Characterization of the interaction between Actinin-Associated LIM Protein (ALP) and the rod domain of α-actinin

Klaavuniemi, Tuula; Alho, Nanna; Hotulainen, Pirta; Kelloniemi, Annina; Havukainen, Heli; Permi, Perttu; Mattila, Sampo; Ylänne, Jari

doi:10.1186/1471-2121-10-22

Cited by 18 publications

(12 citation statements)

References 47 publications

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“…This is in agreement with the observation that 2 min noradrenaline stimulation induced the phosphorylation of ERM, which is required for its binding to EBP50 (68). Among these proteins, ␣-actinin has already been detected as an interacting partner of PDZ domain containing proteins (10,28,36,74). The identification by mass spectrometry of the proteins interacting with EBP50 in noradrenaline-stimulated arteries revealed that almost all the EBP50 interacting partners were related to the cytoskeleton.…”

Section: Discussionsupporting

confidence: 89%

Identification and functional implication of a Rho kinase-dependent moesin-EBP50 interaction in noradrenaline-stimulated artery

Baeyens

Horman

Vertommen

et al. 2010

American Journal of Physiology-Cell Physiology

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Ezrin, radixin, and moesin (ERM) proteins are known to be substrates of Rho kinase (ROCK), a key player in vascular smooth muscle regulation. Their function in arteries remains to be elucidated. The objective of the present study was to investigate ERM phosphorylation and function in rat aorta and mesenteric artery and the influence of ERM-binding phosphoprotein 50 (EBP50), a scaffold partner of ERM proteins in several cell types. In isolated arteries, ERM proteins are phosphorylated by PKC and ROCK with different kinetics after either agonist stimulation or KCl-induced depolarization. Immunoprecipitation of EBP50 in noradrenaline-stimulated arteries allowed identification of its interaction with moesin and several other proteins involved in cytoskeleton regulation. This interaction was inhibited by Y27632, a ROCK inhibitor. Moesin or EBP50 depletion after small interfering RNA transfection by reverse permeabilization in intact mesenteric arteries both potentiated the contractility in response to agonist stimulation without any effect on contractile response induced by high KCl. This effect was preserved in ionomycin-permeabilized arteries. These results indicate that, in agonist-stimulated arteries, the activation of ROCK leads to the binding of moesin to EBP50, which interacts with several components of the cytoskeleton, resulting in a decrease in the contractile response.

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Section: Discussionsupporting

confidence: 89%

Identification and functional implication of a Rho kinase-dependent moesin-EBP50 interaction in noradrenaline-stimulated artery

Baeyens

Horman

Vertommen

et al. 2010

American Journal of Physiology-Cell Physiology

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“…As expected, Zasp52 binds better to α-actinin than to actin. The dissociation constant is very similar to that of human ALP binding to α-actinin [33,34].…”

Section: Plos Onementioning

confidence: 56%

Characterizing the actin-binding ability of Zasp52 and its contribution to myofibril assembly

2020

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In sarcomeres, α-actinin crosslinks thin filaments and anchors them at the Z-disc. Drosophila melanogaster Zasp52 also localizes at Z-discs and interacts with α-actinin via its extended PDZ domain, thereby contributing to myofibril assembly and maintenance, yet the detailed mechanism of Zasp52 function is unknown. Here we show a strong genetic interaction between actin and Zasp52 during indirect flight muscle assembly, indicating that this interaction plays a critical role during myofibril assembly. Our results suggest that Zasp52 contains an actin-binding site, which includes the extended PDZ domain and the ZM region. Zasp52 binds with micromolar affinity to monomeric actin. A co-sedimentation assay indicates that Zasp52 can also bind to F-actin. Finally, we use in vivo rescue assays of myofibril assembly to show that the α-actinin-binding domain of Zasp52 is not sufficient for a full rescue of Zasp52 mutants suggesting additional contributions of Zasp52 actin-binding to myofibril assembly.

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“…However, only for ALP, RIL, CLP36, and ZASP/Cypher, the PDZ domain was confirmed to interact directly with α-actinin [7, 28, 39–41]. To further complicate the issue, the ZM region of ALP has been shown to bind directly to α-actinin [28, 42] and the ZM region, as well as the LIM domains of ZASP/Cypher colocalize with α-actinin [7, 29]. Moreover, LIM domains of CRP proteins can bind α-actinin [43, 44].…”

Section: Discussionmentioning

confidence: 99%

Zasp52, a Core Z-disc Protein in Drosophila Indirect Flight Muscles, Interacts with α-Actinin via an Extended PDZ Domain

2016

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Z-discs are organizing centers that establish and maintain myofibril structure and function. Important Z-disc proteins are α-actinin, which cross-links actin thin filaments at the Z-disc and Zasp PDZ domain proteins, which directly interact with α-actinin. Here we investigate the biochemical and genetic nature of this interaction in more detail. Zasp52 is the major Drosophila Zasp PDZ domain protein, and is required for myofibril assembly and maintenance. We show by in vitro biochemistry that the PDZ domain plus a C-terminal extension is the only area of Zasp52 involved in the interaction with α-actinin. In addition, site-directed mutagenesis of 5 amino acid residues in the N-terminal part of the PDZ domain, within the PWGFRL motif, abolish binding to α-actinin, demonstrating the importance of this motif for α-actinin binding. Rescue assays of a novel Zasp52 allele demonstrate the crucial importance of the PDZ domain for Zasp52 function. Flight assays also show that a Zasp52 mutant suppresses the α-actinin mutant phenotype, indicating that both proteins are core structural Z-disc proteins required for optimal Z-disc function.

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Characterization of the interaction between Actinin-Associated LIM Protein (ALP) and the rod domain of α-actinin

Cited by 18 publications

References 47 publications

Identification and functional implication of a Rho kinase-dependent moesin-EBP50 interaction in noradrenaline-stimulated artery

Identification and functional implication of a Rho kinase-dependent moesin-EBP50 interaction in noradrenaline-stimulated artery

Characterizing the actin-binding ability of Zasp52 and its contribution to myofibril assembly

Zasp52, a Core Z-disc Protein in Drosophila Indirect Flight Muscles, Interacts with α-Actinin via an Extended PDZ Domain

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