Structure, function, and evolution of the beta-thymosin/WH2 (WASP-Homology2) actin-binding module

Marie-France Carlier¹, Maud Hertzog, Dominique Didry

¹Cytoskeleton Dynamics and Motility, LEBS, CNRS, 1 Avenue de la Terrasse, 91198 Gif-sur-Yvette, France. carlier@lebs.cnrs-gif.fr.

Annals of the New York Academy of Sciences

|October 20, 2007

Summary

This study explores how a protein module called the WH2 domain regulates actin. Actin is a key player in cell movement and structure. The WH2 domain appears in many proteins with different functions. Some proteins sequester actin, others promote its assembly. The study used Tbeta4 and Ciboulot as models. Tbeta4 sequesters actin, while Ciboulot promotes assembly. Researchers found that structural changes in the WH2 domain can switch between these functions. Mutations in Tbeta4 can change its behavior. NMR studies showed how dynamic interactions with G-actin subdomains control function. Chimera proteins helped identify key regions. The findings explain how WH2 domains regulate actin dynamics.

Area of Science:

Structural biology of actin-binding proteins
Cell motility mechanisms in developmental biology
Protein evolution in molecular biomedicine

Background:

Actin-binding proteins regulate cytoskeletal dynamics. Thymosin beta4 is known to sequester G-actin. Recent discoveries show the beta-thymosin/WH2 module appears in many proteins with varied functions. These proteins influence cell motility through actin interactions. Prior research has shown that WH2 domains can inhibit or promote actin assembly. However, the structural basis for this functional diversity remains unclear. This gap motivated investigations into how WH2 domains evolve functionally. No prior work had resolved how structural changes affect actin-binding outcomes.

Purpose Of The Study:

The aim is to explore the functional evolution of the WH2 domain. Researchers focused on how structural changes influence actin-binding behavior. Tbeta4 and Ciboulot were used as model proteins for comparison. Tbeta4 sequesters actin, while Ciboulot promotes assembly. The study seeks to identify structural determinants of function. Mutagenesis and structural analysis were used to test functional switches. The goal is to understand how WH2 domains regulate actin dynamics. This work addresses the lack of clarity on WH2 domain versatility.

Main Methods:

The study combined structural and biochemical approaches. Crystallography was used to analyze Ciboulot-actin interactions. NMR spectroscopy tracked dynamic changes in WH2 domain interactions. Point mutagenesis altered Tbeta4's actin-binding behavior. Chimera proteins were engineered from Ciboulot and Tbeta4. Functional assays measured actin assembly promotion or inhibition. Structural analysis focused on subdomains 1 and 2 of G-actin. The approach tested how domain dynamics influence function.

Main Results:

Ciboulot's first repeat promotes actin assembly like profilin. The N-terminal helix of D1 interacts strongly with G-actin. Tbeta4 mutations can switch sequestering to profilin-like activity. NMR data show functional changes correlate with interaction dynamics. Chimera proteins displayed enhanced sequestering or profilin-like functions. Structural analysis revealed key residues in subdomains 1 and 2 of G-actin. Mutagenesis confirmed the role of central and C-terminal regions. These findings clarify how WH2 domains regulate actin-binding outcomes.

Conclusions:

The study clarifies structural determinants of WH2 domain function. Dynamic interactions with G-actin subdomains control activity. The N-terminal helix is critical for actin assembly promotion. Mutations can switch between sequestering and promoting functions. Chimera analysis supports the role of domain regions in function. The findings explain how WH2 domains regulate actin dynamics. Structural changes influence actin-binding outcomes directly. These results provide a framework for understanding WH2 domain versatility.

Keywords:

actin-binding protein function beta-thymosin WH2 domain actin sequestration mechanism structural biology of actin

Frequently Asked Questions

Point mutations in Tbeta4 can switch sequestering to profilin-like activity. NMR studies show this depends on interaction dynamics with G-actin subdomains.

The N-terminal amphipathic helix of Ciboulot's first repeat interacts strongly with G-actin. This region is key for promoting actin assembly.

NMR studies show the central region interacts with subdomain 1 of G-actin. Changes in this interaction correlate with functional switches between inhibition and promotion.

Chimeras of Ciboulot and Tbeta4 reveal structural determinants of function. They show how domain regions influence actin-binding behavior.

Tbeta4 mutations can switch sequestering to profilin-like activity. This depends on changes in interaction dynamics with G-actin subdomains.

The authors conclude that dynamic interactions with G-actin subdomains regulate function. Structural changes influence actin-binding outcomes directly.

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