2009 Nobel Laureates

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4 {$ x& N3 I( L% l' FPhysiology or Medicine
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Cell Press congratulates Cell Editorial Board member Elizabeth H. Blackburn, Cancer Cell Editorial Board member Carol W. Greider, and Chemistry & Biology Editorial Board member Jack W. Szostak for being awarded the 2009 Nobel Prize in Physiology or Medicine.
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Read the award winners' groundbreaking research published in Cell, free!

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The telomere terminal transferase of Tetrahymena is a ribonucleoprotein enzyme with two kinds of primer specificity
5 d  l5 Z9 e) O( \0 U+ [3 gGreider CW, Blackburn EH7 J0 N9 F% C: ~8 A( T
Cell 51(6), 887-898
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/ b( ?8 m. j7 F0 y0 p' w8 R8 Q, gDepartment of Molecular Biology University of California Berkeley, California 94720 USA  B+ r0 X1 [! I+ b2 e3 k3 a
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  K4 ^8 o, o% w- u9 wWe have analyzed the de novo telomere synthesis catalyzed by the enzyme telomere terminal transferase (telomerase) from Tetrahymena. Oligonucleotides representing the G-rich strand of telomeric sequences from five different organisms specifically primed the addition of TTGGGG repeats in vitro, suggesting that primer recognition may involve a DNA structure unique to these oligonucleotides. The sequence at the 3′ end of the oligonucleotide primer specified the first nucleotide added in the reaction. Furthermore, the telomerase was shown to be a ribonucleoprotein complex whose RNA and protein components were both essential for activity. After extensive purification of the enzyme by a series of five different chromatographic steps, a few small low abundance RNAs copurified with the activity.
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Identification of a specific telomere terminal transferase activity in Tetrahymena extracts
. [0 C  Y- ~; ^% EGreider CW, Blackburn EH
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Summary8 H8 g, d" ^0 O9 i5 h
We have found a novel activity in Tetrahymena cell free extracts that adds tandem TTGGGG repeats onto synthetic telomere primers. The single-stranded DNA oligonucleotides (TTGGGG)4 and TGTGTGGGTGTGTGGGTGTGTGGG, consisting of the Tetrahymena and yeast telomeric sequences respectively, each functioned as primers for elongation, while (CCCCAA)4 and two nontelomeric sequence DNA oligomers did not. Efficient synthesis of the TTGGGG repeats depended only on addition of micromolar concentrations of oligomer primer, dGTP, and dTTP to the extract. The activity was sensitive to heat and proteinase K treatment. The repeat addition was independent of both endogenous Tetrahymena DNA and the endogenous α-type DNA polymerase; and a greater elongation activity was present during macronuclear development, when a large number of telomeres are formed and replicated, than during vegetative cell growth. We propose that the novel telomere terminal transferase is involved in the addition of telomeric repeats necessary for the replication of chromosome ends in eukaryotes.
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Cloning yeast telomeres on linear plasmid vectors
4 ~5 n! j: j4 ^0 v- ~6 Y) w' @Szostak JW, Blackburn EH
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Department of Molecular Biology University of California Berkeley, California 94720 USA
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Summary8 a5 w& c: s+ v
We have constructed a linear yeast plasmid by joining fragments from the termini of Tetrahymena ribosomal DNA to a yeast vector. Structural features of the terminus region of the Tetrahymena rDNA plasmid maintained in the yeast linear plasmid include a set of specifically placed single-strand interruptions within the cluster of hexanucleotide (C4A2) repeat units. An artificially constructed hairpin terminus was unable to stabilize a linear plasmid in yeast. The fact that yeast can recognize and use DNA ends from the distantly related organism Tetrahymena suggests that the structural features required for telomere replication and resolution have been highly conserved in evolution. The linear plasmid was used as a vector to clone chromosomal telomeres from yeast. One Tetrahymena end was removed by restriction digestion, and yeast fragments that could function as an end on a linear plasmid were selected. Restriction mapping and hybridization analysis demonstrated that these fragments were yeast telomeres, and suggested that all yeast chromosomes might have a common telomere sequence. Yeast telomeres appear to be similar in structure to the rDNA of Tetrahymena, in which specific nicks or gaps are present within a simple repeated sequence near the terminus of the DNA.% U: z9 |% ?1 y8 W# [
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Chemistry8 U4 V" T  U4 J6 v: Y. Q
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Cell Press congratulates Cell Editorial Board member Venkatraman Ramakrishnan, Structure Editorial Board member Thomas A. Steitz, and Cell Press author Ada E. Yonath for being awarded the 2009 Nobel Prize in Chemistry.
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Read the award winners' groundbreaking research published in Cell, free!

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Selection of tRNA by the ribosome requires a transition from an open to a closed form.
9 F8 z: X% N) XOgle JM, Murphy FV, Tarry MJ, Ramakrishnan V.( P, C5 K+ N- X  t: C
Cell 111(5), 721-32
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" O3 W" v- m& E( SCorrespondence: V. Ramakrishnan, +44 1223 402295 (phone), + 44 1223 213556 (fax)
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A structural and mechanistic explanation for the selection of tRNAs by the ribosome has been elusive. Here, we report crystal structures of the 30S ribosomal subunit with codon and near-cognate tRNA anticodon stem loops bound at the decoding center and compare affinities of equivalent complexes in solution. In ribosomal interactions with near-cognate tRNA, deviation from Watson-Crick geometry results in uncompensated desolvation of hydrogen-bonding partners at the codon-anticodon minor groove. As a result, the transition to a closed form of the 30S induced by cognate tRNA is unfavorable for near-cognate tRNA unless paromomycin induces part of the rearrangement. We conclude that stabilization of a closed 30S conformation is required for tRNA selection, and thereby structurally rationalize much previous data on translational fidelity.+ [' W; g% V$ _2 Q: o" B! n$ |! h# ^7 S3 B

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