Site 2a Research Articles

Crystal structure and crystal chemistry of the τ-Mg32(Al,Zn)49 solid solution using first-principles calculations and thermodynamic modelling

This paper presents a study of the crystal structure and crystal chemistry of the τ-Mg32(Al,Zn)49 phase. We first performed DFT calculations to resolve conflicting data concerning the occupancy of site 2a of the solid solution crystal structure. Subsequently, 16 ordered configurations derived from the mixing of Al/Zn on sites 24g1, 24g2 and 48h of the solid solution structure, as well as the mixing of Mg/Zn on site 12e1 were generated. We then used DFT calculations to derive the formation enthalpies of all the end-members of the solid solution, their elastic constants by imposing 51 finite deformations for each end-member, and their energies as a function of volume. These calculations were used in a Debye-Wang model (in Slater form) to calculate the heat capacities of the 16 end-members and to obtain in fine their Gibbs energies. At last, these calculations were used to support a thermodynamic model based on the Bragg-Williams approximation, enabling all mixed sites occupancies to be calculated at any temperature and chemical composition. For instance, the predictions we carried out at 633 K and 800 K are in very good agreement with the available measurements. On the basis of these new results, we have determined the chemical ordering of the solid solution over a very wide range of chemical composition and temperature, which has enabled us to propose a new sublattice model for the τ-Mg32(Al,Zn)49 solid solution. This sublattice model is both simpler and more accurate than all the other models used in the literature since it agrees with the crystal structure and the crystal chemistry of the τ-Mg32(Al,Zn)49 phase in a large range of temperature and chemical composition.

Atomic resolution transmission electron microscopy visualisation of channel occupancy in beryl in different crystallographic directions

The causes of colour in beryl have been a research topic for decades. For some varieties, such as emerald (green, coloured by Cr3+ and/or V3+), the main cause of colour is substitutions by metal atoms within the framework. However, the causes for the yellow and blue colours in heliodor, golden beryl and aquamarine are still debated. It is generally agreed that Fe ions are responsible for the colour, but there are differing conclusions about the valence states of these ions, the occupied positions and the colour-inducing processes involved. The colour of aquamarine is commonly attributed to intervalence charge transfer (IVCT) between Fe3+ and Fe2+. Various combinations of sites have been proposed to host the Fe ions engaging in this IVCT. Here we present a new approach to address the topic of colour generation: atomic resolution scanning transmission electron microscopy (STEM). For the first time, atomic resolution images of a beryl (natural aquamarine) are presented in the three crystallographic directions [0001], [1-210] and [1-100]. Ions are clearly resolved in the channels. From the ratio of channel occupation and the correlation of the atoms per formula unit (apfu) calculations we conclude that Fe resides in the framework, not in the channels. The projections in the [1-210] direction directly show that the cavity channel site 2a is occupied, most likely by Cs, in agreement with recent results in the literature.

SMYD2 Imparts Gemcitabine Resistance to Pancreatic Adenocarcinoma Cells by Upregulating EVI2A.

Although gemcitabine (GEM) is the first‑line drug for advanced pancreatic adenocarcinoma (PAAD), the development of GEM resistance severely limits the effectiveness of this chemotherapy. This study investigated the mechanisms of ecotropic viral integration site 2A (EVI2A) for resistance to GEM and immune evasion in PAAD. GEM resistance-related biomarkers were predicted using GEO datasets, and GEM-resistant PAAD cells were generated. EVI2A was found expressed highly in GEM-resistant PAAD cells. Gain-of-function analyses revealed that EVI2A encouraged the proliferation and motility of GEM-resistant cells and prevented apoptosis. In addition, EVI2A reduced T cell effector activation. SMYD2 was overexpressed in GEM-resistant cells, and SMYD2 enhanced H3K36me2 modification of EVI2A, thereby promoting EVI2A expression. SMYD2 reduced the sensitivity of GEM-resistant cells, which was reversed by EVI2A knockdown. SMYD2 increased the amount of M2 macrophages (co-cultured with PAAD cells) and decreased T cell effector activation (co-cultured with macrophage supernatant), and the number of M2 macrophages was decreased and T cell effectors were activated following EVI2A knockdown. Our findings indicate that EVI2A, manipulated by the SMYD2-H3K36me2 epigenetic axis, promoted GEM resistance and M2 macrophage-mediated immune evasion in PAAD. Therefore, EVI2A might represent a therapeutic target for overcoming GEM resistance and immunosuppressive environment in PAAD.

CRYSTAL STRUCTURE OF THE Pr3Ag4xGe1,25-xSe7 (x = 0,10; 0,15)

The samples of composition Pr3Ag4xGe1,25-xSe7 (x = 0,10; 0,15), weighing one gram, obtained by sintering elementary components in vacuumed quartz containers at a maximum temperature of 1150 °C. The diffraction patterns for X‑ray phase analysis were recorded at a DRON4–13 diffractometer for 2Q range of 10–100 ° (CuKα radiation, scan step 0,02 °, 20 s exposure in each point). For data processing and the determination of the crystal structure used WinCSD software package. Visualization of the crystal structure, stacking and coordination polyhedra was performed using VESTA program. The crystal structure of the quaternary compound Pr3Ag0,40Ge1,15Se7 (a = 1,06466(4) nm, с = 0,60552(3) nm, RI = 0,0511, Rp = 0,2455) and Pr3Ag0,60Ge1,10Se7 (a = 1,06556(2) nm, c = 0,60583(2) nm, RI = 0,0618, Rp = 0,2068) was investigated by powder X‑ray method. The structure of the compound belongs to the La3CuSiS7 structural type (space group P63, Pearsoncode hP24, a = 1,028 nm, c = 0,575 nm). In this structure, the Pr atom is located in the 6c site and, together with selenium atoms, form trigonal prisms with two additional atoms [Pr Se14Se23Se3] (CN = 8). The atoms of the statistical mixture M1 (0,15 Ge + 0,40 Ag) and M2 (0,10 Ge + 0,60 Ag) occupying one site 2a are located in triangles [M 3Se2] (CN = 3). The Ge atoms are localized in site 2b and have a tetrahedral surrounding of Selenium atoms [Gе Se13Se3] (CN = 4). Atoms Se1 (site 2b) and Se3 (site 6c) also have a tetrahedral surrounding. Se2 atoms (site 6c) are surrounded by five neighbors. Inter-atomic distances are consistent with the sum of the respective ionic radii. Germanium-containing Pr3Ag0,40Ge1,15Se7 and Pr3Ag0,60Ge1,10Se7 selenides based on praseodymium are promising chalcogenide phases on the basis of which materials for nonlinear optics can be created.

Nutrition Program Fidelity Assessment tool: a framework for optimising implementation in military dining facilities.

The aim was to develop, refine and assess the usefulness of the Go for Green® (G4G) 2.0 Program Fidelity Assessment (PFA) tool. G4G 2.0 is a Department of Defense programme designed to optimise access, availability and knowledge of high-performance nutritious foods in military dining facilities (DFAC). During a multi-site study to evaluate G4G 2.0 on meal quality and diner satisfaction, subject matter experts developed and refined a PFA tool based on eight programme requirements (PR). They identified tasks critical to programme success and corresponding benchmarks, then proposed expansion of several PR and developed a scoring system to assess adherence. Three PFA were conducted (Site 1, Site 2A and Site B). Two DFAC in the USA implementing the G4G 2.0 programme. Military DFAC participating in a G4G 2.0 evaluation study. After G4G 2.0 implementation, Site 1 conducted a PFA and met benchmarks for eight of fifteen sections. At Site 2, a PFA was conducted after G4G 2.0 implementation (Site 2A) and one 3 months later (Site 2B) with twelve of fifteen and ten of fifteen sections meeting benchmarks, respectively. Research highlights the need to maximise implementation quality to ensure interventions are effective, achievable and efficient. Using a PFA tool to objectively assess nutrition interventions can inform programme fidelity, successes and opportunities for improvement. Results identify key areas that require additional training and resources to optimise access to nutrient-dense foods that support nutritional fitness. This feedback is critical for assessing potential programme impact on Service Members.

Unusual Site-Selective Doping in Layered Cathode Strengthens Electrostatic Cohesion of Alkali-Metal Layer for Practicable Sodium-Ion Full Cell.

P2-type Na0.67 Ni0.33 Mn0.67 O2 is a dominant cathode material for sodium-ion batteries due to its high theoretical capacity and energy density. However, charging P2-type Na0.67 Ni0.33 Mn0.67 O2 to voltages higher than 4.2 V (vs. Na+ /Na) can induce detrimental structural transformation and severe capacity fading. Herein, stable cycling and moisture resistancy of Na0.67 Ni0.33 Mn0.67 O2 at 4.35 V (vs. Na+ /Na) are achieved through dual-site doping with Cu ion at transition metal site (2a) and unusual Zn ion at Na site (2d) for the first time. The Cu ion doping in 2a site stabilizes the metal layer, while more importantly, the unusual alkali-metal site doping by Zn ion serves as O2- Zn2+ O2- "pillar" for enhancing electrostatic cohesion between two adjacent transition metal layers, preventing the crack of active material along the a-b-plane and restraining the generation of O2 phase upon deep desodiation. This unique dual-site-doped [Na0.67 Zn0.05 ]Ni0.18 Cu0.1 Mn0.67 O2 cathode exhibits a prominent cyclability with 80.6% capacity retention over 2000 cycles at an ultrahigh rate of 10C, demonstrating its great potential for practical applications. Impressively, the full cell devices with [Na0.67 Zn0.05 ]Ni0.18 Cu0.1 Mn0.67 O2 and commercial hard carbon as cathode and anode, respectively, can deliver a high energy density of 217.9Wh kg-1 and excellent cycle life over 1000 cycles.

CRISPR/Cas9-Mediated Specific Integration of Fat-1 and IGF-1 at the pRosa26 Locus.

Many researchers have focused on knock-in pigs for site-specific integration, but little attention has been given to genetically modified pigs with the targeted integration of multiple recombinant genes. To establish a multigene targeted knock-in editing system, we used the internal ribosome entry site (IRES) and self-cleaving 2A peptide technology to construct a plasmid coexpressing the fatty acid desaturase (Fat-1) and porcine insulin-like growth factor-1 (IGF-1) genes at equal levels. In this study, pigs were genetically modified with multiple genes that were precisely inserted into the pRosa26 locus by using the clustered regularly spaced short palindrome repeat sequence (CRISPR)/CRISPR-related 9 (Cas9) system and somatic cell nuclear transfer technology (SCNT) in combination. Single copies of the Fat-1 and IGF-1 genes were expressed satisfactorily in various tissues of F0-generation pigs. Importantly, gas chromatography analysis revealed a significantly increased n-3 polyunsaturated fatty acid (PUFA) level in these genetically modified pigs, which led to a significant decrease of the n-6 PUFA/n-3 PUFA ratio from 6.982 to 3.122 (*** p < 0.001). In conclusion, the establishment of an editing system for targeted double-gene knock-in in this study provides a reference for the precise integration of multiple foreign genes and lays a foundation for the development of new transgenic pig breeds with multiple excellent phenotypes.

NONCENTROSYMMETRIC CRYSTAL STRUCTURE OF SELENIDES M3Si1.75Se7 (M = 0.5Y+0.5La(Pr))

The article studies the problems of the crystal structure of tetrar chalcogenides M3Si1.75Se7 (M = 0.5Y + 0.5La (Pr)). The compounds were obtained by the method of solid-phase reactions by the interaction of elementary components at a temperature of 1323 K. The diffraction patterns of powder samples were taken at room temperature on DRON-4-13. The problem of these structures is not only the localization of a mixture of atoms of rare earth elements (REE), but also the appearance of silicon atoms in two charge states +2 and +4, which has not been previously studied in scientific papers. This approach allows a deeper study of the crystal structure of the obtained phases for the relationship between the charge of the atom and its coordination environment. According to many studies, the localization of atoms carrying the opposite charge relative to the central coordination atom is determined by the position of the atom in the periodic system and its charge. In most studies, there is a variant of the description of the structure, when the composition includes ions with a certain constant conditional charge + or -. In this paper, we propose a solution of the structure in which Silicon with charge 2+ is localized in a site 2a, and Silicon with charge +4 is localized in a site 2b. In addition, this leads to a different coordination environment of these atoms (C.N. = 6 and 4, respectively) The structure was analyzed by the formation of cation-cation pairs between silicon atoms, as the most electronegative atom. A comparative evaluation with known studies that reflect similar issues and found that such pairs in this structure are impossible due to the significant interatomic distances of Si-Si, which leads to the destruction of such a geometric configuration. Silicon atoms in a tetrahedral environment with selenium atoms have relatively short interatomic Si-Se distances, which indicates their predominantly covalent nature. In an octahedral environment, silicon atoms are much larger, indicating an increase in the ionicity of the bond.

Experimental measurement and Monte Carlo code simulation of the gamma heating at different irradiation sites in a nuclear research reactor

Interaction of radiation with matter results in heat generation and in effect, changes in the irradiation sample’s quality. Knowledge about heat deposition within the irradiation sites can go a long way in enhancing the safety and quality of the irradiation condition. To measure such results, SCK-CEN Gamma Thermometer (GT) is used for experimentally measuring Gamma Heating (GH) during the operation of McMaster Nuclear Reactor (MNR) at three irradiation sites. In addition, Monte Carlo reactor physics code (Serpent-2) is also used for modeling the MNR in order to compare as well as validate it against experimental and operational measured data such as the multiplication factor (keff) during the operational day against the actual values (unity), the experimentally measured GH values. After finding a difference of 0.45% at the beginning of the day and 0.32% at the end of the day in the keff, a comparable GH profile was observed between the measurement and the calculation. Additionally, the average axial differences of GH results found in the beryllium irradiation site (2A), and the graphite irradiation sites (8B and 8E) are 4.71%, 10.71%, and 11.72%.

Production and Application of Multicistronic Constructs for Various Human Disease Therapies.

The development of multicistronic vectors has opened up new opportunities to address the fundamental issues of molecular and cellular biology related to the need for the simultaneous delivery and joint expression of several genes. To date, the examples of the successful use of multicistronic vectors have been described for the development of new methods of treatment of various human diseases, including cardiovascular, oncological, metabolic, autoimmune, and neurodegenerative disorders. The safety and effectiveness of the joint delivery of therapeutic genes in multicistronic vectors based on the internal ribosome entry site (IRES) and self-cleaving 2A peptides have been shown in both in vitro and in vivo experiments as well as in clinical trials. Co-expression of several genes in one vector has also been used to create animal models of various inherited diseases which are caused by mutations in several genes. Multicistronic vectors provide expression of all mutant genes, which allows the most complete mimicking disease pathogenesis. This review comprehensively discusses multicistronic vectors based on IRES nucleotide sequence and self-cleaving 2A peptides, including its features and possible application for the treatment and modeling of various human diseases.

Increased expression of ecotropic viral integration site 2A indicates a poor prognosis and promotes osteosarcoma evolution through activating MEK/ERK pathway

Although ecotropic viral integration site 2 A (EVI2A) plays key roles in several cancers, the expression and function of EVI2A in osteosarcoma (OS) have not been investigated. Hence, we explored the expression of EVI2A and its clinical significance of EVI2A of OS. Firstly, we investigated the expression of EVI2A in OS tissues. The relationship of EVI2A expression and survival time was analyzed using Kaplan–Meier plotter. Then, we used quantitative reverse transcription PCR (qRT-PCR) to confirm the expression level of EVI2A in OS cell lines. Cell proliferation, and wound-healing experiments were used to identify the biological function of EVI2A. Moreover, EVI2A-mediated MEK/ERK signaling pathway was evaluated using western blotting. Data suggested that EVI2A was highly expressed in OS tissues, and high-expression of EVI2A was associated with worse overall survival in OS patients. Moreover, the up-regulation of it was observed in OS cell lines (Saos2, and MG63). Knockdown of EVI2A suppressed cell proliferation and migration of OS. Western blotting revealed that the inactivation of MEK/ERK pathway was found in OS cells after EVI2A knockdown. Our data implicated the crucial role of EVI2A in the progression of OS, demonstrating that expression of EVI2A may offer an attractive novel prognostic signature for OS.

Mössbauer-effect study of dynamic, magnetic, and electronic properties of C14 Laves phase Nb0.975Fe2.025

C14 Laves phase Nb0.975Fe2.025 compound was investigated by means of the Mössbauer spectroscopy. Spectra were recorded in the temperature range of 5–300 K. Their analysis in terms of three sub-spectra yielded information on magnetic and lattice dynamical properties of Fe atoms regularly occupying 2a and 6h lattice sites and, excessively, 4f sites. No indication of magnetism was observed down to the temperature of T ≈ 60 K, and spectral parameters, viz., center shift, CS, and the main component of the electric field gradient, Vzz, behave regularly. In particular, analysis of CS(T) in terms of the Debye model yielded the following values of the Debye temperature, TD: 453(5) K for the site 6h, 544(10) K for the site 2a, 479(4) K for the weighted average over 6h and 2a sites, and 363(35) K for the site 4f. Below ∼60 K, anomalous behavior was observed: a broadening of the spectrum appeared indicating thereby a transition into a magnetic phase. Analysis of a temperature dependence of the hyperfine field, B, associated with the 6h and 2a sub-spectra yielded different values of the magnetic ordering temperature. The maximum values of the hyperfine field at 5 K, Bo, were 8.2 kGs and 3.3 kGs, for 6h and 2a, respectively. The Bo-values were used to estimate values of the underlying magnetic moments, μFe(6h) = 0.055–0.065 μB and μFe(2a) = 0.02–0.025 μB. Noteworthy, they are over one order of magnitude lower than those theoretically calculated. The CS and Vzz parameters showed anomalies in the magnetic phase, in particular the former exhibited a strong departure from the Debye model prediction testifying to a significant effect of magnetism on the lattice vibrations.

Revealing the molecular interactions of aptamers that specifically bind to the extracellular domain of HER2 cancer biomarker protein: An in silico assessment

Single-stranded (ss) oligonucleotide aptamers are emerging as the promising substitutes for monoclonal antibodies because of their low production cost and good batch-to-batch consistency. Aptamers vividly bind to a variety of cellular targets and alter their functions with a remarkable degree of specificities. In this study, the positive clones of human epidermal growth factor receptor 2 (HER2) specific binding ssDNA aptamers which were previously identified by in vitro Systematic Evolution of Ligands by EXponential enrichment (SELEX) process, hitherto lacking the putative binding site information and residues crucial for aptamer recognition are studied. Primarily, four putative DNA binding regions present on the HER2 extracellular domain (ECD) were identified using prediction servers and electrostatic potential maps, which were further exploited to delineate the aptamer binding features. Molecular docking and molecular dynamics (MD) simulations revealed stable binding nature of three aptamers (H2>H1>H6), which chose Site 2a as preferred binding site present on the HER2(ECD) protein. Furthermore, amino acid residues viz. Asn37, Gln59, Arg81-Gln84, Asp88, and Lys128 of Site 2a were found to be crucial for high-affinity binding. This knowledge can be utilized as a benchmark for the future studies, in search for better and highly specific anti-HER2 aptamers as cancer therapeutics or as diagnostic agents.

Mass spectrometry for identification of proteins that specifically bind to a distal enhancer of the Oct4 gene

Transcription factor Oct4 is a marker of pluripotent stem cells and has a significant role in their self-renewal. Oct4 gene is controlled by three cis-regulatory elements – proximal promoter, proximal enhancer and distal enhancer. All of these elements are targets for binding of regulatory proteins. Distal enhancer is in our research focus because of its activity in early stages of embryonic development. There are two main sequences called site 2A and site 2B that are presented in distal enhancer. For this moment proteins which bind to a site 2A (CCCCTCCCCCC) remain unknown. Using combination of in vitro method electrophoretic mobility shift assay (EMSA) and mass spectromery we identified several candidates that can regulate Oct4 gene expression through site 2A.

Specific heat of hexagonal Yb1−xSrxMnO3

We have studied influence of the strontium doping in polycrystalline YbMnO3, that is, Yb1−xSrxMnO3 (x = 0.05 and 0.1) multiferroic ceramics, by investigating their magnetic and thermodynamic properties. X-ray powder diffraction measurements on these compounds demonstrated that both samples crystallize in the hexagonal phase with P63cm space group. Antiferromagnetic ordering temperature, as determined by heat capacity measurements, is found to decrease from 86 K (at x = 0.05) to 83 K (at x = 0.1). The compound shows ferromagnetic (FM) ordering at ~4 K due to ordering of the moments of Yb3+ ions in crystallographic site 2a. A Schottky anomaly is observed at around 7 K due to moments of the Yb3+ at the 4b sites as a result of the molecular field present due to Mn3+ ions. Effective molecular fields are estimated to be H mf = 4.6 and 3.0 T for x = 0.05 and x = 0.1, respectively, from the analysis of the observed Schottky anomaly. The magnetic entropy changes (ΔS mag) obtained for samples with x = 0.05 and 0.1 are found to be ~2.24 and ~2.1 J mol−1 K−1, respectively. The relative cooling power is found to be 29 and 28 J mol−1 for a field change of 100 kOe for x = 0.05 and 0.1, respectively.

Magnetic structure of Cu2MnBO5 ludwigite: thermodynamic, magnetic properties and neutron diffraction study

We report on the thermodynamic, magnetic properties and the magnetic structure of ludwigite-type Cu2MnBO5. The specific heat, low-field magnetization and paramagnetic susceptibility were studied on a single crystal and combined with powder neutron diffraction data. The temperature dependence of the specific heat and the neutron diffraction pattern reveal a single magnetic phase transition at T = 92 K, which corresponds to the magnetic ordering into a ferrimagnetic phase. The cation distribution and the values and directions of magnetic moments of ions in different crystallographic sites are established. The magnetic moments of Cu2+ and Mn3+ ions occupying different magnetic sites in the ferrimagnetic phase are pairwise antiparallel and their directions do not coincide with the directions of the principal crystallographic axes. The small value of the magnetic moment of copper ions occupying site 2a is indicative of partial disordering of the magnetic moments on this site. The magnetization measurements show a strong temperature hysteresis of magnetization, which evidences for field-dependent transitions below the phase transition temperature.

Local food environment and fruit and vegetable consumption: An ecological study.

Ecological studies are essential for understanding the environment-diet relationship. The purpose of this study was to describe environmental conditions and their relationship with fruit and vegetable (FV) consumption among Brazilian public health service users in the city of Belo Horizonte. We evaluated food stores contained within 1600 m buffer zones at 18 Health Academy Programme sites, from 2013 to 2014. Variables at the community (density, proximity and type) and the consumer (sectional location of FV; availability, quality, variety, price and advertising of FV and ultra-processed foods) nutrition environment were measured by direct observation, while aggregate data from users (income and FV consumption) were obtained by interview. Data were analysed using the Kernel intensity estimator, average nearest neighbour value and Local Moran's Index for local spatial autocorrelation. We interviewed 3414 users and analysed 336 food stores. Major geographical variations in the FV consumption were identified. Average consumption was higher (site 2A: 410.5 ± 185.7 g vs. site 4B: 311.2 ± 159.9 g) in neighbourhoods with higher income and concentration of food stores, and better index of access to healthy foods. Sites with poor FV consumption had the most stores with poor access to healthy foods (index in the first tertile, ≤ 10). In conclusion, negative characteristics of the food environment, as seen in the present study, may contribute to low FV consumption, suggesting the need for the development and consolidation of public policies aimed at creating healthy environments through built environment interventions that increase access to and consumption of healthy foods like FV.

Type-I silicon clathrates containing lithium

Abstract The intermetallic phase [Li x Ba8−x ][Li y Si46−y ] is the first example of a clathrate-I silicide containing lithium with the peculiarity that the Li atoms occupy both cage and framework positions. The phase was obtained by ambient-pressure solid-state reactions at 400°C between Ba4Li2Si6 and the binary clathrate phase Ba8−x Si46 applying different experimental approaches. In this reaction, Li atoms substitute Si atoms of the framework at site 6c (space group Pm3̅n) and fill up the Ba-deficient dodecahedral Si20 cages at site 2a. The presence of Li atoms in the clathrate phase was proven by combined X-ray powder diffraction, 7Li and 29Si solid-state NMR analyses. Incorporation of lithium markedly increased the lattice parameter of the clathrate phase, e.g. from a=10.3200(2) Å for Ba7.48(2)Si46 to a=10.3715(3) Å for [Li0.59(2)Ba7.41(2)][Li2.3(1)Si43.7(1)]. The critical temperature T c for the transition to the superconducting state decreased from 7.7 K to 3.2 K upon Li incorporation.

The PIN domain endonuclease Utp24 cleaves pre-ribosomal RNA at two coupled sites in yeast and humans

During ribosomal RNA (rRNA) maturation, cleavages at defined sites separate the mature rRNAs from spacer regions, but the identities of several enzymes required for 18S rRNA release remain unknown. PilT N-terminus (PIN) domain proteins are frequently endonucleases and the PIN domain protein Utp24 is essential for early cleavages at three pre-rRNA sites in yeast (A0, A1 and A2) and humans (A0, 1 and 2a). In yeast, A1 is cleaved prior to A2 and both cleavages require base-pairing by the U3 snoRNA to the central pseudoknot elements of the 18S rRNA. We found that yeast Utp24 UV-crosslinked in vivo to U3 and the pseudoknot, placing Utp24 close to cleavage at site A1. Yeast and human Utp24 proteins exhibited in vitro endonuclease activity on an RNA substrate containing yeast site A2. Moreover, an intact PIN domain in human UTP24 was required for accurate cleavages at sites 1 and 2a in vivo, whereas mutation of another potential site 2a endonuclease, RCL1, did not affect 18S production. We propose that Utp24 cleaves sites A1/1 and A2/2a in yeast and human cells.

The PIN domain endonuclease Utp24 cleaves pre-ribosomal RNA at two coupled sites in yeast and humans

During ribosomal RNA (rRNA) maturation, cleavages at defined sites separate the mature rRNAs from spacer regions, but the identities of several enzymes required for 18S rRNA release remain unknown. PilT N-terminus (PIN) domain proteins are frequently endonucleases and the PIN domain protein Utp24 is essential for early cleavages at three pre-rRNA sites in yeast (A0, A1 and A2) and humans (A0, 1 and 2a). In yeast, A1 is cleaved prior to A2 and both cleavages require base-pairing by the U3 snoRNA to the central pseudoknot elements of the 18S rRNA. We found that yeast Utp24 UV-crosslinked in vivo to U3 and the pseudoknot, placing Utp24 close to cleavage at site A1. Yeast and human Utp24 proteins exhibited in vitro endonuclease activity on an RNA substrate containing yeast site A2. Moreover, an intact PIN domain in human UTP24 was required for accurate cleavages at sites 1 and 2a in vivo, whereas mutation of another potential site 2a endonuclease, RCL1, did not affect 18S production. We propose that Utp24 cleaves sites A1/1 and A2/2a in yeast and human cells.