Focal adhesions contain three specialized actin nanoscale layers.
Journal
Nature communications
ISSN: 2041-1723
Titre abrégé: Nat Commun
Pays: England
ID NLM: 101528555
Informations de publication
Date de publication:
21 Mar 2024
21 Mar 2024
Historique:
received:
25
01
2024
accepted:
13
03
2024
medline:
22
3
2024
pubmed:
22
3
2024
entrez:
22
3
2024
Statut:
epublish
Résumé
Focal adhesions (FAs) connect inner workings of cell to the extracellular matrix to control cell adhesion, migration and mechanosensing. Previous studies demonstrated that FAs contain three vertical layers, which connect extracellular matrix to the cytoskeleton. By using super-resolution iPALM microscopy, we identify two additional nanoscale layers within FAs, specified by actin filaments bound to tropomyosin isoforms Tpm1.6 and Tpm3.2. The Tpm1.6-actin filaments, beneath the previously identified α-actinin cross-linked actin filaments, appear critical for adhesion maturation and controlled cell motility, whereas the adjacent Tpm3.2-actin filament layer beneath seems to facilitate adhesion disassembly. Mechanistically, Tpm3.2 stabilizes ACF-7/MACF1 and KANK-family proteins at adhesions, and hence targets microtubule plus-ends to FAs to catalyse their disassembly. Tpm3.2 depletion leads to disorganized microtubule network, abnormally stable FAs, and defects in tail retraction during migration. Thus, FAs are composed of distinct actin filament layers, and each may have specific roles in coupling adhesions to the cytoskeleton, or in controlling adhesion dynamics.
Identifiants
pubmed: 38514695
doi: 10.1038/s41467-024-46868-7
pii: 10.1038/s41467-024-46868-7
doi:
Types de publication
Journal Article
Langues
eng
Sous-ensembles de citation
IM
Pagination
2547Subventions
Organisme : Academy of Finland (Suomen Akatemia)
ID : 346133
Informations de copyright
© 2024. The Author(s).
Références
Revach, O. Y., Grosheva, I. & Geiger, B. Biomechanical regulation of focal adhesion and invadopodia formation. J. Cell Sci. 133, jcs244848 (2020).
pubmed: 33093229
doi: 10.1242/jcs.244848
Kanchanawong, P. & Calderwood, D. A. Organization, dynamics and mechanoregulation of integrin-mediated cell–ECM adhesions. Nat. Rev. Mol. Cell Biol. 24, 142–161 (2022).
pubmed: 36168065
pmcid: 9892292
doi: 10.1038/s41580-022-00531-5
Kanchanawong, P. et al. Nanoscale architecture of integrin-based cell adhesions. Nature 468, 580–584 (2010).
pubmed: 21107430
pmcid: 3046339
doi: 10.1038/nature09621
Liu, J. et al. Talin determines the nanoscale architecture of focal adhesions. Proc. Natl Acad. Sci. USA. 112, E4864–E4873 (2015).
pubmed: 26283369
pmcid: 4568271
doi: 10.1073/pnas.1512025112
Case, L. B. & Waterman, C. M. Integration of actin dynamics and cell adhesion by a three-dimensional, mechanosensitive molecular clutch. Nat. Cell Biol. 17, 955–963 (2015).
pubmed: 26121555
pmcid: 6300998
doi: 10.1038/ncb3191
Riveline, D. et al. Focal contacts as mechanosensors: Externally applied local mechanical force induces growth of focal contacts by an mDia1-dependent and ROCK-independent mechanism. J. Cell Biol. 153, 1175–1185 (2001).
pubmed: 11402062
pmcid: 2192034
doi: 10.1083/jcb.153.6.1175
Zaidel-Bar, R., Ballestrem, C., Kam, Z. & Geiger, B. Early molecular events in the assembly of matrix adhesions at the leading edge of migrating cells. J. Cell Sci. 116, 4605–4613 (2003).
pubmed: 14576354
doi: 10.1242/jcs.00792
Tojkander, S., Gateva, G., Husain, A., Krishnan, R. & Lappalainen, P. Generation of contractile actomyosin bundles depends on mechanosensitive actin filament assembly and disassembly. Elife 4, 1–28 (2015).
doi: 10.7554/eLife.06126
Gardel, M. L., Schneider, I. C., Aratyn-Schaus, Y. & Waterman, C. M. Mechanical integration of actin and adhesion dynamics in cell migration. Annu. Rev. Cell Dev. Biol. 26, 315–333 (2010).
pubmed: 19575647
pmcid: 4437624
doi: 10.1146/annurev.cellbio.011209.122036
Moreno-Layseca, P., Icha, J., Hamidi, H. & Ivaska, J. Integrin trafficking in cells and tissues. Nat. Cell Biol. 21, 122–132 (2019).
pubmed: 30602723
pmcid: 6597357
doi: 10.1038/s41556-018-0223-z
Moreno-Layseca, P. et al. Cargo-specific recruitment in clathrin- and dynamin-independent endocytosis. Nat. Cell Biol. 23, 1073–1084 (2021).
pubmed: 34616024
doi: 10.1038/s41556-021-00767-x
Kaverina, I., Rottner, K. & Small, J. V. Targeting, capture, and stabilization of microtubules at early focal adhesions. J. Cell Biol. 142, 181–190 (1998).
pubmed: 9660872
pmcid: 2133026
doi: 10.1083/jcb.142.1.181
Kaverina, I., Krylyshkina, O. & Small, J. V. Microtubule targeting of substrate contacts promotes their relaxation and dissociation. J. Cell Biol. 146, 1033–1043 (1999).
pubmed: 10477757
pmcid: 2169483
doi: 10.1083/jcb.146.5.1033
Ballestrem, C., Wehrle-Haller, B., Hinz, B. & Imhof, B. A. Actin-dependent lamellipodia formation and microtubule-dependent tail retraction control-directed cell migration. Mol. Biol. Cell 11, 2999–3012 (2000).
pubmed: 10982396
pmcid: 14971
doi: 10.1091/mbc.11.9.2999
Ezratty, E. J., Partridge, M. A. & Gundersen, G. G. Microtubule-induced focal adhesion disassembly is mediated by dynamin and focal adhesion kinase. Nat. Cell Biol. 7, 581–590 (2005).
pubmed: 15895076
doi: 10.1038/ncb1262
Sun, Z. et al. Kank2 activates talin, reduces force transduction across integrins and induces central adhesion formation. Nat. Cell Biol. 18, 941–953 (2016).
pubmed: 27548916
pmcid: 6053543
doi: 10.1038/ncb3402
Bouchet, B. P. et al. Talin-KANK1 interaction controls the recruitment of cortical microtubule stabilizing complexes to focal adhesions. Elife 5, 1–23 (2016).
doi: 10.7554/eLife.18124
Wu, X., Kodama, A. & Fuchs, E. ACF7 Regulates Cytoskeletal-Focal Adhesion Dynamics and Migration and Has ATPase Activity. Cell 135, 137–148 (2008).
pubmed: 18854161
pmcid: 2703712
doi: 10.1016/j.cell.2008.07.045
Voelzmann, A. et al. Drosophila Short stop as a paradigm for the role and regulation of spectraplakins. Semin. Cell Dev. Biol. 69, 40–57 (2017).
pubmed: 28579450
doi: 10.1016/j.semcdb.2017.05.019
Rafiq, N. B. M. et al. A mechano-signalling network linking microtubules, myosin IIA filaments and integrin-based adhesions. Nat. Mater. 18, 638–649 (2019).
pubmed: 31114072
doi: 10.1038/s41563-019-0371-y
Kenific, C. M. et al. NBR 1 enables autophagy-dependent focal adhesion turnover. J. Cell Biol. 212, 577–590 (2016).
pubmed: 26903539
pmcid: 4772495
doi: 10.1083/jcb.201503075
Sharifi, M. N. et al. Autophagy Promotes Focal Adhesion Disassembly and Cell Motility of Metastatic Tumor Cells through the Direct Interaction of Paxillin with LC3. Cell Rep. 15, 1660–1672 (2016).
pubmed: 27184837
pmcid: 4880529
doi: 10.1016/j.celrep.2016.04.065
Juanes, M. A. et al. The role of APC-mediated actin assembly in microtubule capture and focal adhesion turnover. J. Cell Biol. 218, 3415–3435 (2019).
pubmed: 31471457
pmcid: 6781439
doi: 10.1083/jcb.201904165
Zaidel-Bar, R., Itzkovitz, S., Ma, A., Iyengar, R. & Geiger, B. Nat. Cell Biol. 9, 858–867 (2007).
Kuo, J. C. et al. Analysis of the myosin-II-responsive focal adhesion proteome reveals a role for β-Pix in negative regulation of focal adhesion maturation. Nat. Cell Biol. 13, 383–395 (2011).
pubmed: 21423176
pmcid: 3279191
doi: 10.1038/ncb2216
Schiller, H. B. & Fässler, R. Mechanosensitivity and compositional dynamics of cell-matrix adhesions. EMBO Rep. 14, 509–519 (2013).
pubmed: 23681438
pmcid: 3674437
doi: 10.1038/embor.2013.49
Horton, E. R. et al. Definition of a consensus integrin adhesome and its dynamics during adhesion complex assembly and disassembly. Nat. Cell Biol. 17, 1577–1587 (2015).
pubmed: 26479319
pmcid: 4663675
doi: 10.1038/ncb3257
Chastney, M. R. et al. Topological features of integrin adhesion complexes revealed by multiplexed proximity biotinylation. J. Cell Biol. 219, e202003038 (2020).
pubmed: 32585685
pmcid: 7401799
doi: 10.1083/jcb.202003038
Shtengel, G. et al. Interferometric fluorescent super-resolution microscopy resolves 3D cellular ultrastructure. Proc. Natl Acad. Sci. USA. 106, 3125–3130 (2009).
pubmed: 19202073
pmcid: 2637278
doi: 10.1073/pnas.0813131106
Stubb, A. et al. Superresolution architecture of cornerstone focal adhesions in human pluripotent stem cells. Nat. Commun. 10, 4756 (2019).
pubmed: 31628312
pmcid: 6802214
doi: 10.1038/s41467-019-12611-w
Ishizaki, T. et al. Coordination of microtubules and the actin cytoskeleton by the Rho effector mDia1. Nat. Cell Biol. 3, 8–14 (2001).
pubmed: 11146620
doi: 10.1038/35050598
Iskratsch, T. et al. FHOD1 is needed for directed forces and adhesion maturation during cell spreading and migration. Dev. Cell 27, 545–559 (2013).
pubmed: 24331927
pmcid: 3890431
doi: 10.1016/j.devcel.2013.11.003
Chorev, D. S., Moscovitz, O., Geiger, B. & Sharon, M. Regulation of focal adhesion formation by a vinculin-Arp2/3 hybrid complex. Nat. Commun. 5, 3758 (2014).
pubmed: 24781749
doi: 10.1038/ncomms4758
Michelot, A. & Drubin, D. G. Building distinct actin filament networks in a common cytoplasm. Curr. Biol. 21, R560–R569 (2011).
pubmed: 21783039
pmcid: 3384529
doi: 10.1016/j.cub.2011.06.019
Blanchoin, L., Boujemaa-Paterski, R., Sykes, C. & Plastino, J. Actin Dynamics, Architecture, and Mechanics in Cell Motility. Physiol. Rev. 94, 235–263 (2014).
pubmed: 24382887
doi: 10.1152/physrev.00018.2013
Fletcher, D. A. & Mullins, R. D. Cell mechanics and the cytoskeleton. Nature 463, 485–492 (2010).
pubmed: 20110992
pmcid: 2851742
doi: 10.1038/nature08908
Blanchoin, L., Letort, G., Ennomani, H., Gressin, L. & Théry, M. Dynamic reorganization of the actin cytoskeleton. F1000Research 4, 1–11 (2015).
Boiero Sanders, M., Antkowiak, A. & Michelot, A. Diversity from similarity: cellular strategies for assigning particular identities to actin filaments and networks: Actin filaments and networks identities. Open Biol. 10, 200157 (2020).
pubmed: 32873155
pmcid: 7536088
doi: 10.1098/rsob.200157
Gunning, P., O’Neill, G. & Hardeman, E. Tropomyosin-based regulation of the actin cytoskeleton in time and space. Physiol. Rev. 88, 1–35 (2008).
pubmed: 18195081
doi: 10.1152/physrev.00001.2007
Gunning, P. W., Hardeman, E. C., Lappalainen, P. & Mulvihill, D. P. Tropomyosin - master regulator of actin filament function in the cytoskeleton. J. Cell Sci. 128, 2965–2974 (2015).
pubmed: 26240174
Von Der Ecken, J. et al. Structure of the F-actin-tropomyosin complex. Nature 519, 114–117 (2015).
pubmed: 25470062
doi: 10.1038/nature14033
Kumari, R. et al. Tropomodulins Control the Balance between Protrusive and Contractile Structures by Stabilizing Actin-Tropomyosin Filaments. Curr. Biol. 30, 767–778.e5 (2020).
pubmed: 32037094
pmcid: 7065974
doi: 10.1016/j.cub.2019.12.049
Gunning, P. W., Ghoshdastider, U., Whitaker, S., Popp, D. & Robinson, R. C. The evolution of compositionally and functionally distinct actin filaments. J. Cell Sci. 128, 2009–2019 (2015).
pubmed: 25788699
doi: 10.1242/jcs.165563
Gateva, G. et al. Tropomyosin Isoforms Specify Functionally Distinct Actin Filament Populations In Vitro. Curr. Biol. 27, 705–713 (2017).
pubmed: 28216317
pmcid: 5344678
doi: 10.1016/j.cub.2017.01.018
Jansen, S. & Goode, B. L. Tropomyosin isoforms differentially tune actin filament length and disassembly. Mol. Biol. Cell 30, 671–679 (2019).
pubmed: 30650006
pmcid: 6589703
doi: 10.1091/mbc.E18-12-0815
Reindl, T. et al. Distinct actin–tropomyosin cofilament populations drive the functional diversification of cytoskeletal myosin motor complexes. iScience 25, 104484 (2022).
pubmed: 35720262
pmcid: 9204724
doi: 10.1016/j.isci.2022.104484
Tojkander, S. et al. A molecular pathway for myosin II recruitment to stress fibers. Curr. Biol. 21, 539–550 (2011).
pubmed: 21458264
doi: 10.1016/j.cub.2011.03.007
Hu, S. et al. Reciprocal regulation of actomyosin organization and contractility in nonmuscle cells by tropomyosins and alpha-actinins. Mol. Biol. Cell 30, 2025–2036 (2019).
pubmed: 31216217
pmcid: 6727768
doi: 10.1091/mbc.E19-02-0082
Selvaraj, M. et al. Structural basis underlying specific biochemical activities of non-muscle tropomyosin isoforms. Cell Rep. 42, 111900 (2023).
pubmed: 36586407
doi: 10.1016/j.celrep.2022.111900
Schevzov, G., Whittaker, S. P., Fath, T., Lin, J. J. C. & Gunning, P. W. Tropomyosin isoforms and reagents. Bioarchitecture 1, 135–164 (2011).
pubmed: 22069507
pmcid: 3210517
doi: 10.4161/bioa.1.4.17897
Hotulainen, P. & Lappalainen, P. Stress fibers are generated by two distinct actin assembly mechanisms in motile cells. J. Cell Biol. 173, 383–394 (2006).
pubmed: 16651381
pmcid: 2063839
doi: 10.1083/jcb.200511093
Meiring, J. C. M. et al. Co-polymers of Actin and Tropomyosin Account for a Major Fraction of the Human Actin Cytoskeleton Report Co-polymers of Actin and Tropomyosin Account for a Major Fraction of the Human Actin Cytoskeleton. Curr. Biol. 28, 2331–2337.e5 (2018).
pubmed: 29983319
doi: 10.1016/j.cub.2018.05.053
Lilja, J. et al. SHANK proteins limit integrin activation by directly interacting with Rap1 and R-Ras. Nat. Cell Biol. 19, 292–305 (2017).
pubmed: 28263956
pmcid: 5386136
doi: 10.1038/ncb3487
Shattil, S. J., Kim, C. & Ginsberg, M. H. The final steps of integrin activation: The end game. Nat. Rev. Mol. Cell Biol. 11, 288–300 (2010).
pubmed: 20308986
pmcid: 3929966
doi: 10.1038/nrm2871
Rantala, J. K. et al. SHARPIN is an endogenous inhibitor of β1-integrin activation. Nat. Cell Biol. 13, 1315–1324 (2011).
pubmed: 21947080
pmcid: 3257806
doi: 10.1038/ncb2340
Berginski, M. E., Vitriol, E. A., Hahn, K. M. & Gomez, S. M. High-resolution quantification of focal adhesion spatiotemporal dynamics in living cells. PLoS One 6, e22025 (2011).
pubmed: 21779367
pmcid: 3136503
doi: 10.1371/journal.pone.0022025
Wu, C. et al. Arp2/3 is critical for lamellipodia and response to extracellular matrix cues but is dispensable for chemotaxis. Cell 148, 973–987 (2012).
pubmed: 22385962
pmcid: 3707508
doi: 10.1016/j.cell.2011.12.034
Chen, N. P., Sun, Z. & Fässler, R. The Kank family proteins in adhesion dynamics. Curr. Opin. Cell Biol. 54, 130–136 (2018).
pubmed: 29909279
doi: 10.1016/j.ceb.2018.05.015
Yu, M. et al. Force-Dependent Regulation of Talin-KANK1 Complex at Focal Adhesions. Nano Lett. 19, 5982–5990 (2019).
pubmed: 31389241
doi: 10.1021/acs.nanolett.9b01732
Karakesisoglou, I., Yang, Y. & Fuchs, E. An epidermal plakin that integrates actin and microtubule networks at cellular junctions. J. Cell Biol. 149, 195–208 (2000).
pubmed: 10747097
pmcid: 2175090
doi: 10.1083/jcb.149.1.195
Tsvetkov, A. S., Samsonov, A., Akhmanova, A., Galjart, N. & Popov, S. V. Microtubule-binding proteins CLASP1 and CLASP2 interact with actin filaments. Cell Motil. 64, 519–530 (2007).
doi: 10.1002/cm.20201
Rodgers, N. C. et al. CLASP2 facilitates dynamic actin filament organization along the microtubule lattice. Mol. Biol. Cell 34, br3 (2023).
pubmed: 36598814
pmcid: 10011731
doi: 10.1091/mbc.E22-05-0149
Yue, J. et al. In vivo epidermal migration requires focal adhesion targeting of ACF7. Nat. Commun. 7, 11692 (2016).
pubmed: 27216888
pmcid: 5476826
doi: 10.1038/ncomms11692
Patla, I. et al. Dissecting the molecular architecture of integrin adhesion sites by cryo-electron tomography. Nat. Cell Biol. 12, 909–915 (2010).
pubmed: 20694000
doi: 10.1038/ncb2095
Martins, B. et al. Unveiling the polarity of actin filaments by cryo-electron tomography. Structure 29, 488–498.e4 (2021).
pubmed: 33476550
pmcid: 8111420
doi: 10.1016/j.str.2020.12.014
Legerstee, K., Sueters, J., Kremers, G.-J., Hoogenboom, J. P. & Houtsmuller, A. B. Correlative light and electron microscopy reveals fork-shaped structures at actin entry sites of focal adhesions. bioRxiv 2022.04.27.489664 https://doi.org/10.1242/bio.059417 (2022).
Cagigas, M. L. et al. Correlative cryo-ET identifies actin/tropomyosin filaments that mediate cell–substrate adhesion in cancer cells and mechanosensitivity of cell proliferation. Nat. Mater. 21, 120–128 (2022).
pubmed: 34518666
doi: 10.1038/s41563-021-01087-z
Wang, Y. et al. Drug targeting the actin cytoskeleton potentiates the cytotoxicity of low dose vincristine by abrogating actin-mediated repair of spindle defects. Mol. Cancer Res. 18, 1074–1087 (2020).
pubmed: 32269073
doi: 10.1158/1541-7786.MCR-19-1122
Stehbens, S. & Wittmann, T. Targeting and transport: How microtubules control focal adhesion dynamics. J. Cell Biol. 198, 481–489 (2012).
pubmed: 22908306
pmcid: 3514042
doi: 10.1083/jcb.201206050
Seetharaman, S. et al. Microtubules tune mechanosensitive cell responses. Nat. Mater. 21, 366–377 (2021).
pubmed: 34663953
doi: 10.1038/s41563-021-01108-x
Stehn, J. R. et al. A novel class of anticancer compounds targets the actin cytoskeleton in tumor cells. Cancer Res 73, 5169–5182 (2013).
pubmed: 23946473
doi: 10.1158/0008-5472.CAN-12-4501
Janco, M. et al. Molecular integration of the anti-tropomyosin compound ATM-3507 into the coiled coil overlap region of the cancer-associated Tpm3.1. Sci. Rep. 9, 11262 (2019).
pubmed: 31375704
pmcid: 6677793
doi: 10.1038/s41598-019-47592-9
Hendricks, M. & Weintraub, H. Tropomyosin is decreased in transformed cells. Proc. Natl Acad. Sci. USA. 78, 5633–5637 (1981).
pubmed: 6272310
pmcid: 348810
doi: 10.1073/pnas.78.9.5633
Kokate, S. B. et al. Caldesmon controls stress fiber force-balance through dynamic cross-linking of myosin II and actin-tropomyosin filaments. Nat. Commun. 13, 1–20 (2022).
doi: 10.1038/s41467-022-33688-w
Lehtimäki, J. I., Rajakylä, E. K., Tojkander, S. & Lappalainen, P. Generation of stress fibers through myosin-driven reorganization of the actin cortex. Elife 10, 1–43 (2021).
doi: 10.7554/eLife.60710
Hakala, M. et al. Twinfilin uncaps filament barbed ends to promote turnover of lamellipodial actin networks. Nat. Cell Biol. 23, 147–159 (2021).
pubmed: 33558729
doi: 10.1038/s41556-020-00629-y
Ran, F. A. et al. Genome engineering using the CRISPR-Cas9 system. Nat. Protoc. 8, 2281–2308 (2013).
Jimenez, A. J. et al. Acto-myosin network geometry defines centrosome position. Curr. Biol. 31, 1206–1220.e5 (2021).
pubmed: 33609453
doi: 10.1016/j.cub.2021.01.002
Azioune, A., Storch, M., Bornens, M., Théry, M. & Piel, M. Simple and rapid process for single cell micro-patterning. Lab Chip 9, 1640–1642 (2009).
pubmed: 19458875
doi: 10.1039/b821581m
Kopek, B. G. et al. Diverse protocols for correlative super-resolution fluorescence imaging and electron microscopy of chemically fixed samples. Nat. Protoc. 12, 916–946 (2017).
pubmed: 28384138
pmcid: 5514615
doi: 10.1038/nprot.2017.017
Laine, R. F. et al. NanoJ: A high-performance open-source super-resolution microscopy toolbox. J. Phys. D. Appl. Phys. 52, 163001 (2019).
pubmed: 33191949
pmcid: 7655149
doi: 10.1088/1361-6463/ab0261
Han, S. J., Oak, Y., Groisman, A. & Danuser, G. Traction microscopy to identify force modulation in subresolution adhesions. Nat. Methods 12, 653–656 (2015).
pubmed: 26030446
pmcid: 4490115
doi: 10.1038/nmeth.3430
Jiu, Y. et al. Vimentin intermediate filaments control actin stress fiber assembly through GEF-H1 and RhoA. J. Cell Sci. 130, 892–902 (2017). 892–902.
pubmed: 28096473
pmcid: 5358333
Berginski, M. E. & Gomez, S. M. The Focal Adhesion Analysis Server: a web tool for analyzing focal adhesion dynamics. F1000Res. 2, 68 (2013).
pubmed: 24358855
pmcid: 3752736
doi: 10.12688/f1000research.2-68.v1
Bancaud, A., Huet, S., Rabut, G. & Ellenberg, J. Fluorescence perturbation techniques to study mobility and molecular dynamics of proteins in live cells: FRAP, Photoactivation, Photoconversion, and FLIP. Cold Spring Harb. Protoc. 5, 1303–1325 (2010).