TetraThymosinβ Is Required for Actin Dynamics in<i>Caenorhabditis elegans</i>and Acts via Functionally Different Actin-binding Repeats

Troys, Marleen Van; Ono, Kanako; Dewitte, Daisy; Jonckheere, Veronique; Ruyck, Natalie De; Vandekerckhove, Joël; Ono, Shoichiro; Ampè, Christophe

doi:10.1091/mbc.e04-03-0225

Cited by 34 publications

(36 citation statements)

References 54 publications

(77 reference statements)

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“…Therefore, a previous report (9) that the second and third domains of ciboulot do not bind actin may reflect an incorrect definition of domain boundaries. In agreement with this view, all four domains of tetraT␤, a ciboulot-related protein in Caenorhabditis elegans, bind actin with affinities similar to those found here for ciboulot (22).…”

Section: Resultssupporting

confidence: 87%

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Actin-bound structures of Wiskott–Aldrich syndrome protein (WASP)-homology domain 2 and the implications for filament assembly

Chéreau

Kerff

Graceffa

et al. 2005

Proc. Natl. Acad. Sci. U.S.A.

236

375

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Wiskott-Aldrich syndrome protein (WASP)-homology domain 2 (WH2) is a small and widespread actin-binding motif. In the WASP family, WH2 plays a role in filament nucleation by Arp2͞3 complex. Here we describe the crystal structures of complexes of actin with the WH2 domains of WASP, WASP-family verprolin homologous protein, and WASP-interacting protein. Despite low sequence identity, WH2 shares structural similarity with the N-terminal portion of the actin monomer-sequestering thymosin ␤ domain (T␤). We show that both domains inhibit nucleotide exchange by targeting the cleft between actin subdomains 1 and 3, a common binding site for many unrelated actin-binding proteins. Importantly, WH2 is significantly shorter than T␤ but binds actin with Ϸ10-fold higher affinity. WH2 lacks a C-terminal extension that in T␤4 becomes involved in monomer sequestration by interfering with intersubunit contacts in F-actin. Owing to their shorter length, WH2 domains connected in tandem by short linkers can coexist with intersubunit contacts in F-actin and are proposed to function in filament nucleation by lining up actin subunits along a filament strand. The WH2-central region of WASP-family proteins is proposed to function in an analogous way by forming a special class of tandem repeats whose function is to line up actin and Arp2 during Arp2͞3 nucleation. The structures also suggest a mechanism for how profilin-binding Pro-rich sequences positioned N-terminal to WH2 could feed actin monomers directly to WH2, thereby playing a role in filament elongation.x-ray crystallography ͉ isothermal titration calorimetry ͉ nucleotide exchange T he actin cytoskeleton plays an essential role in many cellular functions, including intracellular transport and the control of cell shape and polarity (1). In the cell, a vast number of actin-binding proteins (ABPs) direct the location, rate, and timing for actin assembly into different structures, such as filopodia, lamellipodia, stress fibers, and focal adhesions. ABPs are commonly multidomain proteins, containing signaling domains and structurally conserved actin-binding motifs. One of the most abundant actin-binding motifs is Wiskott-Aldrich syndrome protein (WASP)-homology domain 2 (WH2) (2). The hematopoietic-specific protein, WASP, and its ubiquitously expressed ortholog N-WASP form part of a family that also includes the three WASP-family verprolin homologous protein (WAVE͞SCAR) isoforms: WAVE1, WAVE2, and WAVE3 (1, 3). Members of this family activate Arp2͞3-dependent actin nucleation and branching in response to signals mediated by Rho-family GTPases. Although the domain structure of these proteins varies, reflecting different modes of regulation, they all share a common C-terminal WH2 central-acidic region (CA region) (Fig. 1A), which constitutes the smallest fragment necessary for Arp2͞3 activation (4). WH2 is also present in members of the WASP-interacting protein (WIP) family, which form complexes with WASP͞N-WASP and modulate their functions in vivo (5, 6). Members of this family include ...

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Section: Resultssupporting

confidence: 87%

“…1C), together with published affinity values for various WH2 (6,15,16) and T␤ (21,22) domains, consistently show that WH2 binds actin with Ϸ10-fold higher affinity than T␤. Owing to the presence of the C-terminal ␣-helix, T␤ has a larger actin-binding interface than WH2 (Fig.…”

Section: Resultssupporting

confidence: 62%

Actin-bound structures of Wiskott–Aldrich syndrome protein (WASP)-homology domain 2 and the implications for filament assembly

Chéreau

Kerff

Graceffa

et al. 2005

Proc. Natl. Acad. Sci. U.S.A.

236

375

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“…However, we find that the Ct extension binds to F-actin but does not sequester monomers and only very weakly shifts the actin mobility in native gel assays (data not shown). This is not without precedent, since one of four WH2 domains found within tetrathymosin␤ has been shown to bind F-actin, but only relatively poorly to Gactin (Van Troys et al, 2004). Furthermore, espins are actinbundling proteins that contain a WH2 motif, which although it binds to G-actin also aids in bundle formation (Loomis et al, 2006).…”

Section: Discussionmentioning

confidence: 99%

Characterisation of IRTKS, a novel IRSp53/MIM family actin regulator with distinct filament bundling properties

Millard

Dawson

Machesky

2007

Journal of Cell Science

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IRSp53 is a scaffold protein that contains an IRSp53/MIM homology domain (IMD) that bundles actin filaments and interacts with the small GTPase Rac. IRSp53 also binds to the small GTPase Cdc42 and to Scar/WAVE and Mena/VASP proteins to regulate the actin cytoskeleton. We have characterised a novel IMD-containing protein, insulin receptor tyrosine kinase substrate (IRTKS), which has widespread tissue distribution, is a substrate for the insulin receptor and binds Rac. Unlike IRSp53, IRTKS does not interact with Cdc42. Expression of IRTKS induces clusters of short actin bundles rather than filopodia-like protrusions. This difference may be attributable to a short carboxyl-terminal (Ct) extension present on IRTKS, which resembles a WASP-homology 2 (WH2) motif. Addition of the Ct extension to IRSp53 causes an apparent shortening of bundles induced by the IMD in vitro, and in cultured cells, suggesting that the Ct extension of IRTKS modulates the organising activity of the IMD. Lastly, we could not detect actin monomer sequestration by the Ct extension of IRTKS as would be expected with a conventional WH2 motif, but it did interact with actin filaments.

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“…Since we were unable to demonstrate the presence of the two latter isoforms in early embryos, this absence, or the too low expression levels, may simply explain the lack of rescue of silencing PFN-1. Intriguingly, another protein, tetrathymosin β, with profilin-like activity is expressed in the dividing zygote [Van Troys et al, 2004] but neither is it able to overcome the lethal effects of PFN-1 silencing.…”

Section: Discussionmentioning

confidence: 99%

Caenorhabditis elegans expresses three functional profilins in a tissue‐specific manner

Polet

Lambrechts

Ono

et al. 2005

Cell Motil. Cytoskeleton

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Profilins are actin binding proteins, which also interact with polyphosphoinositides and proline-rich ligands. On the basis of the genome sequence, three diverse profilin homologues (PFN) are predicted to exist in Caenorhabditis elegans. We show that all three isoforms PFN-1, PFN-2, and PFN-3 are expressed in vivo and biochemical studies indicate they bind actin and influence actin dynamics in a similar manner. In addition, they bind poly(L-proline) and phosphatidylinositol 4,5-bisphosphate micelles. PFN-1 is essential whereas PFN-2 and PFN-3 are nonessential. Immunostainings revealed different expression patterns for the profilin isoforms. In embryos, PFN-1 localizes in the cytoplasm and to the cell-cell contacts at the early stages, and in the nerve ring during later stages. During late embryogenesis, expression of PFN-3 was specifically detected in body wall muscle cells. In adult worms, PFN-1 is expressed in the neurons, the vulva, and the somatic gonad, PFN-2 in the intestinal wall, the spermatheca, and the pharynx, and PFN-3 localizes in a striking dot-like fashion in body wall muscle. Thus the model organism Caenorhabditis elegans expresses three profilin isoforms and is the first invertebrate animal with tissue-specific profilin expression.

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TetraThymosinβ Is Required for Actin Dynamics inCaenorhabditis elegansand Acts via Functionally Different Actin-binding Repeats

Cited by 34 publications

References 54 publications

Actin-bound structures of Wiskott–Aldrich syndrome protein (WASP)-homology domain 2 and the implications for filament assembly

Actin-bound structures of Wiskott–Aldrich syndrome protein (WASP)-homology domain 2 and the implications for filament assembly

Characterisation of IRTKS, a novel IRSp53/MIM family actin regulator with distinct filament bundling properties

Caenorhabditis elegans expresses three functional profilins in a tissue‐specific manner

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