The 3D architecture and molecular foundations of de novo centriole assembly via bicentrioles.

Centrioles are structurally conserved organelles, composing both centrosomes and cilia. In animal cycling cells, centrioles often form through a highly characterized process termed canonical duplication. However, a large diversity of eukaryotes assemble centrioles de novo through uncharacterized pathways. This unexplored diversity is key to understanding centriole assembly mechanisms and how they evolved to assist specific cellular functions. Here, we show that, during spermatogenesis of the bryophyte Physcomitrium patens, centrioles are born as a co-axially oriented centriole pair united by a cartwheel. Interestingly, we observe that these centrioles are twisted in opposite orientations. Microtubules emanate from the bicentrioles, which localize to the spindle poles during cell division. After their separation, the two resulting sister centrioles mature asymmetrically, elongating specific microtubule triplets and a naked cartwheel. Subsequently, two motile cilia are assembled that appear to alternate between different motility patterns. We further show that centriolar components SAS6, Bld10, and POC1, which are conserved across eukaryotes, are expressed during spermatogenesis and required for this de novo biogenesis pathway. Our work supports a scenario where centriole biogenesis, while driven by conserved molecular modules, is more diverse than previously thought.

Plk4 triggers autonomous de novo centriole biogenesis and maturation.

Centrioles form centrosomes and cilia. In most proliferating cells, centrioles assemble through canonical duplication, which is spatially, temporally, and numerically regulated by the cell cycle and the presence of mature centrioles. However, in certain cell types, centrioles assemble de novo, yet by poorly understood mechanisms. Herein, we established a controlled system to investigate de novo centriole biogenesis, using Drosophila melanogaster egg explants overexpressing Polo-like kinase 4 (Plk4), a trigger for centriole biogenesis. We show that at a high Plk4 concentration, centrioles form de novo, mature, and duplicate, independently of cell cycle progression and of the presence of other centrioles. Plk4 concentration determines the temporal onset of centriole assembly. Moreover, our results suggest that distinct biochemical kinetics regulate de novo and canonical biogenesis. Finally, we investigated which other factors modulate de novo centriole assembly and found that proteins of the pericentriolar material (PCM), and in particular γ-tubulin, promote biogenesis, likely by locally concentrating critical components.

BiRep: A Reputation Scheme to Mitigate the Effects of Black-Hole Nodes in Delay-Tolerant Internet of Vehicles.

Delay-tolerant networking (DTN) enables communication in disruptive scenarios where issues such as sparse and intermittent connectivity, long and variable delays, high latency, high error rates, or no end-to-end connectivity exist. Internet of Vehicles (IoV) is a network of the future in which integration between devices, vehicles, and users will be unlimited and universal, overcoming the heterogeneity of systems, services, applications, and devices. Delay-tolerant internet of vehicles (DT-IoV) is emerging and becoming a popular research topic due to the critical applications that can be realized, such as software or map update dissemination. For an IoV to work efficiently, a degree of cooperation between nodes is necessary to deliver messages to their destinations. However, nodes might misbehave and silently drop messages, also known as a black-hole attack, degrading network performance. Various solutions have been proposed to deal with black-hole nodes, but most are centralized or require each node to meet every other node. This paper proposes a decentralized reputation scheme called BiRep that identifies and punishes black-hole nodes in DT-IoV. BiRep is tested on the Prophet routing protocol. Simulation results show excellent performance in all scenarios, comparable or better to other reputation schemes, significantly increasing the delivery ratio of messages.

Evolution of centriole assembly.

In this Primer, Nabais et al. discuss the evolution of the structure and function of centrioles and basal bodies, describe conserved centriole assembly features and the diversity in centriole architecture across eukaryotes, and highlight important outstanding evolutionary questions concerning centriole assembly.

A cell-free system of Drosophila egg explants supporting native mitotic cycles.

Mitosis, in a broader sense, is an intracellular mechanical process that is fueled by chemical reactions and regulated by a complex protein interaction network. Research aimed at understanding mitosis in all these aspects is often limited to pharmaceutical treatment or genetic manipulation of single cells or entire tissues. These experimental models entail physical boundaries imposed by the cell membrane, making it extremely challenging to apply mechanical perturbations, or to introduce larger molecules such as peptides, proteins, or genetic transcripts in an acute and specific manner. Here, we present a cell-free experimental assay that is exploiting the properties of a large, multinucleated embryo cell. Drosophila, like almost all insects, initially develops as a syncytial embryo, the task of which is to replicate and distribute the genetic material quickly and regularly. We describe an experimental procedure that allows the isolation of nucleocytoplasm from single embryos that retains the developmental processes, most importantly the native mitotic progression of nuclei.

Noncanonical Biogenesis of Centrioles and Basal Bodies.

Centrioles and basal bodies (CBBs) organize centrosomes and cilia within eukaryotic cells. These organelles are composed of microtubules and hundreds of proteins performing multiple functions such as signaling, cytoskeleton remodeling, and cell motility. The CBB is present in all branches of the eukaryotic tree of life and, despite its ultrastructural and protein conservation, there is diversity in its function, occurrence (i.e., presence/absence), and modes of biogenesis across species. In this review, we provide an overview of the multiple pathways through which CBBs are formed in nature, with a special focus on the less studied, noncanonical ways. Despite the differences among each mechanism herein presented, we highlighted some of their common principles. These principles, governing different steps of biogenesis, ensure that CBBs may perform a multitude of functions in a huge diversity of organisms but yet retained their robustness in structure throughout evolution.

CDK1 Prevents Unscheduled PLK4-STIL Complex Assembly in Centriole Biogenesis.

Centrioles are essential for the assembly of both centrosomes and cilia. Centriole biogenesis occurs once and only once per cell cycle and is temporally coordinated with cell-cycle progression, ensuring the formation of the right number of centrioles at the right time. The formation of new daughter centrioles is guided by a pre-existing, mother centriole. The proximity between mother and daughter centrioles was proposed to restrict new centriole formation until they separate beyond a critical distance. Paradoxically, mother and daughter centrioles overcome this distance in early mitosis, at a time when triggers for centriole biogenesis Polo-like kinase 4 (PLK4) and its substrate STIL are abundant. Here we show that in mitosis, the mitotic kinase CDK1-CyclinB binds STIL and prevents formation of the PLK4-STIL complex and STIL phosphorylation by PLK4, thus inhibiting untimely onset of centriole biogenesis. After CDK1-CyclinB inactivation upon mitotic exit, PLK4 can bind and phosphorylate STIL in G1, allowing pro-centriole assembly in the subsequent S phase. Our work shows that complementary mechanisms, such as mother-daughter centriole proximity and CDK1-CyclinB interaction with centriolar components, ensure that centriole biogenesis occurs once and only once per cell cycle, raising parallels to the cell-cycle regulation of DNA replication and centromere formation.

Effect of lithiation potential and cycling on chemical and morphological evolution of Si thin film electrode studied by ToF-SIMS.

Si thin films obtained by plasma enhanced chemical vapor deposition (PECVD) were used to investigate chemical and morphological modifications induced by lithiation potential and cycling. These modifications were thoughtfully analyzed by time-of-flight secondary ion mass spectrometry (ToF-SIMS) depth profiling, which allows to distinguish the surface and bulk processes related to the formation of the solid electrolyte interphase (SEI) layer, and Li-Si alloying, respectively. The main results are a volume expansion/shrinkage and a dynamic behavior of the SEI layer during the single lithiation/delithiation process and multicycling. Trapping of lithium and other ions corresponding to products of electrolyte decomposition are the major reasons of electrode modifications. It is shown that the SEI layer contributes to 60% of the total volume variation of Si electrodes (100 nm). The apparent diffusion coefficient of lithium (DLi) calculated from the Fick's second law directly from Li-ion ToF-SIMS profiles is of the order of ∼5.9 × 10(-15) cm(2).s(-1). This quite low value can be explained by Li trapping in the bulk of electrode material, at the interfaces, continuous growth of the SEI layer and increase of SiO2 quantity. These modifications can result in limitation the ionic transport of Li.

Polo-like kinases: structural variations lead to multiple functions.

Members of the polo-like kinase (PLK) family are crucial regulators of cell cycle progression, centriole duplication, mitosis, cytokinesis and the DNA damage response. PLKs undergo major changes in abundance, activity, localization and structure at different stages of the cell cycle. They interact with other proteins in a tightly controlled spatiotemporal manner as part of a network that coordinates key cell cycle events. Their essential roles are highlighted by the fact that alterations in PLK function are associated with cancers and other diseases. Recent knowledge gained from PLK crystal structures, evolution and interacting molecules offers important insights into the mechanisms that underlie their regulation and activity, and suggests novel functions unrelated to cell cycle control for this family of kinases.

Comparative cytogenetics of two endangered leuciscine fish, Squalius aradensis and S. torgalensis (Teleostei, Cyprinidae), from the Iberian Peninsula.

In this study, the description of the karyotypes of the endangered chubs Squalius aradensis (Coelho, Bogutskaya, Rodrigues and Collares-Pereira, 1998) and Squalius torgalensis (Coelho, Bogutskaya, Rodrigues and Collares-Pereira, 1998) is presented by means of conventional (Giemsa-staining, Chromomycin A3 (CMA3)-fluorescence, Silver-impregnation (Ag-NORs)) and molecular (fluorescence in situ hybridization (FISH) with 18S rDNA probe) protocols. These endemic sister-species have an allopatric but adjacent distribution in the most southwestern part of the Iberian Peninsula. Diploid chromosome number was invariably 2n = 50 and karyotypes of both species were grossly similar, composed of metacentric and submetacentric elements with a reduced number of acrocentric pairs. Sequential staining using FISH with an 18S rDNA probe, CMA3 and Ag-NORs treatments revealed consistent positive signals located at the end of the short arms of a submetacentric chromosome pair, likely homologous in both species. While providing useful cytogenetic comparative data against other members of the genus Squalius Bonaparte, 1837, the work aimed to draw attention towards the conservation of two narrow-range and highly confined fish species.

Diaminoethane adsorption and water substitution on hydrated TiO2: a thermochemical study based on first-principles calculations.

Epoxy-amines are used as structural adhesives deposited on Ti. The amine adhesion to a Ti surface depends highly on the surface state (oxidation, hydroxylation). Amines may adsorb above preadsorbed water molecules or substitute them to bind directly to surface Ti(4+) Lewis acid sites. The adsorption of a model amine molecule, diaminoethane (DAE), on a model surface, hydrated TiO2-anatase (101) surface, is investigated using Density Functional Theory including Dispersive forces (DFT-D) calculations. DAE adsorption and water substitution by DAE are exothermic processes and turn nearly isoenergetic at high coverage with adsorption-substitution energies around -0.3 eV (including dispersion forces and ZPE). Complementary ab initio molecular dynamics studies also suggest that the formation of an amine-water interaction induces water desorption from the surface at room temperature, a preliminary step towards the amine-Ti bond formation. An atomistic thermodynamic approach is developed to evaluate the interfacial free energy balance of both processes (adsorption and substitution). The main contributions to the energetic balance are dispersive interactions between molecules and the surface on the exergonic side, translational and rotational entropic contributions on the endergonic one. The substitution process is stabilized by 0.55 eV versus the adsorption one when free solvation, rotational and vibrational energies are considered. The main contribution to this free energy gain is due to water solvation. The calculations suggest that in toluene solvent with a water concentration of 10(-4) M or less, a full DAE layer replaces a preadsorbed water layer for a threshold concentration of DAE ≥ 0.1 M.