Live‐cell super‐resolution imaging of actin using LifeAct‐14 with a PAINT‐based approach

Haresh Bhaskar; Dirk‐Jan Kleinjan; Curran Oi; Zoe Gidden; Susan J. Rosser; Mathew H. Horrocks; Lynne Regan

doi:10.1002/pro.4558

Live‐cell super‐resolution imaging of actin using LifeAct‐14 with a PAINT‐based approach

Haresh Bhaskar, Dirk‐Jan Kleinjan, Curran Oi, Zoe Gidden, Susan J. Rosser, Mathew H. Horrocks, Lynne Regan

Research output: Contribution to journal › Article › peer-review

Abstract / Description of output

We present direct-LIVE-PAINT, an easy-to-implement approach for the nanoscopic imaging of protein structures in live cells using labeled binding peptides. We demonstrate the feasibility of direct-LIVE-PAINT with an actin-binding peptide fused to EGFP, the location of which can be accurately determined as it transiently binds to actin filaments. We show that direct-LIVE-PAINT can be used to image actin structures below the diffraction-limit of light and have used it to observe the dynamic nature of actin in live cells. We envisage a similar approach could be applied to imaging other proteins within live mammalian cells.

Original language	English
Article number	e4558
Number of pages	8
Journal	Protein Science
Volume	32
Issue number	2
Early online date	31 Dec 2022
DOIs	https://doi.org/10.1002/pro.4558
Publication status	Published - 1 Feb 2023

Keywords / Materials (for Non-textual outputs)

super-resolution (SR) imaging
single-molecule localization microscopy (SMLM)
actin
live-cell imaging
LifeAct

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10.1002/pro.4558Licence: Creative Commons: Attribution (CC-BY)

ReganEtalPS2022LiveCellSuperResolutionImagingOfActinFinal published version, 1.79 MBLicence: Creative Commons: Attribution (CC-BY)

Cite this

@article{f059a571e2284b07a2dc1f206138dfdc,

title = "Live‐cell super‐resolution imaging of actin using LifeAct‐14 with a PAINT‐based approach",

abstract = "We present direct-LIVE-PAINT, an easy-to-implement approach for the nanoscopic imaging of protein structures in live cells using labeled binding peptides. We demonstrate the feasibility of direct-LIVE-PAINT with an actin-binding peptide fused to EGFP, the location of which can be accurately determined as it transiently binds to actin filaments. We show that direct-LIVE-PAINT can be used to image actin structures below the diffraction-limit of light and have used it to observe the dynamic nature of actin in live cells. We envisage a similar approach could be applied to imaging other proteins within live mammalian cells.",

keywords = "super-resolution (SR) imaging, single-molecule localization microscopy (SMLM), actin, live-cell imaging, LifeAct",

author = "Haresh Bhaskar and Dirk‐Jan Kleinjan and Curran Oi and Zoe Gidden and Rosser, {Susan J.} and Horrocks, {Mathew H.} and Lynne Regan",

note = "Funding Information: Haresh Bhaskar acknowledges the support of Wellcome Trust via the Integrative Cell Mechanisms PhD program (grant number 226437/Z/22/Z). The super‐resolution instrument was funded by UCB Biopharma and a kind donation from Dr. Jim Love. We offer special thanks to members of the Rosser Lab for their tissue culture expertise, Dr. Rebecca Saleeb for help with the SMLM analysis workflow, and Dr. Raef Shams for help with protein structure visualization. Publisher Copyright: {\textcopyright} 2022 The Authors. Protein Science published by Wiley Periodicals LLC on behalf of The Protein Society.",

year = "2023",

month = feb,

day = "1",

doi = "10.1002/pro.4558",

language = "English",

volume = "32",

journal = "Protein Science",

issn = "0961-8368",

publisher = "Cold Spring Harbor Laboratory Press",

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T1 - Live‐cell super‐resolution imaging of actin using LifeAct‐14 with a PAINT‐based approach

AU - Bhaskar, Haresh

AU - Kleinjan, Dirk‐Jan

AU - Oi, Curran

AU - Gidden, Zoe

AU - Rosser, Susan J.

AU - Horrocks, Mathew H.

AU - Regan, Lynne

N1 - Funding Information: Haresh Bhaskar acknowledges the support of Wellcome Trust via the Integrative Cell Mechanisms PhD program (grant number 226437/Z/22/Z). The super‐resolution instrument was funded by UCB Biopharma and a kind donation from Dr. Jim Love. We offer special thanks to members of the Rosser Lab for their tissue culture expertise, Dr. Rebecca Saleeb for help with the SMLM analysis workflow, and Dr. Raef Shams for help with protein structure visualization. Publisher Copyright: © 2022 The Authors. Protein Science published by Wiley Periodicals LLC on behalf of The Protein Society.

PY - 2023/2/1

Y1 - 2023/2/1

N2 - We present direct-LIVE-PAINT, an easy-to-implement approach for the nanoscopic imaging of protein structures in live cells using labeled binding peptides. We demonstrate the feasibility of direct-LIVE-PAINT with an actin-binding peptide fused to EGFP, the location of which can be accurately determined as it transiently binds to actin filaments. We show that direct-LIVE-PAINT can be used to image actin structures below the diffraction-limit of light and have used it to observe the dynamic nature of actin in live cells. We envisage a similar approach could be applied to imaging other proteins within live mammalian cells.

AB - We present direct-LIVE-PAINT, an easy-to-implement approach for the nanoscopic imaging of protein structures in live cells using labeled binding peptides. We demonstrate the feasibility of direct-LIVE-PAINT with an actin-binding peptide fused to EGFP, the location of which can be accurately determined as it transiently binds to actin filaments. We show that direct-LIVE-PAINT can be used to image actin structures below the diffraction-limit of light and have used it to observe the dynamic nature of actin in live cells. We envisage a similar approach could be applied to imaging other proteins within live mammalian cells.

KW - super-resolution (SR) imaging

KW - single-molecule localization microscopy (SMLM)

KW - actin

KW - live-cell imaging

KW - LifeAct

U2 - 10.1002/pro.4558

DO - 10.1002/pro.4558

M3 - Article

SN - 0961-8368

VL - 32

JO - Protein Science

JF - Protein Science

IS - 2

M1 - e4558

ER -

Live‐cell super‐resolution imaging of actin using LifeAct‐14 with a PAINT‐based approach

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Keywords / Materials (for Non-textual outputs)

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