Structure and dynamics of single-isoform recombinant Neuronal Human Tubulin

Microtubules are polymers that cycle stochastically between polymerization and depolymerization i.e., they exhibit 'dynamic instability'. This behavior is crucial for cell division, motility and differentiation. While studies in the last decade have made fundamental breakthroughs in our understanding of how cellular effectors modulate microtubule dynamics, analysis of the relationship between tubulin sequence, structure and dynamics has been held back by a lack of dynamics measurements with and structural characterization of homogenous, isotypically pure, engineered tubulin. Here we report for the first time the cryo-EM structure and in vitro dynamics parameters of recombinant isotypically pure human tubulin. α1A/βIII is a purely neuronal tubulin isoform. The 4.2 Å structure of unmodified human α1A/βIII microtubules shows overall similarity to that of heterogeneous brain microtubules, but is distinguished by subtle differences at polymerization interfaces, which are hotspots for sequence divergence between tubulin isoforms. In vitro dynamics assays show that, like mosaic brain microtubules, recombinant homogenous microtubules undergo dynamic instability but they polymerize slower and catastrophe less frequently. Interestingly, we find that epitaxial growth of α1A/βIII microtubules from heterogeneous brain seeds is inefficient, but can be fully rescued by incorporating as little as 5% of brain tubulin into the homogenous α1A/βIII lattice. Our study establishes a system to examine the structure and dynamics of mammalian microtubules with well-defined tubulin species and is a first and necessary step towards uncovering how tubulin genetic and chemical diversity is exploited to modulate intrinsic microtubule dynamics

Population Pharmacokinetics of Telapristone (CDB-4124) and its Active Monodemethylated Metabolite CDB-4453, with a Mixture Model for Total Clearance

Telapristone is a selective progesterone antagonist that is being developed for the long-term treatment of symptoms associated with endometriosis and uterine fibroids. The population pharmacokinetics of telapristone (CDB-4124) and CDB-4453 was investigated using nonlinear mixed-effects modeling. Data from two clinical studies (n = 32) were included in the analysis. A two-compartment (parent) one compartment (metabolite) mixture model (with two populations for apparent clearance) with first-order absorption and elimination adequately described the pharmacokinetics of telapristone and CDB-4453. Telapristone was rapidly absorbed with an absorption rate constant (Ka) of 1.26 h−1. Moderate renal impairment resulted in a 74% decrease in Ka. The population estimates for oral clearance (CL/F) for the two populations were 11.6 and 3.34 L/h, respectively, with 25% of the subjects being allocated to the high-clearance group. Apparent volume of distribution for the central compartment (V2/F) was 37.4 L, apparent inter-compartmental clearance (Q/F) was 21.9 L/h, and apparent peripheral volume of distribution for the parent (V4/F) was 120 L. The ratio of the fraction of telapristone converted to CDB-4453 to the distribution volume of CDB-4453 (Fmetest) was 0.20/L. Apparent volume of distribution of the metabolite compartment (V3/F) was fixed to 1 L and apparent clearance of the metabolite (CLM/F) was 2.43 L/h. A two-compartment parent-metabolite model adequately described the pharmacokinetics of telapristone and CDB-4453. The clearance of telapristone was separated into two populations and could be the result of metabolism via polymorphic CYP3A5

Tubulin isoform composition tunes microtubule dynamics

Microtubules polymerize and depolymerize stochastically, a behavior essential for cell division, motility and differentiation. While many studies advanced our understanding of how microtubule-associated proteins tune microtubule dynamics in trans, we have yet to understand how tubulin genetic diversity regulates microtubule functions. The majority of in vitro dynamics studies are performed with tubulin purified from brain tissue. This preparation is not representative of tubulin found in many cell types. Here we report the 4.2Å cryo-EM structure and in vitro dynamics parameters of α1B/βI+βIVb microtubules assembled from tubulin purified from a human embryonic kidney cell line with isoform composition characteristic of fibroblasts and many immortalized cell lines. We find that these microtubules grow faster and transition to depolymerization less frequently compared to brain microtubules. Cryo-EM reveals that the dynamic ends of α1B/βI+βIVb microtubules are less tapered and that these tubulin heterodimers display lower curvatures. Interestingly, analysis of EB1 distributions at dynamic ends suggests no differences in GTP cap sizes. Lastly, we show that the addition of recombinant α1A/βIII tubulin, a neuronal isotype overexpressed in many tumors, proportionally tunes the dynamics of α1B/βI+βIVb microtubules. Our study is an important step towards understanding how tubulin isoform composition tunes microtubule dynamics

Integrating Satellite Communication, Navigation and Earth Observation to Achieve a Multi-Hazard Platform for Disaster Management: the PHAROS Platform

Satellite-based assets have the potential to provide high value in all phases of disaster management. This statement is true for each of the three planes on their own, i.e. Earth observation, satellite communication and satellite navigation. However, the potential benefits of using satellite-based assets in disaster management can further increase by exploiting the synergies between them. This paper presents a multi-hazard concept currently under development for all phases of disaster management, integrating all planes of satellite-based assets in a modular and efficient manner. The paper describes the design principles, the synergies and the benefits that can be expected from such a system when it is put into operation.