Ribosome biogenesis and function are ubiquitous processes central to cell growth and regulation. The long-term goal of this work is to understand the mechanisms of coordinate expression and assembly of yeast, ribosomal protein and ribosomal RNA and the mechanisms whereby ribosomes catalyze protein synthesis. The mechanisms of equimolar synthesis of ribosomal proteins and rRNAs or of coordinate changes in their synthesis are not yet fully understood. Ribosomal RNAs, transcribed in the nucleolus, and ribosomal proteins, synthesized in the cytoplasm, assemble into ribosomes within the nucleolus. Little is known about the structure and function of nonribosomal nucleolar molecules or about the positions or function of individual ribosomal molecules in the assembly pathway or within the mature ribosome. Cloned genes for three ribosomal proteins, rp59, L16, and L1, and nucleolar protein genes will be utilized to investigate the following specific aims: (1) Expression of CRY2 encoding rp59 is feedback inhibited by rp59, suggesting a mechanism for coupling ribosome synthesis and assembly. Steps at which CM expression is repressed are being determined. Cis-acting repressor sites and transacting repressors are being defined. (2a) By immunoelectron microscopy L16 was localized near the central protuberance of the 60S subunit. Biochemical and genetic experiments are planned to map L16 at higher resolution and to define nearest neighbors of L16. (b)The effect of rpll6 conditional lethal mutations on ribosome assembly and function will be assayed to assess the role of L16 in these processes. (3)Ribosomal protein L1 (or its homologues) and 5S rRNA form an RNP complex that is a precursor to ribosome assembly in mammals, Xenopus, and bacteria. Using wild type d mutant RPL1 and 5S rRNA genes and antiserum specific for the RNP, we will define the pathway of 5S RNP biogenesis. Domains of Ll and 5S rRNA necessary for their binding to each other will be mapped. Models will be tested whereby 5S rRNA synthesis may be coupled to ribosomal protein synthesis. (4) Nucleolar protein genes will be cloned and mutant alleles isolated to assess functions of nucleolar proteins.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM028301-15
Application #
2175153
Study Section
Biochemistry Study Section (BIO)
Project Start
1980-08-01
Project End
1995-08-31
Budget Start
1994-09-01
Budget End
1995-08-31
Support Year
15
Fiscal Year
1994
Total Cost
Indirect Cost
Name
Carnegie-Mellon University
Department
Biology
Type
Schools of Arts and Sciences
DUNS #
052184116
City
Pittsburgh
State
PA
Country
United States
Zip Code
15213
Biedka, Stephanie; Micic, Jelena; Wilson, Daniel et al. (2018) Hierarchical recruitment of ribosomal proteins and assembly factors remodels nucleolar pre-60S ribosomes. J Cell Biol 217:2503-2518
Biedka, Stephanie; Wu, Shan; LaPeruta, Amber J et al. (2017) Insights into remodeling events during eukaryotic large ribosomal subunit assembly provided by high resolution cryo-EM structures. RNA Biol 14:1306-1313
Ma, Chengying; Wu, Shan; Li, Ningning et al. (2017) Structural snapshot of cytoplasmic pre-60S ribosomal particles bound by Nmd3, Lsg1, Tif6 and Reh1. Nat Struct Mol Biol 24:214-220
Wu, Shan; Tan, Dan; Woolford Jr, John L et al. (2017) Atomic modeling of the ITS2 ribosome assembly subcomplex from cryo-EM together with mass spectrometry-identified protein-protein crosslinks. Protein Sci 26:103-112
Konikkat, Salini; Woolford Jr, John L (2017) Principles of 60S ribosomal subunit assembly emerging from recent studies in yeast. Biochem J 474:195-214
Liu, Zheng; Gutierrez-Vargas, Cristina; Wei, Jia et al. (2016) Structure and assembly model for the Trypanosoma cruzi 60S ribosomal subunit. Proc Natl Acad Sci U S A 113:12174-12179
Ramesh, Madhumitha; Woolford Jr, John L (2016) Eukaryote-specific rRNA expansion segments function in ribosome biogenesis. RNA 22:1153-62
Tutuncuoglu, Beril; Jakovljevic, Jelena; Wu, Shan et al. (2016) The N-terminal extension of yeast ribosomal protein L8 is involved in two major remodeling events during late nuclear stages of 60S ribosomal subunit assembly. RNA 22:1386-99
Talkish, Jason; Biedka, Stephanie; Jakovljevic, Jelena et al. (2016) Disruption of ribosome assembly in yeast blocks cotranscriptional pre-rRNA processing and affects the global hierarchy of ribosome biogenesis. RNA 22:852-66
Kormuth, Karen A; Woolford Jr, John L; Armitage, Bruce A (2016) Homologous PNA Hybridization to Noncanonical DNA G-Quadruplexes. Biochemistry 55:1749-57

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