Tracking Single Cells Motility on Different Substrates

Sharma, Pooja; Lam, Van; Raub, Christopher B.; Chung, Byung Min

doi:10.3390/mps3030056

Cited by 6 publications

(3 citation statements)

References 44 publications

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“…Subsequently, the binarized stack was de-speckled again, and the inconsistencies were fixed using the option “Fill holes”. The wrMTrck plugin was used, as previously described by Sharma et al (2020), to track cells with a migration time of at least 6 h. The plugin was run with the following parameters: minimum particle size at 180, maximum particle size at 2000, maximum particle velocity at 50, the maximum area change at 400, the minimum track length at 18, and fps at 0.0008. The fastest 50% fraction was used for further analyses.…”

Section: Methodsmentioning

confidence: 99%

EphrinA5 regulates cell motility by modulating the targeting of DNMT1 to theNcam1promoter via lncRNA/DNA triplex formation

Yildiz

Kundu

Gehrmann

et al. 2023

Preprint

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Cell-cell communication is mediated by membrane receptors and their cognate ligands, such as the Eph/ephrin system, and dictates physiological processes, including cell proliferation and migration. However, whether and how Eph/ephrin signaling culminates in transcriptional regulation is largely unknown. Epigenetic mechanisms are key for integrating external signals, e.g., from neighboring cells, into the transcriptome. We have previously reported that ephrinA5 stimulation of immortalized cerebellar granule (CB) cells elicits transcriptional changes of lncRNAs and protein-coding genes. LncRNAs represent important adaptors for epigenetic writers through which they regulate gene expression. Here, we investigate the interaction of lncRNA with protein-coding genes by the combined power of in silico modeling of RNA/DNA interactions and respective wet lab approaches, in the context of ephrinA5-dependent regulation of cellular motility. We found that Snhg15, a cancer-related lncRNA, forms a triplex structure with the Ncam1 promoter and interacts with DNMT1. EphrinA5 stimulation leads to reduced Snhg15 expression, diminished Snhg15/DNMT1 interaction and decreased DNMT1 association with the Ncam1 promoter. These findings can explain the attenuated Ncam1 promoter methylation and elevated Ncam1 expression that in turn elicits decreased cell motility of CB cells. Hence, we propose that ephrinA5 influences gene transcription via lncRNA-targeted DNA methylation underlying the regulation of cellular motility.

show abstract

Section: Methodsmentioning

confidence: 99%

EphrinA5 regulates cell motility by modulating the targeting of DNMT1 to theNcam1promoter via lncRNA/DNA triplex formation

Yildiz

Kundu

Gehrmann

et al. 2023

Preprint

View full text Add to dashboard Cite

show abstract

“…Subsequently, the binarized stack was de-speckled again, and the inconsistencies were fixed using the option “Fill holes”. The wrMTrck plugin was used, as previously described by Sharma et al [ 49 ], to track cells with a migration time of at least 6 h. The plugin was run with the following parameters: minimum particle size at 180, maximum particle size at 2000, maximum particle velocity at 50, maximum area change at 400, minimum track length at 18, and fps at 0.0008. The fastest 50% fraction was used for further analyses.…”

Section: Methodsmentioning

confidence: 99%

EphrinA5 regulates cell motility by modulating Snhg15/DNA triplex-dependent targeting of DNMT1 to the Ncam1 promoter

Yildiz,

Kundu,

Gehrmann

et al. 2023

Epigenetics & Chromatin

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Cell–cell communication is mediated by membrane receptors and their ligands, such as the Eph/ephrin system, orchestrating cell migration during development and in diverse cancer types. Epigenetic mechanisms are key for integrating external “signals”, e.g., from neighboring cells, into the transcriptome in health and disease. Previously, we reported ephrinA5 to trigger transcriptional changes of lncRNAs and protein-coding genes in cerebellar granule cells, a cell model for medulloblastoma. LncRNAs represent important adaptors for epigenetic writers through which they regulate gene expression. Here, we investigate a lncRNA-mediated targeting of DNMT1 to specific gene loci by the combined power of in silico modeling of RNA/DNA interactions and wet lab approaches, in the context of the clinically relevant use case of ephrinA5-dependent regulation of cellular motility of cerebellar granule cells. We provide evidence that Snhg15, a cancer-related lncRNA, recruits DNMT1 to the Ncam1 promoter through RNA/DNA triplex structure formation and the interaction with DNMT1. This mediates DNA methylation-dependent silencing of Ncam1, being abolished by ephrinA5 stimulation-triggered reduction of Snhg15 expression. Hence, we here propose a triple helix recognition mechanism, underlying cell motility regulation via lncRNA-targeted DNA methylation in a clinically relevant context. Graphical Abstract

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“…Subsequently, the binarized stack was de-speckled again, and the inconsistencies were xed using the option "Fill holes". The wrMTrck plugin was used, as previously described by Sharma et al [49], to track cells with a migration time of at least 6 h. The plugin was run with the following parameters: minimum particle size at 180, maximum particle size at 2000, maximum particle velocity at 50, the maximum area change at 400, the minimum track length at 18, and fps at 0.0008. The fastest 50% fraction was used for further analyses.…”

Section: Migration Assaymentioning

confidence: 99%

EphrinA5 regulates cell motility by modulating lncRNA/DNA triplex-dependent targeting of DNMT1 to the Ncam1 promoter

Yildiz

Kundu

Gehrmann

et al. 2023

Preprint

View full text Add to dashboard Cite

Background: Cell-cell communication is mediated by membrane receptors and their cognate ligands, such as the Eph/ephrin system, and dictates physiological processes, including cell proliferation and migration. However, whether and how Eph/ephrin signaling culminates in transcriptional regulation is largely unknown. Epigenetic mechanisms are key for integrating external “signals”, e.g., from neighboring cells, into the transcriptome. We have previously reported that ephrinA5 stimulation of immortalized cerebellar granule (CB) cells elicits transcriptional changes of lncRNAs and protein-coding genes. LncRNAs represent important adaptors for epigenetic writers through which they regulate gene expression. Hence, we here aimed to investigate, whether ephrinA5 can act on gene transcription through modulating lncRNA-mediated targeting of the DNA methyltransferase 1 (DNMT1) to gene promoters, thereby regulating cell motility. Results: We analyzed the interaction of lncRNA with protein-coding genes by the combined power of in silico modeling of RNA/DNA interactions and respective wet lab approaches. We found that Snhg15, a cancer-related lncRNA, forms a triplex structure with the Ncam1 promoter and interacts with DNMT1. EphrinA5 stimulation leads to reduced Snhg15 expression, diminished Snhg15/DNMT1 interaction and decreased DNMT1 association with the Ncam1 promoter. These findings can explain the attenuated Ncam1 promoter methylation and elevated Ncam1 expression induced by ephrinA5 stimulation that in turn elicits decreased cell motility of CB cells. Conclusion: Based on our findings, we propose that ephrinA5 influences gene transcription via lncRNA-targeted DNA methylation underlying the regulation of cellular motility. Such mechanism could be relevant in the context of cancerogenic processes, known to involve Eph/ephrin signaling and epigenetic remodelling.

show abstract

Tracking Single Cells Motility on Different Substrates

Cited by 6 publications

References 44 publications

EphrinA5 regulates cell motility by modulating the targeting of DNMT1 to theNcam1promoter via lncRNA/DNA triplex formation

EphrinA5 regulates cell motility by modulating the targeting of DNMT1 to theNcam1promoter via lncRNA/DNA triplex formation

EphrinA5 regulates cell motility by modulating Snhg15/DNA triplex-dependent targeting of DNMT1 to the Ncam1 promoter

EphrinA5 regulates cell motility by modulating lncRNA/DNA triplex-dependent targeting of DNMT1 to the Ncam1 promoter

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