian plasticity, varying their activity with changing dawn–dusk intervals1. How this behaviour evolves is unclear. Here we compare Drosophila melanogaster sechellia, an equatorial, ecological specialist that experiences minimal photoperiod variation, to investigate the mechanistic basis of plasticity evolution2. D. sechellia has lost ability delay its evening peak time under long photoperiods. Screening mutants in melanogaster/D. hybrids identifies a contribution neuropeptide pigment-dispersing factor (Pdf) loss. Pdf exhibits species-specific temporal expression, due part cis-regulatory divergence. RNA interference and rescue experiments using regulatory sequences demonstrate modulation neuropeptide's expression affects degree behavioural plasticity. The region signals selection across populations from different latitudes. We provide evidence confers selective advantage for at elevated latitude, whereas probably suffers fitness costs through reduced copulation success outside range. Our findings highlight gene as hotspot locus evolution might have contributed both melanogaster's global distribution sechellia's specialization. diverged Pdf, conferring

Language: Английский

Citations

5
OME-Zarr: a cloud-optimized bioimaging file format with international community support DOI Creative Commons
Josh Moore, Daniela Basurto-Lozada, Sébastien Besson

et al.

bioRxiv (Cold Spring Harbor Laboratory), Journal Year: 2023, Volume and Issue: unknown

Published: Feb. 21, 2023

A growing community is constructing a next-generation file format (NGFF) for bioimaging to overcome problems of scalability and heterogeneity. Organized by the Open Microscopy Environment (OME), individuals institutes across diverse modalities facing these have designed specification process (OME-NGFF) address needs. This paper brings together wide range those members describe cloud-optimized itself -- OME-Zarr along with tools data resources available today increase FAIR access remove barriers in scientific process. The current momentum offers an opportunity unify key component domain that underlies so many personal, institutional, global management analysis tasks.

Language: Английский

Citations

11
Piscis: a novel loss estimator of the F1 score enables accurate spot detection in fluorescence microscopy images via deep learning DOI Creative Commons
Zijian Niu, Aoife O’Farrell, Jingxin Li

et al.

bioRxiv (Cold Spring Harbor Laboratory), Journal Year: 2024, Volume and Issue: unknown

Published: Jan. 31, 2024

Single-molecule RNA fluorescence in situ hybridization (RNA FISH)-based spatial transcriptomics methods have enabled the accurate quantification of gene expression at single-cell resolution by visualizing transcripts as diffraction-limited spots. While these generally scale to large samples, image analysis remains challenging, often requiring manual parameter tuning. We present Piscis, a fully automatic deep learning algorithm for spot detection trained using novel loss function, SmoothF1 loss, that approximates F1 score directly penalize false positives and negatives but differentiable hence usable training approaches. Piscis was tested on diverse dataset composed 358 manually annotated experimental FISH images representing multiple cell types 240 additional synthetic images. outperforms other state-of-the-art methods, enabling accurate, high-throughput FISH-derived imaging data without need

Language: Английский

Citations

4
Polarity-dependent expression and localization of secretory glucoamylase mRNA in filamentous fungal cells DOI Creative Commons

Yuki Morita,

Kaoru Takegawa, Brett M. Collins

et al.

Microbiological Research, Journal Year: 2024, Volume and Issue: 282, P. 127653 - 127653

Published: Feb. 23, 2024

In multinuclear and multicellular filamentous fungi little is known about how mRNAs encoding secreted enzymes are transcribed localized spatiotemporally. To better understand this process we analyzed mRNA GlaA, a glucoamylase in large amounts by the industrial fungus Aspergillus oryzae, MS2 system, which can be visualized living cells. We found that glaA was significantly near hyphal tip septum, sites of protein secretion, polarity-dependent expression localization manners. also revealed exhibits long-range dynamics vicinity endoplasmic reticulum (ER) manner dependent on microtubule motor proteins kinesin-1 kinesin-3, but independent early endosomes. Moreover, elucidated although to stress granules (SGs) processing bodies (PBs) under high temperature, not seen ER stress, suggesting there different regulatory mechanisms SG PB temperature stress. Collectively, study uncovers dynamic mechanism secretory enzyme fungi.

Language: Английский

Citations

4
High-throughput image processing software for the study of nuclear architecture and gene expression DOI Creative Commons
Adib Keikhosravi,

Faisal Almansour,

Christopher H. Bohrer

et al.

Scientific Reports, Journal Year: 2024, Volume and Issue: 14(1)

Published: Aug. 8, 2024

Abstract High-throughput imaging (HTI) generates complex datasets from a large number of experimental perturbations. Commercial HTI software programs for image analysis workflows typically do not allow full customization and adoption new processing algorithms in the modules. While open-source platforms provide individual modules workflow, like nuclei segmentation, spot detection, or cell tracking, they are often limited integrating novel algorithms. Here, we introduce High-Throughput Image Processing Software (HiTIPS) to expand range existing capabilities. HiTIPS incorporates advanced machine learning automated signal nucleus registration, quantification intensity. Furthermore, features graphical user interface that is open integration pipelines adding To demonstrate utility HiTIPS, present three examples high-throughput DNA FISH, immunofluorescence (IF), live-cell transcription single cells. Altogether, user-friendly, flexible, platform variety biology applications.

Language: Английский

Citations

4
Circadian and circatidal oscillations of clock gene expression in brains of Eurydice pulchra and Parhyale hawaiensis. DOI Creative Commons
Andrew Oliphant,

Chee Ying Sia,

Charalambos P. Kyriacou

et al.

bioRxiv (Cold Spring Harbor Laboratory), Journal Year: 2025, Volume and Issue: unknown

Published: Jan. 13, 2025

Abstract Coastal organisms express daily and tidal rhythms of physiology behaviour to adapt their intertidal environments. Although the molecular-genetic basis circadian clocks driving is well understood, nature mechanism circatidal remain a mystery. Using fluorescent in situ hybridisation, we mapped discrete clusters putative “clock” cells co-expressing canonical clock genes across brains crustaceans Eurydice pulchra Parhyale hawaiensis . Tidally rhythmic, field-collected E. exhibited ∼12-hour ∼24-hours gene expression cell groups. Laboratory-reared P. entrained 24-hour light:dark cycles (LD) exhibiting robust several In showing following entrainment by agitation under LD, was evident some rhythmic groups, but dorsal-lateral cluster ∼12-hour, i.e., circatidal, transcriptional rhythms. This oscillation phased prior agitation, not LD. These brain networks groups with or periodicities tracking LD respectively, reveal neural substrate for interactive generation appropriate complex habitats.

Language: Английский

Citations

0
TrueSpot: A robust automated tool for quantifying signal puncta in fluorescent imaging DOI Creative Commons

Blythe G. Hospelhorn,

Benjamin K. Kesler,

Hossein Jashnsaz

et al.

bioRxiv (Cold Spring Harbor Laboratory), Journal Year: 2025, Volume and Issue: unknown

Published: Jan. 14, 2025

Abstract Characterizing the movement of biomolecules in single cells quantitatively is essential to understanding fundamental biological mechanisms. RNA fluorescent situ hybridization (RNA-FISH) a technique for visualizing fixed using probes. Automated processing resulting images large datasets. Here we demonstrate that our RNA-FISH image tool, TrueSpot, useful automatically detecting locations at molecule resolution. TrueSpot also performs well on with immunofluorescent (IF) and GFP tagged clustered protein targets. Additionally, show 3D spot detection approach substantially outperforms current 2D algorithms.

Language: Английский

Citations

0