189 research outputs found
Antenna mechanism of length control of actin cables
Actin cables are linear cytoskeletal structures that serve as tracks for
myosin-based intracellular transport of vesicles and organelles in both yeast
and mammalian cells. In a yeast cell undergoing budding, cables are in constant
dynamic turnover yet some cables grow from the bud neck toward the back of the
mother cell until their length roughly equals the diameter of the mother cell.
This raises the question: how is the length of these cables controlled? Here we
describe a novel molecular mechanism for cable length control inspired by
recent experimental observations in cells. This antenna mechanism involves
three key proteins: formins, which polymerize actin, Smy1 proteins, which bind
formins and inhibit actin polymerization, and myosin motors, which deliver Smy1
to formins, leading to a length-dependent actin polymerization rate. We compute
the probability distribution of cable lengths as a function of several
experimentally tuneable parameters such as the formin-binding affinity of Smy1
and the concentration of myosin motors delivering Smy1. These results provide
testable predictions of the antenna mechanism of actin-cable length control
Design Principles of Length Control of Cytoskeletal Structures
Cells contain elaborate and interconnected networks of protein polymers which
make up the cytoskeleton. The cytoskeleton governs the internal positioning and
movement of vesicles and organelles, and controls dynamic changes in cell
polarity, shape and movement. Many of these processes require tight control of
the size and shape of cytoskeletal structures, which is achieved despite rapid
turnover of their molecular components. Here we review mechanisms by which
cells control the size of filamentous cytoskeletal structures from the point of
view of simple quantitative models that take into account stochastic dynamics
of their assembly and disassembly. Significantly, these models make
experimentally testable predictions that distinguish different mechanisms of
length-control. While the primary focus of this review is on cytoskeletal
structures, we believe that the broader principles and mechanisms discussed
herein will apply to a range of other subcellular structures whose sizes are
tightly controlled and are linked to their functions.Comment: 61 pages, 11 figure
A novel mode of capping protein-regulation by Twinfilin
Cellular actin assembly is controlled at the barbed ends of actin filaments, where capping protein (CP) limits polymerization. Twinfilin is a conserved in vivo binding partner of CP, yet the significance of this interaction has remained a mystery. Here, we discover that the C-terminal tail of Twinfilin harbors a CP-interacting (CPI) motif, identifying it as a novel CPI-motif protein. Twinfilin and the CPI-motif protein CARMIL have overlapping binding sites on CP. Further, Twinfilin binds competitively with CARMIL to CP, protecting CP from barbed-end displacement by CARMIL. Twinfilin also accelerates dissociation of the CP inhibitor V-1, restoring CP to an active capping state. Knockdowns of Twinfilin and CP each cause similar defects in cell morphology, and elevated Twinfilin expression rescues defects caused by CARMIL hyperactivity. Together, these observations define Twinfilin as the first \u27pro-capping\u27 ligand of CP and lead us to propose important revisions to our understanding of the CP regulatory cycle
GMF Severs Actin-Arp2/3 Complex Branch Junctions by a Cofilin-like Mechanism
SummaryBackgroundBranched actin filament networks driving cell motility, endocytosis, and intracellular transport are assembled in seconds by the Arp2/3 complex and must be equally rapidly debranched and turned over. One of the only factors known to promote debranching of actin networks is the yeast homolog of glia maturation factor (GMF), which is structurally related to the actin filament-severing protein cofilin. However, the identity of the molecular mechanism underlying debranching and whether this activity extends to mammalian GMF have remained open questions.ResultsUsing scanning mutagenesis and total internal reflection fluorescence microscopy, we show that GMF depends on two separate surfaces for debranching. One is analogous to the G-actin and F-actin binding site on cofilin, but we show using fluorescence anisotropy and chemical crosslinking that it instead interacts with actin-related proteins in the Arp2/3 complex. The other is analogous to a second F-actin binding site on cofilin, which in GMF appears to contact the first actin subunit in the daughter filament. We further show that GMF binds to the Arp2/3 complex with low nanomolar affinity and promotes the open conformation. Finally, we show that this debranching activity and mechanism are conserved for mammalian GMF.ConclusionsGMF debranches filaments by a mechanism related to cofilin-mediated severing, but in which GMF has evolved to target molecular junctions between actin-related proteins in the Arp2/3 complex and actin subunits in the daughter filament of the branch. This activity and mechanism are conserved in GMF homologs from evolutionarily distant species
Scaling behaviour in steady-state contracting actomyosin networks
Contractile actomyosin network flows are crucial for many cellular processes including cell division and motility, morphogenesis and transport. How local remodelling of actin architecture tunes stress production and dissipation and regulates large-scale network flows remains poorly understood. Here, we generate contracting actomyosin networks with rapid turnover in vitro, by encapsulating cytoplasmic Xenopus egg extracts into cell-sized ‘water-in-oil’ droplets. Within minutes, the networks reach a dynamic steady-state with continuous inward flow. The networks exhibit homogeneous, density-independent contraction for a wide range of physiological conditions, implying that the myosin-generated stress driving contraction and the effective network viscosity have similar density dependence. We further find that the contraction rate is roughly proportional to the network turnover rate, but this relation breaks down in the presence of excessive crosslinking or branching. Our findings suggest that cells use diverse biochemical mechanisms to generate robust, yet tunable, actin flows by regulating two parameters: turnover rate and network geometry
Centering and symmetry breaking in confined contracting actomyosin networks
Centering and decentering of cellular components is essential for internal
organization of cells and their ability to perform basic cellular functions
such as division and motility. How cells achieve proper localization of their
components is still not well-understood, especially in large cells such as
oocytes. Here, we study actin-based positioning mechanisms in artificial cells
with persistently contracting actomyosin networks, generated by encapsulating
cytoplasmic Xenopus egg extracts into cell-sized water-in-oil droplets. We
observe size-dependent localization of the contraction center, with a symmetric
configuration in larger cells and a polar one in smaller cells. In the
symmetric state, the contraction center is actively centered, via a
hydrodynamic mechanism based on Darcy friction between the contracting network
and the surrounding cytoplasm. During symmetry breaking, transient attachments
to the cell boundary drive the contraction center to a polar location near the
droplet boundary. Our findings demonstrate a robust, yet tunable, mechanism for
subcellular localization
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