86,566 research outputs found
Vetus Latina. Die Reste der altlateinischen Bibel, nach Petrus Sabatier neu gesammelt und herausgegeben von der Erzabtei Beuron
No full textDekkers F. Vetus Latina. Die Reste der altlateinischen Bibel, nach Petrus Sabatier neu gesammelt und herausgegeben von der Erzabtei Beuron. In: Scriptorium, Tome 6 n°1, 1952. pp. 159-161
Nanostructures from Synthetic Genetic Polymers
No full textNanoscale objects of increasing complexity can be constructed
from DNA or RNA. However, the scope of potential applications
could be enhanced by expanding beyond the moderate
chemical diversity of natural nucleic acids. Here, we explore
the construction of nano-objects made entirely from alternative
building blocks: synthetic genetic polymers not found in
nature, also called xeno nucleic acids (XNAs). Specifically, we
describe assembly of 70 kDa tetrahedra elaborated in four
different XNA chemistries (2’-fluro-2’-deoxy-ribofuranose nucleic
acid (2’F-RNA), 2’-fluoroarabino nucleic acids (FANA), hexitol
nucleic acids (HNA), and cyclohexene nucleic acids (CeNA)), as
well as mixed designs, and a ~600 kDa all-FANA octahedron,
visualised by electron microscopy. Our results extend the
chemical scope for programmable nanostructure assembly,
with implications for the design of nano-objects and materials
with an expanded range of structural and physicochemical
properties, including enhanced biostability.sponsorship: Medical Research Council (program no. U105178804 to P.H.)status: Publishe
Origin licensing requires ATP binding and hydrolysis by the MCM replicative helicase.
No full textLoading of the six related Minichromosome Maintenance (MCM) proteins as head-to-head double hexamers during DNA replication origin licensing is crucial for ensuring once-per-cell-cycle DNA replication in eukaryotic cells. Assembly of these prereplicative complexes (pre-RCs) requires the Origin Recognition Complex (ORC), Cdc6, and Cdt1. ORC, Cdc6, and MCM are members of the AAA+ family of ATPases, and pre-RC assembly requires ATP hydrolysis. Here we show that ORC and Cdc6 mutants defective in ATP hydrolysis are competent for origin licensing. However, ATP hydrolysis by Cdc6 is required to release nonproductive licensing intermediates. We show that ATP binding stabilizes the wild-type MCM hexamer. Moreover, by analyzing MCM containing mutant subunits, we show that ATP binding and hydrolysis by MCM are required for Cdt1 release and double hexamer formation. This work alters our view of how ATP is used by licensing factors to assemble pre-RCs
Prereplicative complexes assembled in vitro support origin-dependent and independent DNA replication.
No full textEukaryotic DNA replication initiates from multiple replication origins. To ensure each origin fires just once per cell cycle, initiation is divided into two biochemically discrete steps: the Mcm2-7 helicase is first loaded into prereplicative complexes (pre-RCs) as an inactive double hexamer by the origin recognition complex (ORC), Cdt1 and Cdc6; the helicase is then activated by a set of "firing factors." Here, we show that plasmids containing pre-RCs assembled with purified proteins support complete and semi-conservative replication in extracts from budding yeast cells overexpressing firing factors. Replication requires cyclin-dependent kinase (CDK) and Dbf4-dependent kinase (DDK). DDK phosphorylation of Mcm2-7 does not by itself promote separation of the double hexamer, but is required for the recruitment of firing factors and replisome components in the extract. Plasmid replication does not require a functional replication origin; however, in the presence of competitor DNA and limiting ORC concentrations, replication becomes origin-dependent in this system. These experiments indicate that Mcm2-7 double hexamers can be precursors of replication and provide insight into the nature of eukaryotic DNA replication origins
Structural basis of Ty3 retrotransposon integration at RNA Polymerase III-transcribed genes.
No full textRetrotransposons are endogenous elements that have the ability to mobilise their DNA between different locations in the host genome. The Ty3 retrotransposon integrates with an exquisite specificity in a narrow window upstream of RNA Polymerase (Pol) III-transcribed genes, representing a paradigm for harmless targeted integration. Here we present the cryo-EM reconstruction at 4.0 Å of an active Ty3 strand transfer complex bound to TFIIIB transcription factor and a tRNA gene. The structure unravels the molecular mechanisms underlying Ty3 targeting specificity at Pol III-transcribed genes and sheds light into the architecture of retrotransposon machinery during integration. Ty3 intasome contacts a region of TBP, a subunit of TFIIIB, which is blocked by NC2 transcription regulator in RNA Pol II-transcribed genes. A newly-identified chromodomain on Ty3 integrase interacts with TFIIIB and the tRNA gene, defining with extreme precision the integration site position
Structure of a nucleosome-bound MuvB transcription factor complex reveals DNA remodelling
No full textGenes encoding the core cell cycle machinery are transcriptionally regulated by the MuvB family of protein complexes in a cell cycle-specific manner. Complexes of MuvB with the transcription factors B-MYB and FOXM1 activate mitotic genes during cell proliferation. The mechanisms of transcriptional regulation by these complexes are still poorly characterised. Here, we combine biochemical analysis and in vitro reconstitution, with structural analysis by cryo-electron microscopy and cross-linking mass spectrometry, to functionally examine these complexes. We find that the MuvB:B-MYB complex binds and remodels nucleosomes, thereby exposing nucleosomal DNA. This remodelling activity is supported by B-MYB which directly binds the remodelled DNA. Given the remodelling activity on the nucleosome, we propose that the MuvB:B-MYB complex functions as a pioneer transcription factor complex. In this work, we rationalise prior biochemical and cellular studies and provide a molecular framework of interactions on a protein complex that is key for cell cycle regulation
Variations on the Author
No full text“Variations on the Author” discusses two of Eduardo Coutinho’s recent films (Um Dia na Vida, from 2010, and Últimas Conversas, posthumously released in 2015) and their contribution to the general question of documentary authorship. The director’s filmography is characterized by a consistent yet self-effacing form of authorial self-inscription: Coutinho often features as an interviewer that rather than express opinions propels discourses; an interviewer that is good at listening. This mode of self-inscription characterizes him as an author who is not expressive but who is nonetheless markedly present on the screen. In Um Dia na Vida, however, Coutinho is completely absent form the image, while Últimas Conversas, on the contrary, includes a confessional prologue that moves the director from the margins to the center of his films. This article examines the ways in which these works stand out in the filmography of a director who offers new insights into the notion of cinematic authorship
Nse5/6 is a negative regulator of the ATPase activity of the Smc5/6 complex
No full textThe multi-component Smc5/6 complex plays a critical role in the resolution of recombination intermediates formed during mitosis and meiosis, and in the cellular response to replication stress. Using recombinant proteins, we have reconstituted a series of defined Saccharomyces cerevisiae Smc5/6 complexes, visualised them by negative stain electron microscopy, and tested their ability to function as an ATPase. We find that only the six protein ‘holo-complex’ is capable of turning over ATP and that its activity is significantly increased by the addition of double-stranded DNA to reaction mixes. Furthermore, stimulation is wholly dependent on functional ATP-binding pockets in both Smc5 and Smc6. Importantly, we demonstrate that budding yeast Nse5/6 acts as a negative regulator of Smc5/6 ATPase activity, binding to the head-end of the complex to suppress turnover, irrespective of the DNA-bound status of the complex
Human Condensin I and II Drive Extensive ATP-Dependent Compaction of Nucleosome-Bound DNA
No full textStructural maintenance of chromosomes (SMC) complexes are essential for genome organization from bacteria to humans, but their mechanisms of action remain poorly understood. Here, we characterize human SMC complexes condensin I and II and unveil the architecture of the human condensin II complex, revealing two putative DNA-entrapment sites. Using single-molecule imaging, we demonstrate that both condensin I and II exhibit ATP-dependent motor activity and promote extensive and reversible compaction of double-stranded DNA. Nucleosomes are incorporated into DNA loops during compaction without being displaced from the DNA, indicating that condensin complexes can readily act upon nucleosome-bound DNA molecules. These observations shed light on critical processes involved in genome organization in human cells
A closed conformation of the Caenorhabditis elegans separase-securin complex
No full textThe protease separase plays a key role in sister chromatid disjunction and centriole disengagement. To maintain genomic stability, separase activity is strictly regulated by binding of an inhibitory protein, securin. Despite its central role in cell division, the separase and securin complex is poorly understood at the structural level. This is partly due to the difficulty of generating a sufficient quantity of homogenous, stable protein. Here we report the production of C. elegans separase-securin complex and its characterisation using biochemical methods and by negative staining electron microscopy. Single particle analysis generated a density map at a resolution of 21-24 Å that reveals a close, globular structure of complex connectivity harbouring two lobes. One lobe matches closely a homology model of the N-terminal HEAT repeat domain of separase while the second lobe readily accommodates homology models of the separase C-terminal death and caspase-like domains. The globular structure of the C. elegans separase-securin complex contrasts with the more elongated structure previously described for the H. sapiens complex, which could represent a different functional state of the complex, suggesting a mechanism for the regulation of separase activity through conformational change
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