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The Journal of Cell Biology Feb 2023The lipid phosphatidyl-D-myo-inositol-4,5-bisphosphate [PI(4,5)P2] is a master regulator of plasma membrane (PM) function. Its effector proteins regulate transport,...
The lipid phosphatidyl-D-myo-inositol-4,5-bisphosphate [PI(4,5)P2] is a master regulator of plasma membrane (PM) function. Its effector proteins regulate transport, signaling, and cytoskeletal processes that define PM structure and function. How a single type of lipid regulates so many parallel processes is unclear. We tested the hypothesis that spatially separate PI(4,5)P2 pools associate with different PM complexes. The mobility of PI(4,5)P2 was measured using biosensors by single-particle tracking. We found that PM lipids including PI(4,5)P2 diffuse rapidly (∼0.3 µm2/s) with Brownian motion, although they spend one third of their time diffusing more slowly. Surprisingly, areas of the PM occupied by PI(4,5)P2-dependent complexes did not slow PI(4,5)P2 lateral mobility. Only the spectrin and septin cytoskeletons showed reduced PI(4,5)P2 diffusion. We conclude that even structures with high densities of PI(4,5)P2 effector proteins, such as clathrin-coated pits and focal adhesions, do not corral unbound PI(4,5)P2, questioning a role for spatially segregated PI(4,5)P2 pools in organizing and regulating PM functions.
Topics: Actin Cytoskeleton; Cell Membrane; Diffusion; Membrane Lipids; Spectrin; Phosphatidylinositols
PubMed: 36416724
DOI: 10.1083/jcb.202204099 -
Biophysical Journal Jun 2023Previous studies have documented the formation of a heterodimer between the two protein kinases PDK1 and PKCα on a lipid bilayer containing their target lipids. This...
Previous studies have documented the formation of a heterodimer between the two protein kinases PDK1 and PKCα on a lipid bilayer containing their target lipids. This work investigates the association-dissociation kinetics of this PDK1:PKCα heterodimer. The approach monitors the two-dimensional diffusion of single, membrane-associated PDK1 molecules for diffusivity changes as PKCα molecules bind and unbind. In the absence of PKCα, a membrane-associated PDK1 molecule exhibits high diffusivity (or large diffusion constant, D) because its membrane-contacting PH domain binds the target PIP lipid headgroup with little bilayer penetration, yielding minimal frictional drag against the bilayer. In contrast, membrane-associated PKCα contacts the bilayer via its C1A, C1B, and C2 domains, which each bind at least one target lipid with significant bilayer insertion, yielding a large frictional drag and low diffusivity. The present findings reveal that individual fluor-PDK1 molecules freely diffusing on the membrane surface undergo reversible switching between distinct high and low diffusivity states, corresponding to the PDK1 monomer and the PDK1:PKCα heterodimer, respectively. The observed single-molecule diffusion trajectories are converted to step length time courses, then subjected to two-state, hidden Markov modeling and dwell time analysis. The findings reveal that both the PDK1 monomer state and the PDK1:PKCα heterodimer state decay via simple exponential kinetics, yielding estimates of rate constants for state switching in both directions. Notably, the PDK1:PKCα heterodimer has been shown to competitively inhibit PDK1 phosphoactivation of AKT1, and is believed to play a tumor suppressor role by limiting excess activation of the highly oncogenic PDK1/AKT1/mTOR pathway. Thus, the present elucidation of the PDK1:PKCα association-dissociation kinetics has important biological and medical implications. More broadly, the findings illustrate the power of single-molecule diffusion measurements to reveal the kinetics of association-dissociation events in membrane signaling reactions that yield a large change in diffusive mobility.
Topics: Protein Kinase C-alpha; Lipid Bilayers; Signal Transduction; Protein Binding; Diffusion
PubMed: 36733254
DOI: 10.1016/j.bpj.2023.01.041 -
Biotechnology Progress Jan 2022High-performance countercurrent membrane purification (HPCMP) has recently been presented as a new approach for protein separations, exploiting differences in diffusive...
High-performance countercurrent membrane purification (HPCMP) has recently been presented as a new approach for protein separations, exploiting differences in diffusive transport across a semipermeable membrane to achieve high selectivity for protein separations. This study presents a set of design equations and diagrams that describe the tradeoff between the yield and purification factor in HPCMP processes in terms of two parametric variables: the diffusive membrane selectivity and the ratio of the draw to bulk solution flow rates. Conditions are identified that provide the high yields and purification factors of interest in bioprocessing. In addition, hydrodynamic models for solute transport were used to evaluate the selectivity as a function of the membrane pore size distribution for purely size-based separations. Model calculations demonstrate that diffusive transport provides significantly greater selectivity than traditional pressure-driven membrane separations for the same pore size distribution due to differences in hindered transport rates for diffusion and convection. These results provide a framework that can be used for the development of HPCMP processes for highly selective protein separations.
Topics: Countercurrent Distribution; Diffusion; Membranes; Proteins
PubMed: 34716693
DOI: 10.1002/btpr.3221 -
Science (New York, N.Y.) Dec 1969The diffusivities of ionic potassium, sodium, sulfate, and adenosine triphosphate inside a nmuscle cell are reduced by a factor of 2, relative to diffusivities in...
The diffusivities of ionic potassium, sodium, sulfate, and adenosine triphosphate inside a nmuscle cell are reduced by a factor of 2, relative to diffusivities in aqueous solution. The diffusion coefficients of nonelectrolytes are reduced by the same factor, showing that the diffusion of the ions is retarded by physical, rather than chemical, interactions. In contrast, the diffusivity of the calcium ion, which is taken up by the sarcoplasmic reticulum, is reduced fiftyfold.
Topics: Adenosine Triphosphate; Animals; Anura; Cytoplasm; Diffusion; In Vitro Techniques; Muscles; Myofibrils; Potassium; Sodium; Sorbitol; Sucrose; Sulfates; Water
PubMed: 5350329
DOI: 10.1126/science.166.3910.1297 -
Biophysical Journal Feb 2021Diffusion is a fundamental mechanism for protein distribution in cell membranes. These membranes often exhibit complex shapes, which range from shallow domes to...
Diffusion is a fundamental mechanism for protein distribution in cell membranes. These membranes often exhibit complex shapes, which range from shallow domes to elongated tubular or pearl-like structures. Shape complexity of the membrane influences the diffusive spreading of proteins and molecules. Despite the importance membrane geometry plays in these diffusive processes, it is challenging to establish the dependence between diffusion and membrane morphology. We solve the diffusion equation numerically on various static curved shapes representative for experimentally observed membrane shapes. Our results show that membrane necks become diffusion barriers. We determine the diffusive half-time, i.e., the time that is required to reduce the amount of protein in the budded region by one half, and find a quadratic relation between the diffusive half-time and the averaged mean curvature of the membrane shape, which we rationalize by a scaling law. Our findings thus help estimate the characteristic diffusive timescale based on the simple measure of membrane mean curvature.
Topics: Cell Membrane; Diffusion; Membranes; Proteins
PubMed: 33359464
DOI: 10.1016/j.bpj.2020.12.014 -
Analytical Chemistry Mar 2022The lateral diffusion of transmembrane proteins in cell membranes is an important process that controls the dynamics and functions of the cell membrane. Several...
The lateral diffusion of transmembrane proteins in cell membranes is an important process that controls the dynamics and functions of the cell membrane. Several fluorescence-based techniques have been developed to study the diffusivities of transmembrane proteins. However, it is challenging to measure the diffusivity of a transmembrane protein with slow diffusion because of the photobleaching effect caused by long exposure times or multiple exposures to light. In this study, we developed a cell membrane electrophoresis platform to measure diffusivity. We deposited cell membrane vesicles derived from HeLa cells to form supported cell membrane patches. We demonstrated that the electrophoresis platform can be used to drive the movement of not only a lipid probe but also a native transmembrane protein, GLUT1. The movements were halted by the boundaries of the membrane patches and the concentration profiles reached steady states when the diffusion mass flux was balanced with the electrical mass flux. We used the Nernst-Planck equation as the mass balance equation to describe the steady concentration profiles and fitted these equations to our data to obtain the diffusivities. The obtained diffusivities were comparable to those obtained by fluorescence recovery after photobleaching, suggesting the validity of this new method of diffusivity measurement. Only a single snapshot is required for the diffusivity measurement, addressing the problems associated with photobleaching and allowing researchers to measure the diffusivity of transmembrane proteins with slow diffusion.
Topics: Cell Membrane; Diffusion; Electrophoresis; HeLa Cells; Humans; Membrane Proteins
PubMed: 35230091
DOI: 10.1021/acs.analchem.2c00211 -
Journal of Mathematical Biology Oct 2022In this paper, we investigate the maximization of the total population of a single species which is governed by a stationary diffusive logistic equation with a fixed...
In this paper, we investigate the maximization of the total population of a single species which is governed by a stationary diffusive logistic equation with a fixed amount of resources. For large diffusivity, qualitative properties of the maximizers like symmetry will be addressed. Our results are in line with previous findings which assert that for large diffusion, concentrated resources are favorable for maximizing the total population. Then, an optimality condition for the maximizer is derived based upon rearrangement theory. We develop an efficient numerical algorithm applicable to domains with different geometries in order to compute the maximizer. It is established that the algorithm is convergent. Our numerical simulations give a real insight into the qualitative properties of the maximizer and also lead us to some conjectures about the maximizer.
Topics: Algorithms; Diffusion; Logistic Models
PubMed: 36207613
DOI: 10.1007/s00285-022-01817-0 -
Computers in Biology and Medicine Sep 2023Reliable skin cancer diagnosis models play an essential role in early screening and medical intervention. Prevailing computer-aided skin cancer classification systems...
Reliable skin cancer diagnosis models play an essential role in early screening and medical intervention. Prevailing computer-aided skin cancer classification systems employ deep learning approaches. However, recent studies reveal their extreme vulnerability to adversarial attacks - often imperceptible perturbations to significantly reduce the performances of skin cancer diagnosis models. To mitigate these threats, this work presents a simple, effective, and resource-efficient defense framework by reverse engineering adversarial perturbations in skin cancer images. Specifically, a multiscale image pyramid is first established to better preserve discriminative structures in the medical imaging domain. To neutralize adversarial effects, skin images at different scales are then progressively diffused by injecting isotropic Gaussian noises to move the adversarial examples to the clean image manifold. Crucially, to further reverse adversarial noises and suppress redundant injected noises, a novel multiscale denoising mechanism is carefully designed that aggregates image information from neighboring scales. We evaluated the defensive effectiveness of our method on ISIC 2019, a largest skin cancer multiclass classification dataset. Experimental results demonstrate that the proposed method can successfully reverse adversarial perturbations from different attacks and significantly outperform some state-of-the-art methods in defending skin cancer diagnosis models.
Topics: Humans; Skin Neoplasms; Skin; Diffusion; Normal Distribution
PubMed: 37572441
DOI: 10.1016/j.compbiomed.2023.107310 -
Physical Review Letters Aug 2015Using random walk simulations we explore diffusive transport through monodisperse sphere packings over a range of packing fractions ϕ in the vicinity of the jamming...
Using random walk simulations we explore diffusive transport through monodisperse sphere packings over a range of packing fractions ϕ in the vicinity of the jamming transition at ϕ(c). Various diffusion properties are computed over several orders of magnitude in both time and packing pressure. Two well-separated regimes of normal "Fickian" diffusion, where the mean squared displacement is linear in time, are observed. The first corresponds to diffusion inside individual spheres, while the latter is the long-time bulk diffusion. The intermediate anomalous diffusion regime and the long-time value of the diffusion coefficient are both shown to be controlled by particle contacts, which in turn depend on proximity to ϕ(c). The time required to recover normal diffusion t* scales as (ϕ-ϕ(c))(-0.5) and the long-time diffusivity D(∞)∼(ϕ-ϕ(c))0.5, or D(∞)∼1/t*. It is shown that the distribution of mean first passage times associated with the escape of random walkers between neighboring particles controls both t* and D(∞) in the limit ϕ→ϕ(c).
Topics: Algorithms; Biological Transport; Diffusion; Models, Biological; Models, Theoretical; Stochastic Processes
PubMed: 26340211
DOI: 10.1103/PhysRevLett.115.088002 -
Physical Review Letters Oct 2022We extend the notions of multipole and subsystem symmetries to more general spatially modulated symmetries. We uncover two instances with exponential and (quasi)periodic...
We extend the notions of multipole and subsystem symmetries to more general spatially modulated symmetries. We uncover two instances with exponential and (quasi)periodic modulations and provide simple microscopic models in one, two, and three dimensions. Seeking to understand their effect on the long-time dynamics, we numerically study a stochastic cellular automaton evolution that obeys such symmetries. We prove that, in one dimension, the periodically modulated symmetries lead to a diffusive scaling of correlations modulated by a finite microscopic momentum. In higher dimensions, these symmetries take the form of lines and surfaces of conserved momenta. These give rise to exotic forms of subdiffusive behavior with a rich spatial structure influenced by lattice-scale features. Exponential modulation, on the other hand, can lead to correlations that are infinitely long-lived at the boundary while decaying exponentially in the bulk.
Topics: Diffusion
PubMed: 36332248
DOI: 10.1103/PhysRevLett.129.170601