Browse
Submit Data
Databases
API
Help

Dataset Information

0 Views

0 Connections

0 Citations

0 Reanalyses

0 Downloads

Omics score: 0

Extracellular regulated kinase (ERK) interaction with actin and the calponin homology (CH) domain of actin-binding proteins.

ABSTRACT: An interaction between extracellular regulated kinase 1 (ERK1) and calponin has previously been reported (Menice, Hulvershorn, Adam, Wang and Morgan (1997) J. Biol. Chem. 272 (40), 25157-25161) and has been suggested to reflect a function of calponin as a signalling molecule. We report in this study that calponin binds to both ERK1 and ERK2 under native conditions as well as in an overlay assay. Using chymotryptic fragments of calponin, the binding site of ERK on calponin was identified as the calponin homology (CH) domain, an N-terminal region of calponin found in other actin-binding proteins. ERK also bound, in a gel overlay assay, alpha-actinin, a protein with two tandem CH domains, as well as a 27 kDa thermolysin product of alpha-actinin containing the CH domains of alpha-actinin. The CH domain of calponin could compete with intact calponin or alpha-actinin for ERK binding. Titration of acrylodan-labelled calponin with ERK gave a K(a) of 6x10(6) M(-1) and titration of acrylodan-labelled calponin with a peptide from the alphaL16 helix of ERK gave a K(a) of 1x10(6) M(-1). Recombinant ERK was found to co-sediment with purified actin and induced a fluorescence change in pyrene-labelled F-actin (K(a)=5x10(6) M(-1)). The interaction of ERK with CH domains points to a new potential function for CH domains. The interaction of ERK with actin raises the possibility that actin may provide a scaffold for ERK signalling complexes in both muscle and non-muscle cells.

SUBMITTER: Leinweber BD

PROVIDER: S-EPMC1220621 | biostudies-other | 1999 Nov

REPOSITORIES: biostudies-other

ACCESS DATA

Json Xml

Similar Datasets

Steric regulation of tandem calponin homology domain actin-binding affinity.

Project description:Tandem calponin homology (CH1-CH2) domains are common actin-binding domains in proteins that interact with and organize the actin cytoskeleton. Despite regions of high sequence similarity, CH1-CH2 domains can have remarkably different actin-binding properties, with disease-associated point mutants known to increase as well as decrease affinity for F-actin. To investigate features that affect CH1-CH2 affinity for F-actin in cells and in vitro, we perturbed the utrophin actin-binding domain by making point mutations at the CH1-CH2 interface, replacing the linker domain, and adding a polyethylene glycol (PEG) polymer to CH2. Consistent with a previous model describing CH2 as a steric negative regulator of actin binding, we find that utrophin CH1-CH2 affinity is both increased and decreased by modifications that change the effective "openness" of CH1 and CH2 in solution. We also identified interface mutations that caused a large increase in affinity without changing solution "openness," suggesting additional influences on affinity. Interestingly, we also observe nonuniform subcellular localization of utrophin CH1-CH2 that depends on the N-terminal flanking region but not on bulk affinity. These observations provide new insights into how small sequence changes, such as those found in diseases, can affect CH1-CH2 binding properties.

| S-EPMC6938246 | biostudies-literature

Calponin-homology domain mediated bending of membrane-associated actin filaments.

Project description:Actin filaments are central to numerous biological processes in all domains of life. Driven by the interplay with molecular motors, actin binding and actin modulating proteins, the actin cytoskeleton exhibits a variety of geometries. This includes structures with a curved geometry such as axon-stabilizing actin rings, actin cages around mitochondria and the cytokinetic actomyosin ring, which are generally assumed to be formed by short linear filaments held together by actin cross-linkers. However, whether individual actin filaments in these structures could be curved and how they may assume a curved geometry remains unknown. Here, we show that 'curly', a region from the IQGAP family of proteins from three different organisms, comprising the actin-binding calponin-homology domain and a C-terminal unstructured domain, stabilizes individual actin filaments in a curved geometry when anchored to lipid membranes. Although F-actin is semi-flexible with a persistence length of ~10 μm, binding of mobile curly within lipid membranes generates actin filament arcs and full rings of high curvature with radii below 1 μm. Higher rates of fully formed actin rings are observed in the presence of the actin-binding coiled-coil protein tropomyosin and when actin is directly polymerized on lipid membranes decorated with curly. Strikingly, curly induced actin filament rings contract upon the addition of muscle myosin II filaments and expression of curly in mammalian cells leads to highly curved actin structures in the cytoskeleton. Taken together, our work identifies a new mechanism to generate highly curved actin filaments, which opens a range of possibilities to control actin filament geometries, that can be used, for example, in designing synthetic cytoskeletal structures.

| S-EPMC8315802 | biostudies-literature

Structural Characteristics, Binding Partners and Related Diseases of the Calponin Homology (CH) Domain.

Project description:The calponin homology (CH) domain is one of the most common modules in various actin-binding proteins and is characterized by an α-helical fold. The CH domain plays important regulatory roles in both cytoskeletal dynamics and signaling. The CH domain is required for stability and organization of the actin cytoskeleton, calcium mobilization and activation of downstream pathways. The CH domain has recently garnered increased attention due to its importance in the onset of different diseases, such as cancers and asthma. However, many roles of the CH domain in various protein functions and corresponding diseases are still unclear. Here, we review current knowledge about the structural features, interactome and related diseases of the CH domain.

| S-EPMC7240100 | biostudies-literature

The N-terminal actin-binding tandem calponin-homology (CH) domain of dystrophin is in a closed conformation in solution and when bound to F-actin.

Project description:Deficiency of the vital muscle protein dystrophin triggers Duchenne/Becker muscular dystrophy, but the structure-function relationship of dystrophin is poorly understood. To date, molecular structures of three dystrophin domains have been determined, of which the N-terminal actin-binding domain (N-ABD or ABD1) is of particular interest. This domain is composed of two calponin-homology (CH) domains, which form an important class of ABDs in muscle proteins. A previously determined x-ray structure indicates that the dystrophin N-ABD is a domain-swapped dimer, with each monomer adopting an extended, open conformation in which the two CH domains do not interact. This structure is controversial because it contradicts functional studies and known structures of similar ABDs from other muscle proteins. Here, we investigated the solution conformation of the dystrophin N-ABD using a very simple and elegant technique of pyrene excimer fluorescence. Using the wild-type protein, which contains two cysteines, and the corresponding single-cysteine mutants, we show that the protein is a monomer in solution and is in a closed conformation in which the two CH domains seem to interact, as observed from the excimer fluorescence of pyrene-labeled wild-type protein. Excimer fluorescence was also observed in its actin-bound form, indicating that the dystrophin N-ABD binds to F-actin in a closed conformation. Comparison of the dystrophin N-ABD conformation with other ABDs indicates that the tandem CH domains in general may be monomeric in solution and predominantly occur in closed conformation, whereas their actin-bound conformations may differ.

| S-EPMC3491715 | biostudies-literature

Large-scale opening of utrophin's tandem calponin homology (CH) domains upon actin binding by an induced-fit mechanism.

Project description:We have used site-directed spin labeling and pulsed electron paramagnetic resonance to resolve a controversy concerning the structure of the utrophin-actin complex, with implications for the pathophysiology of muscular dystrophy. Utrophin is a homolog of dystrophin, the defective protein in Duchenne and Becker muscular dystrophies, and therapeutic utrophin derivatives are currently being developed. Both proteins have a pair of N-terminal calponin homology (CH) domains that are important for actin binding. Although there is a crystal structure of the utrophin actin-binding domain, electron microscopy of the actin-bound complexes has produced two very different structural models, in which the CH domains are in open or closed conformations. We engineered a pair of labeling sites in the CH domains of utrophin and used dipolar electron-electron resonance to determine the distribution of interdomain distances with high resolution. We found that the two domains are flexibly connected in solution, indicating a dynamic equilibrium between two distinct open structures. Upon actin binding, the two domains become dramatically separated and ordered, indicating a transition to a single open and extended conformation. There is no trace of this open conformation of utrophin in the absence of actin, providing strong support for an induced-fit model of actin binding.

| S-EPMC3150896 | biostudies-literature

Dynamic Regulation of ?-Actinin's Calponin Homology Domains on F-Actin.

Project description:?-Actinin is an essential actin cross-linker involved in cytoskeletal organization and dynamics. The molecular conformation of ?-actinin's actin-binding domain (ABD) regulates its association with actin and thus mutations in this domain can lead to severe pathogenic conditions. A point mutation at lysine 255 in human ?-actinin-4 to glutamate increases the binding affinity resulting in stiffer cytoskeletal structures. The role of different ABD conformations and the effect of K255E mutation on ABD conformations remain elusive. To evaluate the impact of K255E mutation on ABD binding to actin we use all-atom molecular dynamics and free energy calculation methods and study the molecular mechanism of actin association in both wild-type ?-actinin and in the K225E mutant. Our models illustrate that the strength of actin association is indeed sensitive to the ABD conformation, predict the effect of K255E mutation--based on simulations with the K237E mutant chicken ?-actinin--and evaluate the mechanism of ?-actinin binding to actin. Furthermore, our simulations showed that the calmodulin domain binding to the linker region was important for regulating the distance between actin and ABD. Our results provide valuable insights into the molecular details of this critical cellular phenomenon and further contribute to an understanding of cytoskeletal dynamics in health and disease.

| S-EPMC4816760 | biostudies-literature

Hypertrophic cardiomyopathy mutations in the calponin-homology domain of ACTN2 affect actin binding and cardiomyocyte Z-disc incorporation.

Project description:?-Actinin-2 (ACTN2) is the only muscle isoform of ?-actinin expressed in cardiac muscle. Mutations in this protein have been implicated in mild to moderate forms of hypertrophic cardiomyopathy (HCM). We have investigated the effects of two mutations identified from HCM patients, A119T and G111V, on the secondary and tertiary structure of a purified actin binding domain (ABD) of ACTN2 by circular dichroism and X-ray crystallography, and show small but distinct changes for both mutations. We also find that both mutants have reduced F-actin binding affinity, although the differences are not significant. The full length mEos2 tagged protein expressed in adult cardiomyocytes shows that both mutations additionally affect Z-disc localization and dynamic behaviour. Overall, these two mutations have small effects on structure, function and behaviour, which may contribute to a mild phenotype for this disease.

| S-EPMC4980809 | biostudies-literature

Opening of tandem calponin homology domains regulates their affinity for F-actin.

Project description:Many actin-binding proteins contain calponin homology (CH) domains, but the manner in which these domains interact with F-actin has been controversial. Crystal structures have shown the tandem CH domains of alpha-actinin to be in a compact, closed conformation, but the interpretations of complexes of such tandem CH domains with F-actin have been ambiguous. We show that the tandem CH domains of alpha-actinin bind F-actin in an open conformation, explaining mutations that cause human diseases and suggesting that the opening of these domains may be one of the main regulatory mechanisms for proteins with tandem CH domains.

| S-EPMC2921939 | biostudies-literature

h3/Acidic calponin: an actin-binding protein that controls extracellular signal-regulated kinase 1/2 activity in nonmuscle cells.

Project description:Migration of fibroblasts is important in wound healing. Here, we demonstrate a role and a mechanism for h3/acidic calponin (aCaP, CNN3) in REF52.2 cell motility, a fibroblast line rich in actin filaments. We show that the actin-binding protein h3/acidic calponin associates with stress fibers in the absence of stimulation but is targeted to the cell cortex and podosome-like structures after stimulation with a phorbol ester, phorbol-12,13-dibutyrate (PDBu). By coimmunoprecipitation and colocalization, we show that extracellular signal-regulated kinase (ERK)1/2 and protein kinase C (PKC)alpha constitutively associate with h3/acidic calponin and are cotargeted with h3/acidic calponin in the presence of PDBu. This targeting can be blocked by a PKC inhibitor but does not require phosphorylation of h3/acidic calponin at the PKC sites S175 or T184. Knockdown of h3/acidic calponin results in a loss of PDBu-mediated ERK1/2 targeting, whereas PKCalpha targeting is unaffected. Caldesmon is an actin-binding protein that regulates actomyosin interactions and is a known substrate of ERK1/2. Both ERK1/2 activity and nonmuscle l-caldesmon phosphorylation are blocked by h3/acidic calponin knockdown. Furthermore, h3/acidic calponin knockdown inhibits REF52.2 migration in an in vitro wound healing assay. Our findings are consistent with a model whereby h3/acidic calponin controls fibroblast migration by regulation of ERK1/2-mediated l-caldesmon phosphorylation.

| S-EPMC2854098 | biostudies-literature

A kinesin with calponin-homology domain is involved in premitotic nuclear migration.

Project description:Interaction and cross-talk between microtubules and actin microfilaments are important for numerous processes during plant growth and development, including the control of cell elongation and tissue expansion, but little is known about the molecular components of this interaction. Plant kinesins with the calponin-homology domain (KCH) were recently identified and associated with a putative role in microtubule-microfilament cross-linking. The putative biological role of the rice KCH member OsKCH1 is addressed here using a combined approach with Tos17 kch1 knock-out mutants on the one hand, and a KCH1 overexpression line generated in tobacco BY-2 cells. It is shown that OsKCH1 is expressed in a development and tissue-specific manner in rice and antagonistic cell elongation and division phenotypes as a result of knock-down and overexpression are reported. Further, the dynamic repartitioning of OsKCH1 during the cell cycle is described and it is demonstrated that KCH overexpression delays nuclear positioning and mitosis in BY-2 cells. These findings are discussed with respect to a putative role of KCHs as linkers between actin filaments and microtubules during nuclear positioning.

| S-EPMC2905203 | biostudies-literature

OmicsDI is part of the ELIXIR infrastructure

OmicsDI is an Elixir interoperability service. Learn more ›

Tweets

OmicsDI Databases

PRIDE
PeptideAtlas
MassIVE
JPOST Repository
Physiome Model Repository

EGA
EVA
ENA
LINCS
PAXDB
Cell Collective

MetaboLights
Metabolomics Workbench
MetabolomeExpress
GNPS
BioModels
FAIRDOMHub

ArrayExpress
dbGaP
ExpressionAtlas
GEO
NODE

Information

Databases
Help
API
Contact us
Code on GitHub
Terms of use
Submit Data