Explore the vast range of titles available.

The actin cytoskeleton is one of the most important players in cell motility, adhesion, division, and functioning. The regulation of specific microfilament formation largely determines cellular functions. The main actin-binding protein in animal cells is tropomyosin (Tpm). The unique structural and functional diversity of microfilaments is achieved through the diversity of Tpm isoforms. In our work, we studied the properties of the cytoplasmic isoforms Tpm1.8 and Tpm1.9. The results showed that these isoforms are highly thermostable and differ in the stability of their central and C-terminal fragments. The properties of these isoforms were largely determined by the 6th exons. Thus, the strength of the end-to-end interactions, as well as the affinity of the Tpm molecule for F-actin, differed between the Tpm1.8 and Tpm1.9 isoforms. They were determined by whether an alternative internal exon, 6a or 6b, was included in the Tpm isoform structure. The strong interactions of the Tpm1.8 and Tpm1.9 isoforms with F-actin led to the formation of rigid actin filaments, the stiffness of which was measured using an optical trap. It is quite possible that the structural and functional features of the Tpm isoforms largely determine the appearance of these isoforms in the rigid actin structures of the cell cortex.

Tropomyosins (Tpm) are the family of actin-binding proteins encoded by four genes in humans. Missense mutations in the TPM1 gene associated with cardiomyopathies have been studied in the sarcomeric isoform Tpm1.1. The cardiomyopathy-causing mutations E40K and E54K are located in exon 2b of the TPM1 gene and may be expressed in non-muscle cytoplasmic Tpm isoforms, including Tpm1.7, which is associated with early tissue development. In the present work, we investigate the effects of mutations E40K and E54K on the properties of Tpm1.7. The E40K and E54K mutations caused destabilization of the Tpm1.7 molecule at the N- and C-termini parts. Neither mutation affected the Tpm1.7 affinity for filamentous actin (F-actin). The bending stiffness of F-actin/Tpm1.7 E40K filaments was lower compared to F-actin/Tpm1.7 WT (wild-type). The interplay of Tpm1.7 and motor proteins was studied in an in vitro motility assay with skeletal myosin. Tpm1.7 WT reduced the sliding velocity of F-actin by half; the velocity of F-actin with Tpm1.7 E54K did not differ from that of bare F-actin; and Tpm1.7 E40K decreased the F-actin velocity by approximately threefold. While Tpm1.7 E40K did not affect the protective effect of Tpm1.7 against F-actin severing by cofilin-1, the E54K mutation enhanced protection against cofilin-1. Thus, cardiomyopathic mutations in the TPM1 gene can affect the properties of non-muscle Tpm isoforms, which indicates that this should be taken into account when studying the molecular mechanisms of the pathogenesis of these diseases.

Dynamic cytoskeletal homeostasis drives malignant transformation in tumor cells and represents a therapeutic vulnerability. Therapeutic targeting of this equilibrium may improve outcomes for cancer patients. Rac1 acts as a central molecular switch that controls actin cytoskeleton dynamics. Although multiple biological strategies modulate its spatiotemporal activity to maintain actin cytoskeleton homeostasis, the underlying molecular mechanisms remain unclear. Here, we identify FBXO44 as a critical regulator of Rac1 nucleocytoplasmic trafficking via its interaction with MTSS1. In gastric cancer (GC), FBXO44 directs two distinct ubiquitination programs on MTSS1: K63-linked polyubiquitination of MTSS1 promotes Rac1 nuclear translocation, whereas K11-linked polyubiquitination induces proteasomal degradation of MTSS1, restricting Rac1 nuclear entry. This ubiquitin-mediated coordination reprograms nucleocytoplasmic Rac1 signaling distribution and remodels the actin cytoskeleton. Structural analysis demonstrates that FBXO44 binds MTSS1 via distinct domains, dynamically balancing these opposing ubiquitination events by controlling MTSS1 abundance, thereby fine-tuning actin cytoskeletal dynamics. Clinically, this regulatory axis supports an oncogenic phenotype: FBXO44 overexpression correlates with enhanced Rac1 signaling and activation of associated pathways in advanced GC. Importantly, the expression balance of the FBXO44/MTSS1 axis significantly influences patient prognosis. Our findings provide mechanistic insights into cytoskeletal regulation and establish a translational framework for GC therapy.

Actin-binding proteins play a critical role in regulating the dynamic rearrangement of the actin cytoskeleton, which is essential for maintaining cellular homeostasis and facilitating various processes in eukaryotic cells. Cofilin-1 (Cfl1), an actin-binding protein, promotes the severing and depolymerization of actin filaments. To investigate the function of Cfl1 in mast cells, we generated Mcpt5-Cre-nf-Cfl1 fl/fl knock-in mice, expressing a non-functional form of Cfl1 (nf-Cfl1) instead of wildtype Cfl1 under the control of the connective tissue mast cell (CTMC)-specific promoter mast cell protease 5 (Mcpt5). Expression of nf-Cfl1 resulted in the complete absence of CTMCs. Notably, normal numbers of basophils were observed, in contrast to other mast cell-deficient mice. Interestingly, an inducible knock-in of nf-Cfl1 in mature mast cells did not affect the survival of mature mast cells. The Mcpt5-Cre-nf-Cfl1 fl/fl mice lacking CTMCs showed impaired induction of systemic anaphylaxis. However, they remained fully susceptible to 1-fluoro-2,4-dinitrobenzene-induced contact hypersensitivity and imiquimod-induced psoriasis-like dermatitis. In addition, clearance of vaccinia virus skin infection was unaltered. Thus, this study demonstrates that CTMCs are not essential in these inflammatory skin diseases. Deviating results in some other mast cell-deficient models suggest that the concomitant lack of basophils or residual CTMCs in these mouse models influence disease outcome. Taken together, the complete absence of CTMCs and the preserved presence of basophils in Mcpt5-Cre-nf-Cfl1 fl/fl mice establishes this model as a valuable tool for studying the specific role of CTMCs in different diseases.

Malfunction of the actin cytoskeleton is linked to numerous human diseases including neurological disorders and cancer. LIMK1 (LIM domain kinase 1) and its paralogue LIMK2 are two closely related kinases that control actin cytoskeleton dynamics. Consequently, they are potential therapeutic targets for the treatment of such diseases. In the present review, we describe the LIMK conformational space and its dependence on ligand binding. Furthermore, we explain the unique catalytic mechanism of the kinase, shedding light on substrate recognition and how LIMK activity is regulated. The structural features are evaluated for implications on the drug discovery process. Finally, potential future directions for targeting LIMKs pharmacologically, also beyond just inhibiting the kinase domain, are discussed.

A theory for diffusivity estimation for spatially extended activator–inhibitor dynamics modeling the evolution of intracellular signaling networks is developed in the mathematical framework of stochastic reaction–diffusion systems. In order to account for model uncertainties, we extend the results for parameter estimation for semilinear stochastic partial differential equations, as developed in Pasemann and Stannat (Electron J Stat 14(1):547–579, 2020), to the problem of joint estimation of diffusivity and parametrized reaction terms. Our theoretical findings are applied to the estimation of effective diffusivity of signaling components contributing to intracellular dynamics of the actin cytoskeleton in the model organism Dictyostelium discoideum .

Actin cytoskeleton dynamics is critical for variety of cellular events including cell elongation, division and morphogenesis, and is tightly regulated by numerous groups of actin binding proteins. However it is not well understood how these actin binding proteins are modulated in a physiological condition by their interaction proteins. In this study, we describe that Arabidopsis 14-3-3 λ protein interacted with actin depolymerizing factor 1 (ADF1) in plant to regulate F-actin stability and dynamics. Loss of 14-3-3 λ in Arabidopsis resulted in longer etiolated hypocotyls in dark and changed actin cytoskeleton architecture in hypocotyl cells. Overexpression of ADF1 repressed 14-3-3 λ mutant hypocotyl elongation and actin dynamic phenotype. In addition, the phosphorylation level of ADF1 was increased and the subcellular localization of ADF1 was altered in 14-3-3 λ mutant. Consistent with these observations, the actin filaments were more stable in 14-3-3 λ mutant. Our results indicate that 14-3-3 λ protein mediates F-actin dynamics possibly through inhibiting ADF1 phosphorylation in vivo .

This chapter contains sections titled: Definition Historical Perspectives Molecular Composition of the Domain Functional Implications and Roles for Domain Organization Future Perspectives Abbreviations References

SignificanceActin filaments are biopolymers that reside close to the plasma membrane in mammalian cells and are essential for its mechanical stability and shape. In vitro experiments have demonstrated that direct interactions between actin strands and lipid membranes exist and that the lipid composition and the ions in the buffer are of major importance. Here, we utilize molecular dynamics simulations to systematically analyze these effects on a close-to-atomistic resolution. We find that the charge of the lipid membrane determines the attraction/repulsion of the actin filaments, which can be reversed by the inclusion of divalent buffer ions. Our results are verified by an experimental reconstitution assay. These results suggest that direct actin binding can be used for engineering synthetic cells. The cytoskeletal protein actin polymerizes into filaments that are essential for the mechanical stability of mammalian cells. In vitro experiments showed that direct interactions between actin filaments and lipid bilayers are possible and that the net charge of the bilayer as well as the presence of divalent ions in the buffer play an important role. In vivo, colocalization of actin filaments and divalent ions are suppressed, and cells rely on linker proteins to connect the plasma membrane to the actin network. Little is known, however, about why this is the case and what microscopic interactions are important. A deeper understanding is highly beneficial, first, to obtain understanding in the biological design of cells and, second, as a possible basis for the building of artificial cortices for the stabilization of synthetic cells. Here, we report the results of coarse-grained molecular dynamics simulations of monomeric and filamentous actin in the vicinity of differently charged lipid bilayers. We observe that charges on the lipid head groups strongly determine the ability of actin to adsorb to the bilayer. The inclusion of divalent ions leads to a reversal of the binding affinity. Our in silico results are validated experimentally by reconstitution assays with actin on lipid bilayer membranes and provide a molecular-level understanding of the actin–membrane interaction.