Abstract
Focal adhesions (FAs) are contact points of the cell with the extracellular matrix (ECM) and play a major role in several cellular functions including migration, proliferation, differentiation, and growth. During cell migration, FAs are continuously assembled and disassembled. It is well established that FA dynamics are regulated by the cytoskeleton, motor proteins, small GTPases, and specific kinases and phosphatases. However, more recently, the establishment of contacts between FAs and the endoplasmic reticulum (ER) has been shown to be another factor implicated in the regulation of FA dynamics. The transport of ER tubules along microtubules to contact FAs is indeed crucial to support FA growth. Alteration of such ER-FA contacts affects FA growth, dynamics, and thus cell migration. Here, we present a protocol for live-cell imaging and analysis of ER-FA contact points during cell migration. Our analysis pipeline includes two examples showing physiological conditions and disruption of ER-FA contacts upon nocodazole treatment. The described method can be adapted to different cell lines.
This is a preview of subscription content, log in via an institution to check access.
Access this chapter
Tax calculation will be finalised at checkout
Purchases are for personal use only
Similar content being viewed by others
References
Janiszewska M, Primi MC, Izard T (2020) Cell adhesion in cancer: beyond the migration of single cells. J Biol Chem 295(8):2495–2505. https://doi.org/10.1074/jbc.REV119.007759
Worbs T, Hammerschmidt SI, Forster R (2017) Dendritic cell migration in health and disease. Nat Rev Immunol 17(1):30–48. https://doi.org/10.1038/nri.2016.116
Maziveyi M, Alahari SK (2017) Cell matrix adhesions in cancer: the proteins that form the glue. Oncotarget 8(29):48471–48487. https://doi.org/10.18632/oncotarget.17265
Kechagia JZ, Ivaska J, Roca-Cusachs P (2019) Integrins as biomechanical sensors of the microenvironment. Nat Rev Mol Cell Biol 20(8):457–473. https://doi.org/10.1038/s41580-019-0134-2
Zhang X, Tee YH, Heng JK, Zhu Y, Hu X, Margadant F, Ballestrem C, Bershadsky A, Griffiths G, Yu H (2010) Kinectin-mediated endoplasmic reticulum dynamics supports focal adhesion growth in the cellular lamella. J Cell Sci 123(Pt 22):3901–3912. https://doi.org/10.1242/jcs.069153
Guadagno NA, Margiotta A, Bjørnestad SA, Haugen LH, Kjos I, Xu X, Hu X, Bakke O, Margadant F, Progida C (2020) Rab18 regulates focal adhesion dynamics by interacting with kinectin-1 at the endoplasmic reticulum. J Cell Biol 219(7):e201809020. https://doi.org/10.1083/jcb.201809020
Friedman JR, Webster BM, Mastronarde DN, Verhey KJ, Voeltz GK (2010) ER sliding dynamics and ER-mitochondrial contacts occur on acetylated microtubules. J Cell Biol 190(3):363–375. https://doi.org/10.1083/jcb.200911024
Kerselidou D, Dohai BS, Nelson DR, Daakour S, De Cock N, Hassoun ZAO, Kim DK, Olivet J, El Assal DC, Jaiswal A, Alzahmi A, Saha D, Pain C, Matthijssens F, Lemaitre P, Herfs M, Chapuis J, Ghesquiere B, Vertommen D, Kriechbaumer V, Knoops K, Lopez-Iglesias C, van Zandvoort M, Lambert JC, Hanson J, Desmet C, Thiry M, Lauersen KJ, Vidal M, Van Vlierberghe P, Dequiedt F, Salehi-Ashtiani K, Twizere JC (2021) Alternative glycosylation controls endoplasmic reticulum dynamics and tubular extension in mammalian cells. Sci Adv 7(19). https://doi.org/10.1126/sciadv.abe8349
Garcia-Pardo ME, Simpson JC, O’Sullivan NC (2021) A novel automated image analysis pipeline for quantifying morphological changes to the endoplasmic reticulum in cultured human cells. BMC Bioinform 22(1):427. https://doi.org/10.1186/s12859-021-04334-x
Fokkelman M, Balcioglu HE, Klip JE, Yan K, Verbeek FJ, Danen EH, van de Water B (2016) Cellular adhesome screen identifies critical modulators of focal adhesion dynamics, cellular traction forces and cell migration behaviour. Sci Rep 6:31707. https://doi.org/10.1038/srep31707
Acknowledgments
The work in the authors’ laboratory is supported by grants from the Norwegian Cancer Society (grants 198094 and 223181), the Research Council of Norway (grant 287560), the Anders Jahre Fond til Vitenskapens Fremme, the S. G. Sønneland Foundation, the Nansenfondet og de dermed forbundne fond, and UNIFOR-FRIMED.
Author information
Authors and Affiliations
Corresponding author
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 2023 The Author(s), under exclusive license to Springer Science+Business Media, LLC, part of Springer Nature
About this protocol
Cite this protocol
Guadagno, N.A., Progida, C. (2023). Probing the ER-Focal Adhesion Link During Cell Migration. In: Margadant, C. (eds) Cell Migration in Three Dimensions. Methods in Molecular Biology, vol 2608. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-2887-4_3
Download citation
DOI: https://doi.org/10.1007/978-1-0716-2887-4_3
Published:
Publisher Name: Humana, New York, NY
Print ISBN: 978-1-0716-2886-7
Online ISBN: 978-1-0716-2887-4
eBook Packages: Springer Protocols