Fast Dynamic in vivo Monitoring of Erk Activity at Single Cell Resolution in DREKA Zebrafish

Mayr, Vanessa; Sturtzel, Caterina; Stadler, Manuela; Grissenberger, Sarah; Distel, Martin

doi:10.3389/fcell.2018.00111

Cited by 35 publications

(49 citation statements)

References 30 publications

(43 reference statements)

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“…Blood vessel development was unaffected in Tg(fli1aep:EKC) transgenic embryos and larvae ( Figures 1B-E). Furthermore, transgenic adults displayed no adverse morphological features and were fertile (data not shown), indicating that EKC does not inhibit Erk-signalling in vivo, or cause developmental phenotypes and consistent with previous findings (Mayr et al, 2018).…”

Section: Generation Of a Zebrafish Ec Ekc Transgenic Linesupporting

confidence: 90%

“…The EKC module that we took advantage of here relies upon the well characterised ERK-dependent transcription factor ELK1, utilising the ERK docking site ( Figure 1A) (Chang et al, 2002;Regot et al, 2014). This reporter has previously been shown to report Erk activity in vivo (de la Cova et al, 2017;Mayr et al, 2018). To visualise real-time Erk-signalling in ECs, we expressed this reporter under the control of an EC-specific promoter (fli1aep (Villefranc et al, 2007)) ( Figures 1A-E).…”

Section: Generation Of a Zebrafish Ec Ekc Transgenic Linementioning

confidence: 99%

“…We chose this model because vessel wounding in 4 dpf larvae results in highly reproducible Vegfa/Kdr/Kdrl signalling-dependent vessel regeneration (Gurevich et al, 2018). Importantly, cell wounding induces rapid ERK-signalling waves in vitro and in vivo in other settings (Matsubayashi et al, 2004;Li et al, 2013;Hiratsuka et al, 2015;Aoki et al, 2017;Mayr et al, 2018). Figure 3G).…”

Section: Vessel Wounding Induces Rapid Erk Activationmentioning

confidence: 99%

“…Thus, the KTR system allows rapid quantifiable measurements of ERK activity based upon subcellular localisation of a fluorescent fusion protein, and is more sensitive to phosphatase-mediated kinase activity downregulation when compared to other commonly used kinase activity reporters. This has been applied to enable dynamic ERK-signalling pulses to be analysed in single-cell resolution both in vitro and also in vivo (Regot et al, 2014;de la Cova et al, 2017;Mayr et al, 2018;Goglia et al, 2020), where it has demonstrated to be of high utility.…”

Section: Introductionmentioning

confidence: 99%

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Live-imaging of endothelial Erk activity reveals dynamic and sequential signalling events during regenerative angiogenesis

Okuda

Keyser

Gurevich

et al. 2020

Preprint

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The formation of new blood vessel networks occurs via angiogenesis during development, tissue repair and disease. Angiogenesis is regulated by intracellular endothelial signalling pathways, induced downstream of Vascular endothelial growth factors (VEGFs) and their receptors (VEGFRs). A major challenge in understanding angiogenesis is interpreting how signalling events occur dynamically within endothelial cell populations during sprouting, proliferation and migration. Erk is a central downstream effector of Vegf-signalling and reports the signalling that drives angiogenesis. We generated a vascular Erk biosensor transgenic line in zebrafish using a kinase translocation reporter that allows live-imaging of Erk-signalling dynamics. We demonstrate the utility of this line to live-image Erk activity during physiologically relevant angiogenic events. Further, we reveal dynamic and sequential endothelial cell Erk-signalling events following blood vessel wounding. Initial signalling is dependent upon Ca2+ in the earliest responding endothelial cells, but is independent of Vegfr-signalling and local inflammation. The sustained regenerative response however, involves a Vegfr-dependent mechanism that initiates concomitant with the wound inflammatory response. This work thus reveals a highly dynamic sequence in regenerative angiogenesis that was not previously appreciated. Altogether, this study demonstrates the utility of a unique biosensor strain for analysing dynamic endothelial Erk-signalling events and validates a new resource for the study of vascular signalling in real-time.

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Section: Generation Of a Zebrafish Ec Ekc Transgenic Linesupporting

confidence: 90%

Section: Generation Of a Zebrafish Ec Ekc Transgenic Linementioning

confidence: 99%

Section: Vessel Wounding Induces Rapid Erk Activationmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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Live-imaging of endothelial Erk activity reveals dynamic and sequential signalling events during regenerative angiogenesis

Okuda

Keyser

Gurevich

et al. 2020

Preprint

Self Cite

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“…An exciting perspective coming up in the near future is the use of kinase FRET biosensors in vivo. This is becoming highly feasible for the substrate-based ERK FRET biosensor (EKAR), which has been employed in C. elegans [54], in zebrafish [55], and in mice [56]. In parallel, conformational FRET biosensors containing the full-length sequence of the kinase could become a promising approach to sense the activation of the kinase.…”

Section: Accepted Manuscriptmentioning

confidence: 99%

Insights into the non-mitotic functions of Aurora kinase A: more than just cell division

Bertolin

Tramier

2019

Cell. Mol. Life Sci.

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AURKA is a serine/threonine kinase overexpressed in several cancers. Originally identified as a protein with multifaceted roles during mitosis, improvements in quantitative microscopy uncovered several nonmitotic roles as well. In physiological conditions, AURKA regulates cilia disassembly, neurite extension, cell motility, DNA replication and senescence programs. In cancer-like contexts, AURKA actively promotes DNA repair, it acts as a transcription factor, promotes cell migration and invasion, and it localises at mitochondria to regulate mitochondrial dynamics and ATP production. Here we review the non-mitotic roles of AURKA, and its partners outside of cell division. In addition, we make an insight on how structural data and quantitative fluorescence microscopy allowed to understand AURKA activation and its interaction with new substrates, highlighting future developments in fluorescence microscopy needed to better understand AURKA functions in vivo. Last, we will recapitulate the most significant AURKA inhibitors currently in clinical trials, and we will explore how the non-mitotic roles of the kinase may provide new insights to ameliorate current pharmacological strategies against AURKA overexpression.

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Intracellular signaling dynamics and their role in coordinating tissue repair

Ghilardi

O'Reilly

Sgro

2020

WIREs Mechanisms of Disease

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Tissue repair is a complex process that requires effective communication and coordination between cells across multiple tissues and organ systems. Two of the initial intracellular signals that encode injury signals and initiate tissue repair responses are calcium and extracellular signal-regulated kinase (ERK). However, calcium and ERK signaling control a variety of cellular behaviors important for injury repair including cellular motility, contractility, and proliferation, as well as the activity of several different transcription factors, making it challenging to relate specific injury signals to their respective repair programs. This knowledge gap ultimately hinders the development of new wound healing therapies that could take advantage of native cellular signaling programs to more effectively repair tissue damage. The objective of this review is to highlight the roles of calcium and ERK signaling dynamics as mechanisms that link specific injury signals to specific cellular repair programs during epithelial and stromal injury repair. We detail how the signaling networks controlling calcium and ERK can now also be dissected using classical signal processing techniques with the advent of new biosensors and optogenetic signal controllers. Finally, we advocate the importance of recognizing calcium and ERK dynamics as key links between injury detection and injury repair programs that both organize and execute a coordinated tissue repair response between cells across different tissues and organs.

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Fast Dynamic in vivo Monitoring of Erk Activity at Single Cell Resolution in DREKA Zebrafish

Cited by 35 publications

References 30 publications

Live-imaging of endothelial Erk activity reveals dynamic and sequential signalling events during regenerative angiogenesis

Live-imaging of endothelial Erk activity reveals dynamic and sequential signalling events during regenerative angiogenesis

Insights into the non-mitotic functions of Aurora kinase A: more than just cell division

Intracellular signaling dynamics and their role in coordinating tissue repair

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