17
views
0
recommends
+1 Recommend
0 collections
    0
    shares
      • Record: found
      • Abstract: found
      • Article: found
      Is Open Access

      Structure and folding of the Tetrahymena telomerase RNA pseudoknot

      research-article
      , *
      Nucleic Acids Research
      Oxford University Press

      Read this article at

      Bookmark
          There is no author summary for this article yet. Authors can add summaries to their articles on ScienceOpen to make them more accessible to a non-specialist audience.

          Abstract

          Telomerase maintains telomere length at the ends of linear chromosomes using an integral telomerase RNA (TER) and telomerase reverse transcriptase (TERT). An essential part of TER is the template/pseudoknot domain (t/PK) which includes the template, for adding telomeric repeats, template boundary element (TBE), and pseudoknot, enclosed in a circle by stem 1. The Tetrahymena telomerase holoenzyme catalytic core (p65-TER-TERT) was recently modeled in our 9 Å resolution cryo-electron microscopy map by fitting protein and TER domains, including a solution NMR structure of the Tetrahymena pseudoknot. Here, we describe in detail the structure and folding of the isolated pseudoknot, which forms a compact structure with major groove U•A-U and novel C•G-A + base triples. Base substitutions that disrupt the base triples reduce telomerase activity in vitro. NMR studies also reveal that the pseudoknot does not form in the context of full-length TER in the absence of TERT, due to formation of a competing structure that sequesters pseudoknot residues. The residues around the TBE remain unpaired, potentially providing access by TERT to this high affinity binding site during an early step in TERT-TER assembly. A model for the assembly pathway of the catalytic core is proposed.

          Related collections

          Most cited references85

          • Record: found
          • Abstract: found
          • Article: not found

          Identification of a specific telomere terminal transferase activity in Tetrahymena extracts.

          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 alpha-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.
            Bookmark
            • Record: found
            • Abstract: found
            • Article: not found

            Automated 3D structure composition for large RNAs

            Understanding the numerous functions that RNAs play in living cells depends critically on knowledge of their three-dimensional structure. Due to the difficulties in experimentally assessing structures of large RNAs, there is currently great demand for new high-resolution structure prediction methods. We present the novel method for the fully automated prediction of RNA 3D structures from a user-defined secondary structure. The concept is founded on the machine translation system. The translation engine operates on the RNA FRABASE database tailored to the dictionary relating the RNA secondary structure and tertiary structure elements. The translation algorithm is very fast. Initial 3D structure is composed in a range of seconds on a single processor. The method assures the prediction of large RNA 3D structures of high quality. Our approach needs neither structural templates nor RNA sequence alignment, required for comparative methods. This enables the building of unresolved yet native and artificial RNA structures. The method is implemented in a publicly available, user-friendly server RNAComposer. It works in an interactive mode and a batch mode. The batch mode is designed for large-scale modelling and accepts atomic distance restraints. Presently, the server is set to build RNA structures of up to 500 residues.
              Bookmark
              • Record: found
              • Abstract: found
              • Article: not found

              Secondary structure of vertebrate telomerase RNA.

              Telomerase is a ribonucleoprotein enzyme that maintains telomere length by adding telomeric sequence repeats onto chromosome ends. The essential RNA component of telomerase provides the template for telomeric repeat synthesis. To determine the secondary structure of vertebrate telomerase RNA, 32 new telomerase RNA genes were cloned and sequenced from a variety of vertebrate species including 18 mammals, 2 birds, 1 reptile, 7 amphibians, and 4 fishes. Using phylogenetic comparative analysis, we propose a secondary structure that contains four structural domains conserved in all vertebrates. Ten helical regions of the RNA are universally conserved while other regions vary significantly in length and sequence between different classes of vertebrates. The proposed vertebrate telomerase RNA structure displays a strikingly similar topology to the previously determined ciliate telomerase RNA structure, implying an evolutionary conservation of the global architecture of telomerase RNA.
                Bookmark

                Author and article information

                Journal
                Nucleic Acids Res
                Nucleic Acids Res
                nar
                nar
                Nucleic Acids Research
                Oxford University Press
                0305-1048
                1362-4962
                09 January 2017
                28 November 2016
                28 November 2016
                : 45
                : 1
                : 482-495
                Affiliations
                Department of Chemistry and Biochemistry, and Molecular Biology Institute, University of California, Los Angeles, CA 90095-1569, USA
                Author notes
                [* ]To whom correspondence should be addressed. Tel: +1 310 206 6922; Fax: +1 310 825 0982; Email: feigon@ 123456mbi.ucla.edu
                Article
                10.1093/nar/gkw1153
                5224487
                27899638
                4c3f6990-9b0d-4f18-b20b-08ddac683bda
                © The Author(s) 2016. Published by Oxford University Press on behalf of Nucleic Acids Research.

                This is an Open Access article distributed under the terms of the Creative Commons Attribution License ( http://creativecommons.org/licenses/by-nc/4.0/), which permits non-commercial re-use, distribution, and reproduction in any medium, provided the original work is properly cited. For commercial re-use, please contact journals.permissions@ 123456oup.com

                History
                : 03 November 2016
                : 26 October 2016
                : 24 June 2016
                Page count
                Pages: 14
                Categories
                Structural Biology
                Custom metadata
                09 January 2017

                Genetics
                Genetics

                Comments

                Comment on this article