A teamwork promotion of formin-mediated actin nucleation by Bud6 and Aip5 in Saccharomyces cerevisiae

2021 ◽  
Author(s):  
Ying Xie ◽  
Feng Zhou ◽  
Qianqian Ma ◽  
Lanyuan Lu ◽  
Yansong Miao
Keyword(s):  
Actin Polymerization ◽  
Protein Complexes ◽  
Budding Yeast ◽  
Intramolecular Interactions ◽  
Functional Protein ◽  
Actin Nucleation ◽  
Intrinsically Disordered ◽  
Multiple Partners ◽  
Functional Complex ◽  
Actin Nucleator

Actin nucleation is achieved by collaborative teamwork of actin nucleator factors (NFs) and nucleation-promoting factors (NPFs) into functional protein complexes. Selective inter- and intramolecular interactions between the nucleation complex constituents enable diverse modes of complex assembly in initiating actin polymerization upon demand. Budding yeast has two formins, Bni1 and Bnr1, which are teamed up with different NPFs. However, the selective pairing between formin NFs and NPFs into the nucleation core for actin polymerization is not completely understood. By examining the functions and interactions of NPFs and NFs via biochemistry, genetics, and mathematical modeling approaches, we found that two NPFs, Aip5 and Bud6, showed joint teamwork effort with Bni1 and Bnr1, respectively, by interacting with the C-terminal intrinsically disordered region (IDR) of formin, in which two NPFs work together to promote formin-mediated actin nucleation. Although the C-terminal IDRs of Bni1 and Bnr1 are distinct in length, each formin IDR orchestrates the recruitment of Bud6 and Aip5 cooperatively by different positioning strategies to form a functional complex. Our study demonstrated the dynamic assembly of the actin nucleation complex by recruiting multiple partners in budding yeast, which may be a general feature for effective actin nucleation by formins. [Media: see text]

scholarly journals Rickettsia Sca2 Recruits Two Actin Subunits for Nucleation but Lacks WH2 Domains

2018 ◽  
Author(s):  
SS Alqassim ◽  
IG Lee ◽  
R Dominguez
Keyword(s):  
Amino Acid ◽  
Actin Filament ◽  
Actin Polymerization ◽  
Intramolecular Interactions ◽  
Actin Binding ◽  
Globular Domain ◽  
Comet Tail ◽  
Capping Protein ◽  
High Affinity ◽  
Actin Nucleator

AbstractThe Rickettsia ~1,800 amino acid autotransporter protein Sca2 promotes actin polymerization on the surface of the bacterium to drive its movement using an actin comet tail mechanism. Sca2 mimics eukaryotic formins in that it promotes both actin filament nucleation and elongation and competes with capping protein to generate filaments that are long and unbranched. However, despite these functional similarities, Sca2 is structurally unrelated to eukaryotic formins and achieves these functions through an entirely different mechanism. Thus, while formins are dimeric, Sca2 functions as a monomer. However, Sca2 displays intramolecular interactions and functional cooperativity between its N- and C-terminal domains that are crucial for actin nucleation and elongation. Here, we map the interaction of N- and C-terminal fragments of Sca2 and their contributions to actin binding and nucleation. We find that both the N- and C-terminal regions of Sca2 interact with actin monomers, but only weakly, whereas the full-length protein binds two actin monomers with high affinity. Moreover, deletions at both ends of the N- and C-terminal regions disrupt their ability to interact with each other, suggesting that they form a contiguous ring-like structure that wraps around two actin subunits, analogous to the formin homology-2 (FH2) domain. The discovery of Sca2 as an actin nucleator followed the identification of what appeared to be a repeat of three WH2 domains in the middle of the molecule, consistent with the presence of WH2 domains in most actin nucleators. However, we show here that contrary to previous assumptions Sca2 does not contain WH2 domains, and that the corresponding region is folded as a globular domain that cooperates with other parts of the Sca2 molecule for actin binding and nucleation.

scholarly journals Molecular and Biochemical Techniques for Deciphering p53-MDM2 Regulatory Mechanisms

Biomolecules ◽  
2020 ◽  
Vol 11 (1) ◽  
pp. 36
Author(s):  
Konstantinos Karakostis ◽  
Ignacio López ◽  
Ana M. Peña-Balderas ◽  
Robin Fåhareus ◽  
Vanesa Olivares-Illana
Keyword(s):  
Protein Complexes ◽  
Post Translational Modifications ◽  
Intrinsically Disordered ◽  
Advantages And Disadvantages ◽  
Intrinsically Disordered Regions ◽  
Multiple Partners ◽  
Allosteric Interactions ◽  
Spatio Temporal

The p53 and Mouse double minute 2 (MDM2) proteins are hubs in extensive networks of interactions with multiple partners and functions. Intrinsically disordered regions help to adopt function-specific structural conformations in response to ligand binding and post-translational modifications. Different techniques have been used to dissect interactions of the p53-MDM2 pathway, in vitro, in vivo, and in situ each having its own advantages and disadvantages. This review uses the p53-MDM2 to show how different techniques can be employed, illustrating how a combination of in vitro and in vivo techniques is highly recommended to study the spatio-temporal location and dynamics of interactions, and to address their regulation mechanisms and functions. By using well-established techniques in combination with more recent advances, it is possible to rapidly decipher complex mechanisms, such as the p53 regulatory pathway, and to demonstrate how protein and nucleotide ligands in combination with post-translational modifications, result in inter-allosteric and intra-allosteric interactions that govern the activity of the protein complexes and their specific roles in oncogenesis. This promotes elegant therapeutic strategies that exploit protein dynamics to target specific interactions.

scholarly journals Polarisome assembly mediates actin remodeling during polarized yeast and fungal growth

2021 ◽  
Vol 134 (1) ◽  
pp. jcs247916
Author(s):  
Ying Xie ◽  
Yansong Miao
Keyword(s):  
Actin Polymerization ◽  
Budding Yeast ◽  
Fungal Growth ◽  
System Response ◽  
Macromolecular Complex ◽  
Cell Axis ◽  
Intrinsically Disordered ◽  
Intrinsically Disordered Regions ◽  
Dynamic Assembly ◽  
Actin Cables

ABSTRACTDynamic assembly and remodeling of actin is critical for many cellular processes during development and stress adaptation. In filamentous fungi and budding yeast, actin cables align in a polarized manner along the mother-to-daughter cell axis, and are essential for the establishment and maintenance of polarity; moreover, they rapidly remodel in response to environmental cues to achieve an optimal system response. A formin at the tip region within a macromolecular complex, called the polarisome, is responsible for driving actin cable polymerization during polarity establishment. This polarisome undergoes dynamic assembly through spatial and temporally regulated interactions between its components. Understanding this process is important to comprehend the tuneable activities of the formin-centered nucleation core, which are regulated through divergent molecular interactions and assembly modes within the polarisome. In this Review, we focus on how intrinsically disordered regions (IDRs) orchestrate the condensation of the polarisome components and the dynamic assembly of the complex. In addition, we address how these components are dynamically distributed in and out of the assembly zone, thereby regulating polarized growth. We also discuss the potential mechanical feedback mechanisms by which the force-induced actin polymerization at the tip of the budding yeast regulates the assembly and function of the polarisome.

scholarly journals Evolutionary Perspectives on the Moonlighting Functions of Bacterial Factors That Support Actin-Based Motility

mBio ◽  
2019 ◽  
Vol 10 (4) ◽  
Author(s):  
Volkan K. Köseoğlu ◽  
Hervé Agaisse
Keyword(s):  
Cell Adhesion ◽  
Host Cell ◽  
Biofilm Formation ◽  
Actin Polymerization ◽  
Current Knowledge ◽  
Bacterial Pathogens ◽  
Actin Nucleation ◽  
Bacterial Aggregation ◽  
Moonlighting Functions ◽  
Evolutionary Perspectives

ABSTRACT Various bacterial pathogens display an intracellular lifestyle and spread from cell to cell through actin-based motility (ABM). ABM requires actin polymerization at the bacterial pole and is mediated by the expression of bacterial factors that hijack the host cell actin nucleation machinery or exhibit intrinsic actin nucleation properties. It is increasingly recognized that bacterial ABM factors, in addition to having a crucial task during the intracellular phase of infection, display “moonlighting” adhesin functions, such as bacterial aggregation, biofilm formation, and host cell adhesion/invasion. Here, we review our current knowledge of ABM factors and their additional functions, and we propose that intracellular ABM functions have evolved from ancestral, extracellular adhesin functions.

scholarly journals Intrinsically disordered proteins identified in the aggregate proteome serve as biomarkers of neurodegeneration

2021 ◽  
Author(s):  
Srinivas Ayyadevara ◽  
Akshatha Ganne ◽  
Meenakshisundaram Balasubramaniam ◽  
Robert J. Shmookler Reis
Keyword(s):  
Intrinsically Disordered Proteins ◽  
Protein Complexes ◽  
Disordered Proteins ◽  
Intrinsically Disordered ◽  
Intrinsically Disordered Regions ◽  
Physiological Processes ◽  
Protein Partners ◽  
And Function ◽  
Computational Analyses ◽  
Disordered Regions

AbstractA protein’s structure is determined by its amino acid sequence and post-translational modifications, and provides the basis for its physiological functions. Across all organisms, roughly a third of the proteome comprises proteins that contain highly unstructured or intrinsically disordered regions. Proteins comprising or containing extensive unstructured regions are referred to as intrinsically disordered proteins (IDPs). IDPs are believed to participate in complex physiological processes through refolding of IDP regions, dependent on their binding to a diverse array of potential protein partners. They thus play critical roles in the assembly and function of protein complexes. Recent advances in experimental and computational analyses predicted multiple interacting partners for the disordered regions of proteins, implying critical roles in signal transduction and regulation of biological processes. Numerous disordered proteins are sequestered into aggregates in neurodegenerative diseases such as Alzheimer’s disease (AD) where they are enriched even in serum, making them good candidates for serum biomarkers to enable early detection of AD.

scholarly journals EGF stimulates an increase in actin nucleation and filament number at the leading edge of the lamellipod in mammary adenocarcinoma cells

1998 ◽  
Vol 111 (2) ◽  
pp. 199-211 ◽  
Author(s):  
A.Y. Chan ◽  
S. Raft ◽  
M. Bailly ◽  
J.B. Wyckoff ◽  
J.E. Segall ◽  
...  
Keyword(s):  
Actin Polymerization ◽  
De Novo ◽  
Leading Edge ◽  
Mammary Adenocarcinoma ◽  
Actin Dynamics ◽  
Actin Nucleation ◽  
Nucleation Sites ◽  
Nucleation Activity ◽  
Pointed End ◽  
Stimulation Of

Stimulation of metastatic MTLn3 cells with EGF causes the rapid extension of lamellipods, which contain a zone of F-actin at the leading edge. In order to establish the mechanism for accumulation of F-actin at the leading edge and its relationship to lamellipod extension in response to EGF, we have studied the kinetics and location of EGF-induced actin nucleation activity in MTLn3 cells and characterized the actin dynamics at the leading edge by measuring the changes at the pointed and barbed ends of actin filaments upon EGF stimulation of MTLn3 cells. The major result of this study is that stimulation of MTLn3 cells with EGF causes a transient increase in actin nucleation activity resulting from the appearance of free barbed ends very close to the leading edge of extending lamellipods. In addition, cytochalasin D causes a significant decrease in the total F-actin content in EGF-stimulated cells, indicating that both actin polymerization and depolymerization are stimulated by EGF. Pointed end incorporation of rhodamine-labeled actin by the EGF stimulated cells is 2.12+/−0.47 times higher than that of control cells. Since EGF stimulation causes an increase in both barbed and pointed end incorporation of rhodamine-labeled actin in the same location, the EGF-stimulated nucleation sites are more likely due either to severing of pre-existing filaments or de novo nucleation of filaments at the leading edge thereby creating new barbed and pointed ends. The timing and location of EGF-induced actin nucleation activity in MTLn3 cells can account for the observed accumulation of F-actin at the leading edge and demonstrate that this F-actin rich zone is the primary actin polymerization zone after stimulation.

scholarly journals Progressive Phosphorylation Modulates the Self-Association of a Variably Modified Histone H3 Peptide

2021 ◽  
Vol 8 ◽  
Author(s):  
George V. Papamokos ◽  
George Tziatzos ◽  
Dimitrios G. Papageorgiou ◽  
Spyros Georgatos ◽  
Efthimios Kaxiras ◽  
...  
Keyword(s):  
Intrinsically Disordered Proteins ◽  
Histone H3 ◽  
Intramolecular Interactions ◽  
Disordered Proteins ◽  
Post Translational Modifications ◽  
Intrinsically Disordered ◽  
Self Association ◽  
Dynamics Simulations ◽  
Peptide Charge

Protein phosphorylation is a key regulatory mechanism in eukaryotic cells. In the intrinsically disordered histone tails, phosphorylation is often a part of combinatorial post-translational modifications and an integral part of the “histone code” that regulates gene expression. Here, we study the association between two histone H3 tail peptides modified to different degrees, using fully atomistic molecular dynamics simulations. Assuming that the initial conformations are either α-helical or fully extended, we compare the propensity of the two peptides to associate with one another when both are unmodified, one modified and the other unmodified, or both modified. The simulations lead to the identification of distinct inter- and intramolecular interactions in the peptide dimer, highlighting a prominent role of a fine-tuned phosphorylation rheostat in peptide association. Progressive phosphorylation appears to modulate peptide charge, inducing strong and specific intermolecular interactions between the monomers, which do not result in the formation of amorphous or ordered aggregates, as documented by experimental evidence derived from Circular Dichroism and NMR spectroscopy. However, upon complete saturation of positive charges by phosphate groups, this effect is reversed: intramolecular interactions prevail and dimerization of zero-charge peptides is markedly reduced. These findings underscore the role of phosphorylation thresholds in the dynamics of intrinsically disordered proteins. Phosphorylation rheostats might account for the divergent effects of histone modifications on the modulation of chromatin structure.

scholarly journals Systematic analysis of the molecular architecture of endocytosis reveals a nanoscale actin nucleation template that drives efficient vesicle formation

2017 ◽  
Author(s):  
Markus Mund ◽  
Johannes Albertus van der Beek ◽  
Joran Deschamps ◽  
Serge Dmitrieff ◽  
Jooske Louise Monster ◽  
...  
Keyword(s):  
Actin Polymerization ◽  
Design Principle ◽  
Vesicle Formation ◽  
Molecular Architecture ◽  
Actin Nucleation ◽  
Systematic Analysis ◽  
Superresolution Microscopy ◽  
Membrane Invagination ◽  
Wasp Family Proteins ◽  
Endocytic Vesicles

Clathrin-mediated endocytosis is an essential cellular function in all eukaryotes that is driven by a self-assembled macromolecular machine of over 50 different proteins in tens to hundreds of copies. How these proteins are organized to produce endocytic vesicles with high precision and efficiency is not understood. Here, we developed high-throughput superresolution microscopy to reconstruct the nanoscale structural organization of 23 endocytic proteins from over 100,000 endocytic sites in yeast. We found that proteins assemble by radially-ordered recruitment according to function. WASP family proteins form a circular nano-scale template on the membrane to spatially control actin nucleation during vesicle formation. Mathematical modeling of actin polymerization showed that this WASP nano-template creates sufficient force for membrane invagination and substantially increases the efficiency of endocytosis. Such nanoscale pre-patterning of actin nucleation may represent a general design principle for directional force generation in membrane remodeling processes such as during cell migration and division.

scholarly journals Phase transition of RNA−protein complexes into ordered hollow condensates

2020 ◽  
Vol 117 (27) ◽  
pp. 15650-15658 ◽  
Author(s):  
Ibraheem Alshareedah ◽  
Mahdi Muhammad Moosa ◽  
Muralikrishna Raju ◽  
Davit A. Potoyan ◽  
Priya R. Banerjee
Keyword(s):  
Phase Separation ◽  
Protein Complexes ◽  
Intrinsically Disordered Protein ◽  
Liquid Droplets ◽  
Isotropic Liquid ◽  
Size Dependent ◽  
Intrinsically Disordered ◽  
Condensate Phase ◽  
Rna Complexes ◽  
Rna Protein Complexes

Liquid−liquid phase separation of multivalent intrinsically disordered protein−RNA complexes is ubiquitous in both natural and biomimetic systems. So far, isotropic liquid droplets are the most commonly observed topology of RNA−protein condensates in experiments and simulations. Here, by systematically studying the phase behavior of RNA−protein complexes across varied mixture compositions, we report a hollow vesicle-like condensate phase of nucleoprotein assemblies that is distinct from RNA−protein droplets. We show that these vesicular condensates are stable at specific mixture compositions and concentration regimes within the phase diagram and are formed through the phase separation of anisotropic protein−RNA complexes. Similar to membranes composed of amphiphilic lipids, these nucleoprotein−RNA vesicular membranes exhibit local ordering, size-dependent permeability, and selective encapsulation capacity without sacrificing their dynamic formation and dissolution in response to physicochemical stimuli. Our findings suggest that protein−RNA complexes can robustly create lipid-free vesicle-like enclosures by phase separation.

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