ArticleJournal of visualized experiments : JoVE2018
Micromanipulation Techniques Allowing Analysis of Morphogenetic Dynamics and Turnover of Cytoskeletal Regulators.
Article in Journal of visualized experiments : JoVE, 2018. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 5 papers.
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The abstract states no effect estimate the extractor could read, or names no intervention and outcome on the map, so this paper lights no cell and moves no belief. It is still indexed, cited and linked below.
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Who cites it
5 citing papers in PubMed, 10 citations in OpenAlex.
- Profilin promotes lamellipodium protrusion by tuning the antagonistic activities of capping protein and VASP.Nature communications · 2026Article
- Lamellipodia-like actin networks in cells lacking WAVE regulatory complex.Journal of cell science · 2022Article
- Induced Arp2/3 Complex Depletion Increases FMNL2/3 Formin Expression and Filopodia Formation.Frontiers in cell and developmental biology · 2021Article
- Loss of Ena/VASP interferes with lamellipodium architecture, motility and integrin-dependent adhesion.eLife · 2020Article
- Lamellipodin tunes cell migration by stabilizing protrusions and promoting adhesion formation.Journal of cell science · 2020Article
Corrections and comments
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Authors and funding
2 authors at 1 institution in 2 countries.
Funding
No grant is acknowledged in the PubMed record.
Abstract
Examining the spatiotemporal dynamics of proteins can reveal their functional importance in various contexts. In this article, it is discussed how fluorescent recovery after photobleaching (FRAP) and photoactivation techniques can be used to study the spatiotemporal dynamics of proteins in subcellular locations. We also show how these techniques enable straightforward determination of various parameters linked to actin cytoskeletal regulation and cell motility. Moreover, the microinjection of cells is additionally described as an alternative treatment (potentially preceding or complementing the aforementioned photomanipulation techniques) to trigger instantaneous effects of translocated proteins on cell morphology and function. Micromanipulation such as protein injection or local application of plasma membrane-permeable drugs or cytoskeletal inhibitors can serve as powerful tool to record immediate consequences of a given treatment on cell behavior at the single cell and subcellular level. This is exemplified here by immediate induction of lamellipodial cell edge protrusion by the injection of recombinant Rac1 protein, as established a quarter-century ago. In addition, we provide a protocol for determining the turnover of enhanced green fluorescent protein (EGFP)-VASP, an actin filament polymerase prominently accumulating at lamellipodial tips of B16-F1 cells, employing FRAP and including associated data analysis and curve fitting. We also present guidelines for estimating the rates of lamellipodial actin network polymerization, as exemplified by cells expressing EGFP-tagged β-actin. Finally, instructions are given for how to investigate the rates of actin monomer mobility within the cell cytoplasm, followed by actin incorporation at sites of rapid filament assembly, such as the tips of protruding lamellipodia, using photoactivation approaches. None of these protocols is restricted to components or regulators of the actin cytoskeleton, but can easily be extended to explore in analogous fashion the spatiotemporal dynamics and function of proteins in various different subcellular structures or functional contexts.
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Registered trials
Read under generation 80e0d062 · epoch 390. Bibliography from PubMed, PubMed Central and OpenAlex; grants from NIH RePORTER; trial links from ClinicalTrials.gov; estimates, votes and beliefs from the OpenQuestion graph.