Nucleic Acids Research, 2000, Vol. 28, No. 18 3524-3534
© 2000 Oxford University Press
Complete deletion of yeast chromosomal rDNA repeats and integration of a new rDNA repeat: use of rDNA deletion strains for functional analysis of rDNA promoter elements in vivo
Department of Biological Chemistry, University of California at Irvine, Irvine, CA 92697-1700, USA
Strains of Saccharomyces cerevisiae were constructed in which chromosomal rDNA repeats are completely deleted and their growth is supported by a plasmid carrying a single rDNA repeat, either a plasmid carrying the 35S rRNA gene transcribed from the native promoter by RNA polymerase I or a plasmid carrying the 35S rRNA gene fused to the GAL7 promoter for transcription by RNA polymerase II. This system has made it possible to assess the expression of rDNA by measuring the ability of synthesized rRNA to support cell growth as well as by measuring the actual rRNA synthesized rather than by the use of reporter mini-rDNA genes. Using this system, deletion analysis of the rDNA promoter confirmed the presence of two elements, the upstream element and the core promoter, and showed that basal transcription from the core promoter, if it takes place in vivo as was observed in vitro, is not sufficient to allow cell growth. We have also succeeded in integration of a rDNA repeat and its copy number expansion at the original chromosomal locus, which will allow future mutational analysis not only of rRNA but also other DNA elements involved in rRNA transcription, rDNA replication and recombination within a repeated rDNA structure.
* To whom correspondence should be addressed. Tel: +1 949 824 4564; Fax: +1 949 824 3201; Email: mnomura@uci.edu
CiteULike 
Connotea 
Del.icio.us What's this?
This article has been cited by other articles:
|
|
 |
|
  B.-S. Shin, J.-R. Kim, M. G. Acker, K. N. Maher, J. R. Lorsch, and T. E. Dever rRNA Suppressor of a Eukaryotic Translation Initiation Factor 5B/Initiation Factor 2 Mutant Reveals a Binding Site for Translational GTPases on the Small Ribosomal Subunit Mol. Cell. Biol., February 1, 2009; 29(3): 808 - 821. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 C. Luttermann and G. Meyers The importance of inter- and intramolecular base pairing for translation reinitiation on a eukaryotic bicistronic mRNA Genes & Dev., February 1, 2009; 23(3): 331 - 344. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
M. Sullivan, L. Holt, and D. O. Morgan Cyclin-Specific Control of Ribosomal DNA Segregation Mol. Cell. Biol., September 1, 2008; 28(17): 5328 - 5336. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
J. White, Z. Li, R. Sardana, J. M. Bujnicki, E. M. Marcotte, and A. W. Johnson Bud23 Methylates G1575 of 18S rRNA and Is Required for Efficient Nuclear Export of Pre-40S Subunits Mol. Cell. Biol., May 15, 2008; 28(10): 3151 - 3161. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 A. S. Bommakanti, L. Lindahl, and J. M. Zengel Mutation from guanine to adenine in 25S rRNA at the position equivalent to E. coli A2058 does not confer erythromycin sensitivity in Sacchromyces cerevisae RNA, March 1, 2008; 14(3): 460 - 464. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 R. Rakauskaite and J. D. Dinman rRNA mutants in the yeast peptidyltransferase center reveal allosteric information networks and mechanisms of drug resistance Nucleic Acids Res., March 1, 2008; 36(5): 1497 - 1507. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
H. Fan-Minogue and D. M. Bedwell Eukaryotic ribosomal RNA determinants of aminoglycoside resistance and their role in translational fidelity RNA, January 1, 2008; 14(1): 148 - 157. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
S. N. Hobbie, S. K. Kalapala, S. Akshay, C. Bruell, S. Schmidt, S. Dabow, A. Vasella, P. Sander, and E. C. Bottger Engineering the rRNA decoding site of eukaryotic cytosolic ribosomes in bacteria Nucleic Acids Res., September 25, 2007; 35(18): 6086 - 6093. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
R. Rakauskaite and J. D. Dinman An Arc of Unpaired "Hinge Bases" Facilitates Information Exchange among Functional Centers of the Ribosome Mol. Cell. Biol., December 1, 2006; 26(23): 8992 - 9002. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
M. L. Oakes, K. Johzuka, L. Vu, K. Eliason, and M. Nomura Expression of rRNA Genes and Nucleolus Formation at Ectopic Chromosomal Sites in the Yeast Saccharomyces cerevisiae. Mol. Cell. Biol., August 1, 2006; 26(16): 6223 - 6238. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 S. A. Chappell, J. Dresios, G. M. Edelman, and V. P. Mauro Ribosomal shunting mediated by a translational enhancer element that base pairs to 18S rRNA PNAS, June 20, 2006; 103(25): 9488 - 9493. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
K. Johzuka, M. Terasawa, H. Ogawa, T. Ogawa, and T. Horiuchi Condensin Loaded onto the Replication Fork Barrier Site in the rRNA Gene Repeats during S Phase in a FOB1-Dependent Fashion To Prevent Contraction of a Long Repetitive Array in Saccharomyces cerevisiae. Mol. Cell. Biol., March 1, 2006; 26(6): 2226 - 2236. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
F. Di Felice, F. Cioci, and G. Camilloni FOB1 affects DNA topoisomerase I in vivo cleavages in the enhancer region of the Saccharomyces cerevisiae ribosomal DNA locus Nucleic Acids Res., November 3, 2005; 33(19): 6327 - 6337. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
D. A. Schneider and M. Nomura RNA polymerase I remains intact without subunit exchange through multiple rounds of transcription in Saccharomyces cerevisiae PNAS, October 19, 2004; 101(42): 15112 - 15117. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 T. Weitao, M. Budd, L. L. M. Hoopes, and J. L. Campbell Dna2 Helicase/Nuclease Causes Replicative Fork Stalling and Double-strand Breaks in the Ribosomal DNA of Saccharomyces cerevisiae J. Biol. Chem., June 13, 2003; 278(25): 22513 - 22522. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
A. G. Hinnebusch Unleashing yeast genetics on a factor-independent mechanism of internal translation initiation PNAS, November 6, 2001; 98(23): 12866 - 12868. [Full Text] [PDF]
|
|
|
|
|
|
H. Wai, K. Johzuka, L. Vu, K. Eliason, T. Kobayashi, T. Horiuchi, and M. Nomura Yeast RNA Polymerase I Enhancer Is Dispensable for Transcription of the Chromosomal rRNA Gene and Cell Growth, and Its Apparent Transcription Enhancement from Ectopic Promoters Requires Fob1 Protein Mol. Cell. Biol., August 15, 2001; 21(16): 5541 - 5553. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 M. NOMURA Ribosomal RNA Genes, RNA Polymerases, Nucleolar Structures, and Synthesis of rRNA in the Yeast Saccharomyces cerevisiae Cold Spring Harb Symp Quant Biol, January 1, 2001; 66(0): 555 - 566. [Abstract] [PDF]
|
|