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BMC Biology Jun 2022The establishment of tissue architecture requires coordination between distinct processes including basement membrane assembly, cell adhesion, and polarity; however, the...
BACKGROUND
The establishment of tissue architecture requires coordination between distinct processes including basement membrane assembly, cell adhesion, and polarity; however, the underlying mechanisms remain poorly understood. The actin cytoskeleton is ideally situated to orchestrate tissue morphogenesis due to its roles in mechanical, structural, and regulatory processes. However, the function of many pivotal actin-binding proteins in mammalian development is poorly understood.
RESULTS
Here, we identify a crucial role for anillin (ANLN), an actin-binding protein, in orchestrating epidermal morphogenesis. In utero RNAi-mediated silencing of Anln in mouse embryos disrupted epidermal architecture marked by adhesion, polarity, and basement membrane defects. Unexpectedly, these defects cannot explain the profoundly perturbed epidermis of Anln-depleted embryos. Indeed, even before these defects emerge, Anln-depleted epidermis exhibits abnormalities in mitotic rounding and its associated processes: chromosome segregation, spindle orientation, and mitotic progression, though not in cytokinesis that was disrupted only in Anln-depleted cultured keratinocytes. We further show that ANLN localizes to the cell cortex during mitotic rounding, where it regulates the distribution of active RhoA and the levels, activity, and structural organization of the cortical actomyosin proteins.
CONCLUSIONS
Our results demonstrate that ANLN is a major regulator of epidermal morphogenesis and identify a novel role for ANLN in mitotic rounding, a near-universal process that governs cell shape, fate, and tissue morphogenesis.
Topics: Actin Cytoskeleton; Animals; Contractile Proteins; Cytokinesis; Mammals; Mice; Microfilament Proteins
PubMed: 35710398
DOI: 10.1186/s12915-022-01345-9 -
The Journal of Biological Chemistry Apr 2012SCAB1 is a novel plant-specific actin-binding protein that binds, bundles, and stabilizes actin filaments and regulates stomatal movement. Here, we dissected the...
SCAB1 is a novel plant-specific actin-binding protein that binds, bundles, and stabilizes actin filaments and regulates stomatal movement. Here, we dissected the structure and function of SCAB1 by structural and biochemical approaches. We show that SCAB1 is composed of an actin-binding domain, two coiled-coil (CC) domains, and a fused immunoglobulin and pleckstrin homology (Ig-PH) domain. We determined crystal structures for the CC1 and Ig-PH domains at 1.9 and 1.7 Å resolution, respectively. The CC1 domain adopts an antiparallel helical hairpin that further dimerizes into a four-helix bundle. The CC2 domain also mediates dimerization. At least one of the coiled coils is required for actin binding, indicating that SCAB1 is a bivalent actin cross-linker. The key residues required for actin binding were identified. The PH domain lacks a canonical basic phosphoinositide-binding pocket but can bind weakly to inositol phosphates via a basic surface patch, implying the involvement of inositol signaling in SCAB1 regulation. Our results provide novel insights into the functional organization of SCAB1.
Topics: Actins; Amino Acid Motifs; Amino Acid Sequence; Arabidopsis; Arabidopsis Proteins; Binding Sites; Chromatography, Gel; Conserved Sequence; Crystallography, X-Ray; Hydrogen Bonding; Inositol Phosphates; Microfilament Proteins; Models, Molecular; Molecular Sequence Data; Protein Binding; Protein Interaction Domains and Motifs; Protein Multimerization; Protein Structure, Quaternary; Surface Properties
PubMed: 22356912
DOI: 10.1074/jbc.M111.338525 -
Cell Structure and Function Dec 2023Directional cell rearrangement is a critical process underlying correct tissue deformation during morphogenesis. Although the involvement of F-actin regulation in cell...
Directional cell rearrangement is a critical process underlying correct tissue deformation during morphogenesis. Although the involvement of F-actin regulation in cell rearrangement has been established, the role and regulation of actin binding proteins (ABPs) in this process are not well understood. In this study, we investigated the function of Coronin-1, a WD-repeat actin-binding protein, in controlling directional cell rearrangement in the Drosophila pupal wing. Transgenic flies expressing Coronin-1-EGFP were generated using CRISPR-Cas9. We observed that Coronin-1 localizes at the reconnecting junction during cell rearrangement, which is dependent on actin interacting protein 1 (AIP1) and cofilin, actin disassemblers and known regulators of wing cell rearrangement. Loss of Coronin-1 function reduces cell rearrangement directionality and hexagonal cell fraction. These results suggest that Coronin-1 promotes directional cell rearrangement via its interaction with AIP1 and cofilin, highlighting the role of ABPs in the complex process of morphogenesis.Key words: morphogenesis, cell rearrangement, actin binding proteins (ABPs).
Topics: Animals; Drosophila; Microfilament Proteins; Actins; Actin Depolymerizing Factors; Epithelium
PubMed: 38030242
DOI: 10.1247/csf.23049 -
Molecular Biology of the Cell Jan 2000Toxoplasma gondii relies on its actin cytoskeleton to glide and enter its host cell. However, T. gondii tachyzoites are known to display a strikingly low amount of actin...
Toxoplasma gondii relies on its actin cytoskeleton to glide and enter its host cell. However, T. gondii tachyzoites are known to display a strikingly low amount of actin filaments, which suggests that sequestration of actin monomers could play a key role in parasite actin dynamics. We isolated a 27-kDa tachyzoite protein on the basis of its ability to bind muscle G-actin and demonstrated that it interacts with parasite G-actin. Cloning and sequence analysis of the gene coding for this protein, which we named Toxofilin, showed that it is a novel actin-binding protein. In in vitro assays, Toxofilin not only bound to G-actin and inhibited actin polymerization as an actin-sequestering protein but also slowed down F-actin disassembly through a filament end capping activity. In addition, when green fluorescent protein-tagged Toxofilin was overexpressed in mammalian nonmuscle cells, the dynamics of actin stress fibers was drastically impaired, whereas green fluorescent protein-Toxofilin copurified with G-actin. Finally, in motile parasites, during gliding or host cell entry, Toxofilin was localized in the entire cytoplasm, including the rear end of the parasite, whereas in intracellular tachyzoites, especially before they exit from the parasitophorous vacuole of their host cell, Toxofilin was found to be restricted to the apical end.
Topics: Actin Capping Proteins; Actins; Animals; Base Sequence; Cytosol; DNA, Protozoan; HeLa Cells; Humans; Microfilament Proteins; Molecular Sequence Data; Protozoan Proteins; Recombinant Fusion Proteins; Toxoplasma
PubMed: 10637313
DOI: 10.1091/mbc.11.1.355 -
Acta Crystallographica. Section F,... Feb 2016Asymmetric stem-cell divisions are fundamental for morphogenesis and tissue homeostasis. They rely on the coordination between cortical polarity and the orientation of...
Asymmetric stem-cell divisions are fundamental for morphogenesis and tissue homeostasis. They rely on the coordination between cortical polarity and the orientation of the mitotic spindle, which is orchestrated by microtubule pulling motors recruited at the cortex by NuMA-LGN-Gαi complexes. LGN has emerged as a central component of the spindle-orientation pathway that is conserved throughout species. Its domain structure consists of an N-terminal TPR domain associating with NuMA, followed by four GoLoco motifs binding to Gαi subunits. The LGN(TPR) region is also involved in interactions with other membrane-associated proteins ensuring the correct cortical localization of microtubule motors, among which is the junctional protein afadin. To investigate the architecture of LGN(TPR) in complex with afadin, a chimeric fusion protein with a native linker derived from the region of afadin upstream of the LGN-binding domain was generated. The fusion protein behaves as a globular monomer in solution and readily crystallizes in the presence of sulfate-containing reservoirs. The crystals diffracted to 3.0 Å resolution and belonged to the cubic space group P213, with unit-cell parameter a = 170.3 Å. The structure of the engineered protein revealed that the crystal packing is promoted by the coordination of sulfate ions by residues of the afadin linker region and LGN(TPR).
Topics: Actins; Amino Acid Sequence; Crystallization; Crystallography, X-Ray; Humans; Intracellular Signaling Peptides and Proteins; Microfilament Proteins; Models, Molecular; Molecular Sequence Data; Protein Binding; Protein Conformation; Protein Stability; X-Ray Diffraction
PubMed: 26841766
DOI: 10.1107/S2053230X16000807 -
Trends in Cell Biology Jan 2011Nebulin, a giant, actin-binding protein, is the largest member of a family of proteins (including N-RAP, nebulette, lasp-1 and lasp-2) that are assembled in a variety of... (Review)
Review
Nebulin, a giant, actin-binding protein, is the largest member of a family of proteins (including N-RAP, nebulette, lasp-1 and lasp-2) that are assembled in a variety of cytoskeletal structures, and expressed in different tissues. For decades, nebulin has been thought to act as a molecular ruler, specifying the precise length of actin filaments in skeletal muscle. However, emerging evidence suggests that nebulin should not be viewed as a ruler but as an actin filament stabilizer required for length maintenance. Nebulin has also been implicated recently in an array of regulatory functions independent of its role in actin filament length regulation. In this review, we discuss the current evolutionary, biochemical, and functional data for the nebulin family of proteins - a family whose members, both large and small, function as cytoskeletal scaffolds and stabilizers.
Topics: Actin Cytoskeleton; Animals; Evolution, Molecular; Humans; Microfilament Proteins; Muscle Proteins; Muscle, Skeletal; Myocardium
PubMed: 20951588
DOI: 10.1016/j.tcb.2010.09.005 -
Small GTPases 2014Eukaryotic cells have evolved a variety of actin-binding proteins to regulate the architecture and the dynamics of the actin cytoskeleton in time and space. The... (Review)
Review
Eukaryotic cells have evolved a variety of actin-binding proteins to regulate the architecture and the dynamics of the actin cytoskeleton in time and space. The Diaphanous-related formins (DRF) represent a diverse group of Rho-GTPase-regulated actin regulators that control a range of actin structures composed of tightly-bundled, unbranched actin filaments as found in stress fibers and in filopodia. Under resting conditions, DRFs are auto-inhibited by an intra-molecular interaction between the C-terminal and the N-terminal domains. The auto-inhibition is thought to be released by binding of an activated RhoGTPase to the N-terminal GTPase-binding domain (GBD). However, there is growing evidence for more sophisticated variations from this simplified linear activation model. In this review we focus on the formin homology domain-containing proteins (FHOD), an unconventional group of DRFs. Recent findings on the molecular control and cellular functions of FHOD proteins in vivo are discussed in the light of the phylogeny of FHOD proteins.
Topics: Actin Cytoskeleton; Animals; Humans; Microfilament Proteins; Protein Structure, Tertiary; rho GTP-Binding Proteins
PubMed: 25483300
DOI: 10.4161/21541248.2014.973765 -
The Journal of Cell Biology Dec 1997Yeast verprolin, encoded by VRP1, is implicated in cell growth, cytoskeletal organization, endocytosis and mitochondrial protein distribution and function. We show that...
Yeast verprolin, encoded by VRP1, is implicated in cell growth, cytoskeletal organization, endocytosis and mitochondrial protein distribution and function. We show that verprolin is also required for bipolar bud-site selection. Previously we reported that additional actin suppresses the temperature-dependent growth defect caused by a mutation in VRP1. Here we show that additional actin suppresses all known defects caused by vrp1-1 and conclude that the defects relate to an abnormal cytoskeleton. Using the two-hybrid system, we show that verprolin binds actin. An actin-binding domain maps to the LKKAET hexapeptide located in the first 70 amino acids. A similar hexapeptide in other acting-binding proteins was previously shown to be necessary for actin-binding activity. The entire 70- amino acid motif is conserved in novel higher eukaryotic proteins that we predict to be actin-binding, and also in the actin-binding proteins, WASP and N-WASP. Verprolin-GFP in live cells has a cell cycle-dependent distribution similar to the actin cortical cytoskeleton. In fixed cells hemagglutinin-tagged Vrp1p often co-localizes with actin in cortical patches. However, disassembly of the actin cytoskeleton using Latrunculin-A does not alter verprolin's location, indicating that verprolin establishes and maintains its location independent of the actin cytoskeleton. Verprolin is a new member of the actin-binding protein family that serves as a polarity development protein, perhaps by anchoring actin. We speculate that the effects of verprolin upon the actin cytoskeleton might influence mitochondrial protein sorting/function via mRNA distribution.
Topics: Actins; Alleles; Amino Acid Sequence; Binding Sites; Cell Polarity; Cytoskeleton; Endocytosis; Fungal Proteins; Microfilament Proteins; Mitochondria; Molecular Sequence Data; Morphogenesis; Nucleic Acid Hybridization; Phenotype; Saccharomyces cerevisiae; Saccharomyces cerevisiae Proteins; Sequence Homology, Amino Acid
PubMed: 9412475
DOI: 10.1083/jcb.139.7.1821 -
The Journal of Biological Chemistry Dec 1986Actobindin is a new actin-binding protein isolated from Acanthamoeba castellanii. It is composed of two possibly identical polypeptide chains of approximately 13,000...
Actobindin is a new actin-binding protein isolated from Acanthamoeba castellanii. It is composed of two possibly identical polypeptide chains of approximately 13,000 daltons, as determined by sodium dodecyl sulfate polyacrylamide gel electrophoresis, and with isoelectric points of 5.9. In the native state, actobindin appears to be a dimer of about 25,000 daltons by sedimentation equilibrium analysis. It contains no tryptophan and probably no tyrosine. Actobindin reduces the concentration of F-actin at steady state and inhibits the rate of filament elongation to extents consistent with the formation of a 1:1 actobindin-G-actin complex in a reaction with a KD of about 5 microM. The available data do not eliminate the possibility of other stoichiometries for the complex, but they are not consistent with any significant interaction between actobindin and F-actin. Despite the similarities between the effects of actobindin and Acanthamoeba profilin on the polymerization of Acanthamoeba actin, the two proteins are quite distinct with different native and subunit molecular weights, different isoelectric points, and different amino acid compositions. Also, unlike profilin, actobindin binds as well to rabbit skeletal muscle G-actin and to pyrenyl-labeled G-actin as it does to unmodified Acanthamoeba G-actin.
Topics: Actins; Amino Acids; Amoeba; Animals; Carrier Proteins; Gelsolin; Kinetics; Microfilament Proteins; Molecular Weight; Protozoan Proteins
PubMed: 3782158
DOI: No ID Found -
The Journal of Biological Chemistry Nov 2010Coronin is a conserved actin-binding protein that co-functions with ADF/cofilin and Arp2/3 complex to govern cellular actin dynamics. Despite emerging roles for coronin...
Coronin is a conserved actin-binding protein that co-functions with ADF/cofilin and Arp2/3 complex to govern cellular actin dynamics. Despite emerging roles for coronin in a range of physiological processes and disease states, a detailed understanding of the molecular interactions of coronin with actin and other binding partners has been lacking. Here, we performed a systematic mutational analysis of surfaces on the yeast coronin β-propeller domain, which binds to F-actin and is conserved in all coronin family members. We generated 21 mutant alleles and analyzed their biochemical effects on actin binding and ADF/cofilin activity. Conserved actin-binding residues mapped to a discrete ridge stretching across one side of the β-propeller. Mutants defective in actin binding showed loss of synergy with ADF/cofilin in severing filaments, diminished localization to actin structures in vivo, and loss of coronin overexpression growth defects. In addition, one allele showed normal actin binding but impaired functional interactions with ADF/cofilin. Another allele showed normal actin binding but failed to cause coronin overexpression defects. Together, these results indicate that actin binding is critical for many of the biochemical and cellular functions of coronin and that the β-propeller domain mediates additional functional interactions with ADF/cofilin and possibly other ligands. Conservation of the actin-binding surfaces across distant species and in all three major classes of coronin isoforms suggests that the nature of the coronin-actin association may be similar in other family members.
Topics: Actins; Alleles; Destrin; Microfilament Proteins; Mutagenesis; Peptide Mapping; Protein Binding; Protein Structure, Secondary; Protein Structure, Tertiary; Saccharomyces cerevisiae
PubMed: 20813846
DOI: 10.1074/jbc.M110.171496