Optimizing microtubule arrangements for rapid cargo capture

Cellular functions such as autophagy, cell signaling and vesicular trafficking involve the retrograde transport of motor-driven cargo along microtubules. Typically, newly formed cargo engages in slow diffusive movement from its point of origin before attaching to a microtubule. In some cell types, cargo destined for delivery to the perinuclear region relies on capture at dynein-enriched loading zones located near microtubule plus-ends. Such systems include extended cell regions of neurites and fungal hyphae, where the efficiency of the initial diffusive loading process depends on the axial distribution of microtubule plus-ends relative to the initial cargo position. We use analytic mean first passage time calculations and numerical simulations to model diffusive capture processes in tubular cells, exploring how the spatial arrangement of microtubule plus-ends affects the efficiency of retrograde cargo transport. Our model delineates the key features of optimal microtubule arrangements that minimize mean cargo capture times. Namely, we show that configurations with a single long microtubule and broad distribution of additional microtubule plus-ends allow for efficient capture in a variety of different scenarios for retrograde transport. Live-cell imaging of microtubule plus-ends in Aspergillus nidulans hyphae indicates that their distributions exhibit these optimal qualitative features. Our results highlight important coupling effects between microtubule length distribution and retrograde cargo transport, providing guiding principles for the spatial arrangement of microtubules within tubular cell regions.

Random sub-diffusion and capture of genes by the nuclear pore reduces dynamics and coordinates interchromosomal movement

Hundreds of genes interact with the yeast nuclear pore complex (NPC), localizing at the nuclear periphery and clustering with co-regulated genes. Dynamic tracking of peripheral genes shows that they cycle on and off the NPC and that interaction with the NPC slows their sub-diffusive movement. Furthermore, NPC-dependent inter-chromosomal clustering leads to coordinated movement of pairs of loci separated by hundreds of nanometers. We developed Fractional Brownian Motion simulations for chromosomal loci in the nucleoplasm and interacting with NPCs. These simulations predict the rate and nature of random sub-diffusion during repositioning from nucleoplasm to periphery and match measurements from two different experimental models, arguing that recruitment to the nuclear periphery is due to random sub-diffusion and transient capture by NPCs. Finally, the simulations do not lead to inter-chromosomal clustering or coordinated movement, suggesting that interaction with the NPC is necessary, but not sufficient, to cause clustering.

Random sub-diffusion and capture of genes by the nuclear pore reduces dynamics and coordinates interchromosomal movement

ABSTRACTHundreds of genes interact with the yeast nuclear pore complex (NPC), localizing at the nuclear periphery and clustering with co-regulated genes. Dynamic tracking of peripheral genes shows that they cycle on and off the NPC and that interaction with the NPC slows their sub-diffusive movement. Furthermore, NPC-dependent inter-chromosomal clustering leads to coordinated movement of pairs of loci separated by hundreds of nanometers. We developed Fractional Brownian Motion simulations for chromosomal loci in the nucleoplasm and interacting with NPCs. These simulations predict the rate and nature of random sub-diffusion during repositioning from nucleoplasm to periphery and match measurements from two different experimental models, arguing that recruitment to the nuclear periphery is due to random subdiffusion, collision, and capture by NPCs. Finally, the simulations do not lead to inter-chromosomal clustering or coordinated movement, suggesting that interaction with the NPC is necessary, but not sufficient, to cause clustering.

Short-Term Effect of the Addition of Rice Husk Gasification Slag on the Movement and Transformation of Phosphorus in Different Soil Types

Rice husk gasification slag (RS) is a type of biochar that is one of the main by-products generated from the production of biomass power with rice husk as the feed. This study aimed to explore the short-term effect of the application of RS on the movement and transformation of fertilizer P in two different soil types through an incubation experiment. The results showed that the RS addition had a significant influence on the diffusive movement of P in soil microsites close to fertilizer placements both in latosolic red soil and fluvo-aquic soil. After 50 d of incubation, most of the WE-P (water-extractable P), AE-P (acid-extractable P), and Olsen-P (available P) were concentrated within 0–5 mm from the fertilization site. WE-P, Olsen-P, and the movement amount of the P in the 0–5 mm soil section were significantly increased at all levels of the RS application in the fertilizer P both in the two soil types. The application of the RS reduced the sorption and precipitation of the fertilizer P in the soil and improved the efficiency of the fertilizer P. The findings presented in this study may be used as references in developing RS applications that reduce losses of fertilizer P and reduce environmental risks.

Diffusive Transport in the Vitreous Humor: Experimental and Analytical Studies

In relation to intravitreal drug delivery, predictive mathematical models for drug transport are being developed, and to effectively implement these for retinal delivery, the information on biophysical properties of various ocular tissues is fundamentally important. It is therefore necessary to accurately measure the diffusion coefficient of drugs and drug surrogates in the vitreous humor. In this review, we present the studies conducted by various researchers on such measurements over the last several decades. These include imaging techniques (fluorescence and magnetic resonance imaging (MRI)) that make use of introducing a contrast agent or a labeled drug into the vitreous and tracking its diffusive movement at various time points. A predictive model for the same initial conditions when matched with the experimental measurements provides the diffusion coefficient, leading to results for various molecules ranging in size from approximately 0.1 to 160 kDa. For real drugs, the effectiveness of this system depends on the successful labeling of the drugs with suitable contrast agents such as fluorescein and gadolinium or manganese so that fluorescence or MR imagining could be conducted. Besides this technique, some work has been carried out using the diffusion apparatus for measuring permeation of a drug across an excised vitreous body from a donor chamber to the receptor by sampling assays from the chambers at various time intervals. This has the advantage of not requiring labeling but is otherwise more disruptive to the vitreous. Some success with nanoparticles has been achieved using dynamic light scattering (DLS), and presently, radioactive labeling is being explored.

Biometric Image Analysis for Quantitation of Dividing Platelets

(1) Background: Quantification of platelet division is challenging because automated Coulter cell counters produce equivocal platelet counts. (2) Methods: We applied the flow cytometric cell tracking dye dilution assay as a popular immunological method to evaluate lymphocyte proliferation to prove and quantitate platelet division. We also devised a method relying on platelet culture in a semisolid medium which enabled dividing platelets to be identified by limiting the diffusive movement of platelets. Mixing platelets of different labeling colors in semisolid medium and counting the platelet doublets of each color combination enabled us to prove and quantitate platelet division. (3) Results: The tracking dye dilution assay revealed that 75.5 to 85.6% of platelets were dividing after 20 hours in culture. Platelets labeled with two different tracking dyes were mixed and cultured in semisolid medium for differential doublet counting. We counted platelet singlets and doublets of each color and color combination using confocal microscopy after six hours of culture and compared the relative number of two-colored doublets with binomial prediction to prove platelet division (P < 0.01). Division was suppressed by taxol, nocodazole, or cytochalasin D treatment. We derived a formula for determining the fraction of dividing platelets using the numbers of singlets and doublets of each color and color combination. The platelet division fraction ranged from 8.8 to 17.5%. (4) Conclusion: We successfully measured platelet division using a simple biometric image analysis method with possible future application to microfluidic devices.

Shear effects on the diffusive movement of oil in triacylglycerol networks

Correlation between the macroscopic Deff obtained through the Ziegleder model and Dmol obtained by NMR in fat crystal networks.

Single-molecule imaging of Hedgehog pathway protein Smoothened in primary cilia reveals binding events regulated by Patched1

Accumulation of the signaling protein Smoothened (Smo) in the membrane of primary cilia is an essential step in Hedgehog (Hh) signal transduction, yet the molecular mechanisms of Smo movement and localization are poorly understood. Using ultrasensitive single-molecule tracking with high spatial/temporal precision (30 nm/10 ms), we discovered that binding events disrupt the primarily diffusive movement of Smo in cilia at an array of sites near the base. The affinity of Smo for these binding sites was modulated by the Hh pathway activation state. Activation, by either a ligand or genetic loss of the negatively acting Hh receptor Patched-1 (Ptch), reduced the affinity and frequency of Smo binding at the base. Our findings quantify activation-dependent changes in Smo dynamics in cilia and highlight a previously unknown step in Hh pathway activation.