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Control of Actin Polymerization in Cells Through Activation and Inhibition of the Arp2/3 Complex

Control of Actin Polymerization in Cells Through Activation and Inhibition of the Arp2/3 Complex

Roberto Domínguez

Department of Physiology. University of Pennsylvania. Perelman School of Medicine (Philadelphia, USA)

24/06/2021 to 01/01/0001
Time: 16:30
on-line
Streaming link: OPEN
Host: Miguel Vicente Manzanares
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Mechanisms of Arp2/3 complex activation and inhibition

Arp2/3 complex: Actin filament nucleators catalyze the rate-limiting step in actin polymerization, i.e. nucleation [1, 2]. The Arp2/3 complex is the only nucleator that can generate branched actin networks. A family of Arp2/3 complex activators, collectively known as nucleation promoting factors (NPFs), recruit the Arp2/3 complex to specific cellular membranes. There, the complex generates so-called dendritic actin networks, i.e. networks of branched actin filaments. These networks exert pushing forces on membranes during processes such as cell motility, vesicular trafficking and membrane scission. The Arp2/3 complex consists of seven proteins, including two actin related proteins (Arp2 and Arp3) that act as a pseudo-actin dimer during nucleation, and five scaffolding subunits (ArpC1-ArpC5) that hold the Arps in place [4] and mediate interactions with the mother actin filament in the branch [5].

 

Nucleation promoting factors (NPFs) and Arp2/3 complex activation: Isolated mammalian Arp2/3 complex is inactive, with the Arps splayed apart [4]. In the presence of actin monomers and filaments, NPFs perform three main functions: a) they promote a conformational change that repositions the Arps into a filament-like (short-pitch) conformation [5-9], b) they recruit actin subunits to the barbed end of the Arps to form the initial polymerization seed [10, 11] and c) they promote binding of the complex to the side of a pre-existing (mother) filament to form a branch (daughter) filament that emerges at a ~70° angle [5, 12, 13]. The NPFs are generally unrelated, except for their C-terminal WCA region, comprising one to three WASP-homology 2 (WH2) domains (W) that bind actin, and Central (C) and Acidic (A) segments that bind Arp2/3 complex [14-18]. The WCA region of NPFs is necessary and sufficient to activate Arp2/3 complex in vitro, whereas their N-terminal domains typically diverge and are involved in regulation/localization, allowing for the spatiotemporal control of Arp2/3 complex activity in cells [3, 19]. In response to signaling, NPFs trigger a burst of branched actin polymerization

 

Arpin and Arp2/3 complex inhibition: The activity of Arp2/3 complex is additionally regulated in time and space by Arp2/3 complex inhibitors. Arpin, a major inhibitor of Arp2/3 complex, controls the persistence of lamellipodial protrusions and cell migration [20].

 

New developments discussed in my lecture: Despite intensive interest, the mechanisms that control Arp2/3 complex activation by NPFs and inhibition by Arpin remained poorly understood. Several cryo-electron microscopy (cryo-EM) structures of Arp2/3 complex, including structure of Arp2/3 complex in the branch and with bound NPFs or Arpin, have been recently obtained, and substantially help to illuminate the mechanisms that control Arp2/3 complex activity. These structures reveal the conformational changes that the complex undergoes during activation, as well as the binding sites of NPFs and Arpin on the complex. Strikingly, despite their opposite functions, NPFs and Arpin use similar structural mechanisms. During my lecture, I will describe these developments and their implications for our understanding of Arp2/3 complex function in cells.

 

References cited

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  2. Dominguez, R. (2016) The WH2 Domain and Actin Nucleation: Necessary but Insufficient. Trends Biochem Sci 41 (6), 478-90.
  3. Molinie, N. and Gautreau, A. (2018) The Arp2/3 Regulatory System and Its Deregulation in Cancer. Physiol Rev 98 (1), 215-238.
  4. Robinson, R.C. et al. (2001) Crystal structure of Arp2/3 complex. Science 294 (5547), 1679-84.
  5. Rouiller, I. et al. (2008) The structural basis of actin filament branching by the Arp2/3 complex. The Journal of cell biology 180 (5), 887-95.
  6. Goley, E.D. et al. (2004) Critical conformational changes in the Arp2/3 complex are induced by nucleotide and nucleation promoting factor. Mol Cell 16 (2), 269-79.
  7. Rodal, A.A. et al. (2005) Conformational changes in the Arp2/3 complex leading to actin nucleation. Nat Struct Mol Biol 12 (1), 26-31.
  8. Rodnick-Smith, M. et al. (2016) Role and structural mechanism of WASP-triggered conformational changes in branched actin filament nucleation by Arp2/3 complex. Proc Natl Acad Sci U S A 113 (27), E3834-43.
  9. Espinoza-Sanchez, S. et al. (2018) Conformational changes in Arp2/3 complex induced by ATP, WASp-VCA, and actin filaments. Proc Natl Acad Sci U S A 115 (37), E8642-E8651.
  10. Boczkowska, M. et al. (2008) X-ray scattering study of activated Arp2/3 complex with bound actin-WCA. Structure 16 (5), 695-704.
  11. Padrick, S.B. et al. (2011) Arp2/3 complex is bound and activated by two WASP proteins. Proceedings of the National Academy of Sciences of the United States of America 108 (33), E472-9.
  12. Smith, B.A. et al. (2013) Pathway of actin filament branch formation by Arp2/3 complex revealed by single-molecule imaging. Proceedings of the National Academy of Sciences of the United States of America 110 (4), 1285-90.
  13. Smith, B.A. et al. (2013) Three-color single molecule imaging shows WASP detachment from Arp2/3 complex triggers actin filament branch formation. Elife 2, e01008.
  14. Marchand, J.B. et al. (2001) Interaction of WASP/Scar proteins with actin and vertebrate Arp2/3 complex. Nat Cell Biol 3 (1), 76-82.
  15. Miki, H. and Takenawa, T. (1998) Direct binding of the verprolin-homology domain in N-WASP to actin is essential for cytoskeletal reorganization. Biochem Biophys Res Commun 243 (1), 73-8.
  16. Panchal, S.C. et al. (2003) A conserved amphipathic helix in WASP/Scar proteins is essential for activation of Arp2/3 complex. Nat Struct Biol 10 (8), 591-8.
  17. Chereau, D. et al. (2005) Actin-bound structures of Wiskott-Aldrich syndrome protein (WASP)-homology domain 2 and the implications for filament assembly. Proceedings of the National Academy of Sciences of the United States of America 102 (46), 16644-9.
  18. Higgs, H.N. et al. (1999) Influence of the C terminus of Wiskott-Aldrich syndrome protein (WASp) and the Arp2/3 complex on actin polymerization. Biochemistry 38 (46), 15212-22.
  19. Kast, D.J. et al. (2015) WHAMM Directs the Arp2/3 Complex to the ER for Autophagosome Biogenesis through an Actin Comet Tail Mechanism. Curr Biol 25 (13), 1791-7.
  20. Dang, I. et al. (2013) Inhibitory signalling to the Arp2/3 complex steers cell migration. Nature 503 (7475), 281-4.
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