Interactions with Actin Monomers, Actin Filaments, and Arp2/3 Complex Define the Roles of WASP Family Proteins and Cortactin in Coordinately Regulating Branched Actin Networks

Helgeson, Luke A.; Prendergast, Julianna G.; Wagner, Andrew R.; Rodnick-Smith, Max; Nolen, Brad J.

doi:10.1074/jbc.m114.587527

Cited by 54 publications

(50 citation statements)

References 77 publications

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“…The direction of synaptic morphologic changes, to retract or to sprout, may depend on the balance of actin depolymerization and polymerization, the latter being regulated by Rac/Cdc42 pathways as stated above. [39][40][41] In our study, the contribution of Pak to both retraction and growth suggests that although process growth in isolated rod cells appears to be a late-stage development, the Rac/Cdc42 growth pathways may be activated early on. However, when and where actin filament nucleation and elongation begins still needs to be examined.…”

Section: Discussionmentioning

confidence: 58%

“…19,20,[35][36][37] The role of Rac and Cdc42 in promoting growth is achieved through actin filament nucleation by ARP2/3 and Pak activities. [39][40][41] We speculate that the role of Pak in retraction may be to promote depolymerization of actin filaments through LIMK activity. Such depolymerization would be necessary to form new actin cytoskeleton in either filopodia or lamellipodia.…”

Section: Discussionmentioning

confidence: 92%

“…5,7,38 LIM kinase is well known to function in regulating actin polymerization in cooperation with the pathways that promote nucleation and extension of new actin filament branches. [39][40][41] Our study therefore also investigates the effect of LIMK inhibition on process outgrowth and explores the possibility that using inhibition of a single kinase, LIMK, can reduce both forms of structural plasticity, axonal retraction and neuritic sprouting.…”

mentioning

confidence: 99%

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LIM Kinase, a Newly Identified Regulator of Presynaptic Remodeling by Rod Photoreceptors After Injury

Wang

Townes‐Anderson

2015

Invest. Ophthalmol. Vis. Sci.

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METHODS. Phosphorylated LIMK (p-LIMK), the active form of LIMK, was examined in salamander retina with Western blot and confocal microscopy. Axon length within the first 7 hours and process growth after 3 days of culture were assessed in isolated rod photoreceptors treated with inhibitors of upstream regulators ROCK and p21-activated kinase (Pak) (Y27632 and IPA-3) and a direct LIMK inhibitor (BMS-5). Porcine retinal explants were also treated with BMS-5 and analyzed 24 hours after detachment. Because Ca 2þ influx contributes to axonal retraction, L-type channels were blocked in some experiments with nicardipine. RESULTS.Phosphorylated LIMK is present in rod terminals during retraction and in newly formed processes. Axonal retraction over 7 hours was significantly reduced by inhibition of LIMK or its regulators, ROCK and Pak. Process growth was reduced by LIMK or Pak inhibition especially at the basal (axon-bearing) region of the rod cells. Combining Ca 2þ channel and LIMK inhibition had no additional effect on retraction but did further inhibit sprouting after 3 days. In detached porcine retina, LIMK inhibition reduced rod axonal retraction and improved retinal morphology.CONCLUSIONS. Thus structural remodeling, in the form of either axonal retraction or neuritic growth, requires LIMK activity. LIM kinase inhibition may have therapeutic potential for reducing pathologic rod terminal plasticity after retinal injury.

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Section: Discussionmentioning

confidence: 58%

Section: Discussionmentioning

confidence: 92%

mentioning

confidence: 99%

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LIM Kinase, a Newly Identified Regulator of Presynaptic Remodeling by Rod Photoreceptors After Injury

Wang

Townes‐Anderson

2015

Invest. Ophthalmol. Vis. Sci.

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“…Although little is known about this class of NPFs, we previously used the cross-linking assay to show that WDS protein Dip1 stimulates formation of the shortpitch conformation, and our data here suggest this stimulation is sufficient for Dip1-mediated activation of the complex (5). Like the WDS family proteins, type II NPFs such as cortactin and fission yeast myosin I interact with the Arp2/3 complex and actin filaments but not with actin monomers (37,(42)(43)(44). Unlike WDS proteins, type II NPFs are generally weak activators on their own (7).…”

Section: Discussionmentioning

confidence: 60%

Role and structural mechanism of WASP-triggered conformational changes in branched actin filament nucleation by Arp2/3 complex

Rodnick-Smith

Luan

Liu

et al. 2016

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

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The Arp2/3 (Actin-related proteins 2/3) complex is activated by WASP (Wiskott-Aldrich syndrome protein) family proteins to nucleate branched actin filaments that are important for cellular motility. WASP recruits actin monomers to the complex and stimulates movement of Arp2 and Arp3 into a "short-pitch" conformation that mimics the arrangement of actin subunits within filaments. The relative contribution of these functions in Arp2/3 complex activation and the mechanism by which WASP stimulates the conformational change have been unknown. We purified budding yeast Arp2/3 complex held in or near the short-pitch conformation by an engineered covalent cross-link to determine if the WASP-induced conformational change is sufficient for activity. Remarkably, cross-linked Arp2/3 complex bypasses the need for WASP in activation and is more active than WASP-activated Arp2/3 complex. These data indicate that stimulation of the short-pitch conformation is the critical activating function of WASP and that monomer delivery is not a fundamental requirement for nucleation but is a specific requirement for WASP-mediated activation. During activation, WASP limits nucleation rates by releasing slowly from nascent branches. The cross-linked complex is inhibited by WASP's CA region, even though CA potently stimulates cross-linking, suggesting that slow WASP detachment masks the activating potential of the short-pitch conformational switch. We use structure-based mutations and WASP-Arp fusion chimeras to determine how WASP stimulates movement toward the short-pitch conformation. Our data indicate that WASP displaces the autoinhibitory Arp3 C-terminal tail from a hydrophobic groove at Arp3′s barbed end to destabilize the inactive state, providing a mechanism by which WASP stimulates the short-pitch conformation and activates Arp2/3 complex.actin | Arp2/3 | WASP

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“…It was previously thought that Arp2/3 complex was activated by a single NPF, but a series of studies have now shown that the complex is maximally activated by two NPFs [47-50]. Thus, like in other nucleators, the WH2 domains of NPFs play an auxiliary role in nucleation, by delivering actin subunits at the barbed end of the nucleus formed by Arp2 and Arp3 ( Figure 5 ).…”

Section: Opinionmentioning

confidence: 95%

The WH2 Domain and Actin Nucleation: Necessary but Insufficient

Domínguez

2016

Trends in Biochemical Sciences

117

144

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Two types of sequences, proline-rich domains (PRDs) and the WASP-Homology 2 (WH2) domain, are found in most actin filament nucleation and elongation factors discovered thus far. PRDs serve as a platform for protein-protein interactions, often mediating the binding of profilin-actin. The WH2 domain is an abundant actin monomer-binding motif consisting of ~17-aa. It frequently occurs in tandem repeats, and functions in nucleation by recruiting actin subunits to form the polymerization nucleus. It is found in Spire, Cobl, Lmod, Arp2/3 complex activators (WASP, WHAMM, WAVE, etc.), the bacterial nucleators VopL/VopF and Sca2 and some formins. Yet, it is argued here that the WH2 domain plays only an auxiliary role in nucleation, always synergizing with other domains or proteins for this activity.

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Interactions with Actin Monomers, Actin Filaments, and Arp2/3 Complex Define the Roles of WASP Family Proteins and Cortactin in Coordinately Regulating Branched Actin Networks

Cited by 54 publications

References 77 publications

LIM Kinase, a Newly Identified Regulator of Presynaptic Remodeling by Rod Photoreceptors After Injury

LIM Kinase, a Newly Identified Regulator of Presynaptic Remodeling by Rod Photoreceptors After Injury

Role and structural mechanism of WASP-triggered conformational changes in branched actin filament nucleation by Arp2/3 complex

The WH2 Domain and Actin Nucleation: Necessary but Insufficient

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