Genes Encoding Ribosomal Proteins Rps0A/B of Saccharomyces cerevisiae Interact With TOM1 Mutants Defective in Ribosome Synthesis

Genetics ◽  
2001 ◽  
Vol 157 (3) ◽  
pp. 1107-1116
Author(s):  
Amy L Tabb ◽  
Takahiko Utsugi ◽  
Clavia R Wooten-Kee ◽  
Takeshi Sasaki ◽  
Steven A Edling ◽  
...  
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Ribosomal Proteins ◽  
Elevated Temperatures ◽  
Synergistic Effects ◽  
Ribosomal Subunit ◽  
Hect Domain ◽  
Ribosome Synthesis ◽  
Ubiquitin Protein Ligase ◽  
40S Ribosomal Subunit ◽  
Genes Encoding

Abstract The Saccharomyces cerevisiae RPS0A/B genes encode proteins of the 40S ribosomal subunit that are required for the maturation of 18S rRNA. We show here that the RPS0 genes interact genetically with TOM1. TOM1 encodes a member of the hect-domain-containing E3 ubiquitin-protein ligase family that is required for growth at elevated temperatures. Mutant alleles of the RPS0 and TOM1 genes have synergistic effects on cell growth at temperatures permissive for TOM1 mutants. Moreover, the growth arrest of TOM1 mutants at elevated temperatures is partially suppressed by overexpression of RPS0A/B. Strains with mutant alleles of TOM1 are defective in multiple steps in rRNA processing, and interactions between RPS0A/B and TOM1 stem, in part, from their roles in the maturation of ribosomal subunits. Ribosome synthesis is therefore included among the cellular processes governed by members of the hect-domain-containing E3 ubiquitin-protein ligase family.

scholarly journals Depletion of yeast ribosomal proteins L16 or rp59 disrupts ribosome assembly.

1990 ◽  
Vol 111 (6) ◽  
pp. 2261-2274 ◽  
Author(s):  
M Moritz ◽  
A G Paulovich ◽  
Y F Tsay ◽  
J L Woolford
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Ribosomal Proteins ◽  
Ribosomal Subunit ◽  
Ribosome Assembly ◽  
Ribosomal Rnas ◽  
Subunit Protein ◽  
40S Ribosomal Subunit ◽  
Ribosomal Subunit Protein ◽  
The Stability

Two strains of Saccharomyces cerevisiae were constructed that are conditional for synthesis of the 60S ribosomal subunit protein, L16, or the 40S ribosomal subunit protein, rp59. These strains were used to determine the effects of depriving cells of either of these ribosomal proteins on ribosome assembly and on the synthesis and stability of other ribosomal proteins and ribosomal RNAs. Termination of synthesis of either protein leads to diminished accumulation of the subunit into which it normally assembles. Depletion of L16 or rp59 has no effect on synthesis of most other ribosomal proteins or ribosomal RNAs. However, most ribosomal proteins and ribosomal RNAs that are components of the same subunit as L16 or rp59 are rapidly degraded upon depletion of L16 or rp59, presumably resulting from abortive assembly of the subunit. Depletion of L16 has no effect on the stability of most components of the 40S subunit. Conversely, termination of synthesis of rp59 has no effect on the stability of most 60S subunit components. The implications of these findings for control of ribosome assembly and the order of assembly of ribosomal proteins into the ribosome are discussed.

scholarly journals Rsp5p, a New Link between the Actin Cytoskeleton and Endocytosis in the Yeast Saccharomyces cerevisiae

2002 ◽  
Vol 22 (20) ◽  
pp. 6946-6948 ◽  
Author(s):  
Joanna Kamińska ◽  
Beata Gajewska ◽  
Anita K. Hopper ◽  
Teresa ˙Zołądek
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Actin Cytoskeleton ◽  
Cytoskeletal Proteins ◽  
Wild Type ◽  
Yeast Saccharomyces Cerevisiae ◽  
Ww Domains ◽  
Ubiquitin Protein Ligase ◽  
Growth Defects ◽  
Genes Encoding ◽  
Cytoskeleton Dynamics

ABSTRACT Rsp5p is an ubiquitin-protein ligase of Saccharomyces cerevisiae that has been implicated in numerous processes including transcription, mitochondrial inheritance, and endocytosis. Rsp5p functions at multiple steps of endocytosis, including ubiquitination of substrates and other undefined steps. We propose that one of the roles of Rsp5p in endocytosis involves maintenance and remodeling of the actin cytoskeleton. We report the following. (i) There are genetic interactions between rsp5 and several mutant genes encoding actin cytoskeletal proteins. rsp5 arp2, rsp5 end3, and rsp5 sla2 double mutants all show synthetic growth defects. Overexpressed wild-type RSP5 or mutant rsp5 genes with lesions of some WW domains suppress growth defects of arp2 and end3 cells. The defects in endocytosis, actin cytoskeleton, and morphology of arp2 are also suppressed. (ii) Rsp5p and Sla2p colocalize in abnormal F-actin-containing clumps in arp2 and pan1 mutants. Immunoprecipitation experiments confirmed that Rsp5p and Act1p colocalize in pan1 mutants. (iii) Rsp5p and Sla2p coimmunoprecipitate and partially colocalize to punctate structures in wild-type cells. These studies provide the first evidence for an interaction of an actin cytoskeleton protein with Rsp5p. (iv) rsp5-w1 mutants are resistant to latrunculin A, a drug that sequesters actin monomers and depolymerizes actin filaments, consistent with the fact that Rsp5p is involved in actin cytoskeleton dynamics.

Ribosomal protein S14 is altered by two-step emetine resistance mutations in Chinese hamster cells

1983 ◽  
Vol 3 (2) ◽  
pp. 190-197
Author(s):  
J J Madjar ◽  
M Frahm ◽  
S McGill ◽  
D J Roufa
Keyword(s):  
Ribosomal Protein ◽  
Ribosomal Proteins ◽  
Ribosomal Subunit ◽  
Chinese Hamster ◽  
Complementation Group ◽  
Resistance Mutations ◽  
Cho Cell ◽  
Ribosomal Subunits ◽  
40S Ribosomal Subunit ◽  
One Step

Four two-dimensional polyacrylamide gel electrophoresis systems were used to identify 78 Chinese hamster cell ribosomal proteins by the uniform nomenclature based on rat liver ribosomal proteins. The 40S ribosomal subunit protein affected by Chinese hamster ovary (CHO) cell one-step emetine resistance mutations is designated S14 in the standard nomenclature. To seek unambiguous genetic evidence for a cause and effect relationship between CHO cell emetine resistance and mutations in the S14 gene, we mutagenized a one-step CHO cell mutant and isolated second-step mutant clones resistant to 10-fold-higher concentrations of emetine. All of the highly resistant, two-step CHO cell mutants obtained displayed additional alterations in ribosomal protein S14. Hybridization complementation tests revealed that the two-step CHO cell emetine resistance mutants were members of the same complementation group defined by one-step CHO cell mutants, EmtB. Two-step mutants obtained from a Chinese hamster lung cell emetine-resistant clone belong to the EmtA complementation group. The two-step and EmtB mutants elaborated 40S ribosomal subunits, which dissociated to 32S and 40S core particles in buffers containing 0.5 M KCl at 4 degrees C. In contrast, 40S ribosomal subunits purified from all EmtA, one-step EmtB EmtC mutants, and wild-type CHO and lung cells were stable at this temperature in buffers containing substantially higher concentrations of salt. Thus, two-step emtB mutations affect the structure of S14 protein directly and the stability of the 40S ribosomal subunit indirectly.

scholarly journals Disruption of single-copy genes encoding acidic ribosomal proteins in Saccharomyces cerevisiae.

1990 ◽  
Vol 10 (5) ◽  
pp. 2182-2190 ◽  
Author(s):  
M Remacha ◽  
C Santos ◽  
J P Ballesta
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Ribosomal Proteins ◽  
Minimal Medium ◽  
Single Copy ◽  
Diploid Strain ◽  
Tetrad Analysis ◽  
Genes Encoding ◽  
Acidic Proteins ◽  
Dna Fragment ◽  
His3 Gene

Using the cloned genes coding for the ribosomal acidic proteins L44 and L45, constructions were made which deleted part of the coding sequence and inserted a DNA fragment at that site carrying either the URA3 or HIS3 gene. By gene disruption techniques with linearized DNA from these constructions, strains of Saccharomyces cerevisiae were obtained which lacked a functional gene for either protein L44 or protein L45. The disrupted genes in the transformants were characterized by Southern blots. The absence of the proteins was verified by electrofocusing and immunological techniques, but a compensating increase of the other acidic ribosomal proteins was not detected. The mutant lacking L44 grew at a rate identical to the parental strain in complex as well as in minimal medium. The L45-disrupted strain also grew well in both media but at a slower rate than the parental culture. A diploid strain was obtained by crossing both transformants, and by tetrad analysis it was shown that the double transformant lacking both genes is not viable. These results indicated that proteins L44 and L45 are independently dispensable for cell growth and that the ribosome is functional in the absence of either of them.

scholarly journals Mapping of the Saccharomyces cerevisiae Oxa1-Mitochondrial Ribosome Interface and Identification of MrpL40, a Ribosomal Protein in Close Proximity to Oxa1 and Critical for Oxidative Phosphorylation Complex Assembly

2009 ◽  
Vol 8 (11) ◽  
pp. 1792-1802 ◽  
Author(s):  
Lixia Jia ◽  
Jasvinder Kaur ◽  
Rosemary A. Stuart
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Oxidative Phosphorylation ◽  
Ribosomal Protein ◽  
Ribosomal Proteins ◽  
Close Association ◽  
Ribosomal Subunit ◽  
Inner Membrane ◽  
Mitochondrial Inner Membrane ◽  
Mitochondrial Ribosome ◽  
Encoded Proteins

ABSTRACT The Oxa1 protein plays a central role in facilitating the cotranslational insertion of the nascent polypeptide chains into the mitochondrial inner membrane. Mitochondrially encoded proteins are synthesized on matrix-localized ribosomes which are tethered to the inner membrane and in physical association with the Oxa1 protein. In the present study we used a chemical cross-linking approach to map the Saccharomyces cerevisiae Oxa1-ribosome interface, and we demonstrate here a close association of Oxa1 and the large ribosomal subunit protein, MrpL40. Evidence to indicate that a close physical and functional relationship exists between MrpL40 and another large ribosomal protein, the Mrp20/L23 protein, is also provided. MrpL40 shares sequence features with the bacterial ribosomal protein L24, which like Mrp20/L23 is known to be located adjacent to the ribosomal polypeptide exit site. We propose therefore that MrpL40 represents the Saccharomyces cerevisiae L24 homolog. MrpL40, like many mitochondrial ribosomal proteins, contains a C-terminal extension region that bears no similarity to the bacterial counterpart. We show that this C-terminal mitochondria-specific region is important for MrpL40's ability to support the synthesis of the correct complement of mitochondrially encoded proteins and their subsequent assembly into oxidative phosphorylation complexes.

scholarly journals Why Dom34 Stimulates Growth of Cells with Defects of 40S Ribosomal Subunit Biosynthesis

2010 ◽  
Vol 30 (23) ◽  
pp. 5562-5571 ◽  
Author(s):  
Arpita Bhattacharya ◽  
Kerri B. McIntosh ◽  
Ian M. Willis ◽  
Jonathan R. Warner
Keyword(s):  
Translation Initiation ◽  
Ribosomal Proteins ◽  
Slow Growth ◽  
Ribosomal Subunit ◽  
Ribosomal Subunits ◽  
Growth Defects ◽  
Structural Problems ◽  
Genome Wide ◽  
40S Ribosomal Subunit ◽  
Severe Shortage

ABSTRACT A set of genome-wide screens for proteins whose absence exacerbates growth defects due to pseudo-haploinsufficiency of ribosomal proteins in Saccharomyces cerevisiae identified Dom34 as being particularly important for cell growth when there is a deficit of 40S ribosomal subunits. In contrast, strains with a deficit of 60S ribosomal proteins were largely insensitive to the loss of Dom34. The slow growth of cells lacking Dom34 and haploinsufficient for a protein of the 40S subunit is caused by a severe shortage of 40S subunits available for translation initiation due to a combination of three effects: (i) the natural deficiency of 40S subunits due to defective synthesis, (ii) the sequestration of 40S subunits due to the large accumulation of free 60S subunits, and (iii) the accumulation of ribosomes “stuck” in a distinct 80S form, insensitive to the Mg2+ concentration, and at least temporarily unavailable for further translation. Our data suggest that these stuck ribosomes have neither mRNA nor tRNA. We postulate, based on our results and on previously published work, that the stuck ribosomes arise because of the lack of Dom34, which normally resolves a ribosome stalled due to insufficient tRNAs, to structural problems with its mRNA, or to a defect in the ribosome itself.

Disruption of single-copy genes encoding acidic ribosomal proteins in Saccharomyces cerevisiae

1990 ◽  
Vol 10 (5) ◽  
pp. 2182-2190
Author(s):  
M Remacha ◽  
C Santos ◽  
J P Ballesta
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Ribosomal Proteins ◽  
Minimal Medium ◽  
Single Copy ◽  
Diploid Strain ◽  
Tetrad Analysis ◽  
Genes Encoding ◽  
Acidic Proteins ◽  
Dna Fragment ◽  
His3 Gene

Using the cloned genes coding for the ribosomal acidic proteins L44 and L45, constructions were made which deleted part of the coding sequence and inserted a DNA fragment at that site carrying either the URA3 or HIS3 gene. By gene disruption techniques with linearized DNA from these constructions, strains of Saccharomyces cerevisiae were obtained which lacked a functional gene for either protein L44 or protein L45. The disrupted genes in the transformants were characterized by Southern blots. The absence of the proteins was verified by electrofocusing and immunological techniques, but a compensating increase of the other acidic ribosomal proteins was not detected. The mutant lacking L44 grew at a rate identical to the parental strain in complex as well as in minimal medium. The L45-disrupted strain also grew well in both media but at a slower rate than the parental culture. A diploid strain was obtained by crossing both transformants, and by tetrad analysis it was shown that the double transformant lacking both genes is not viable. These results indicated that proteins L44 and L45 are independently dispensable for cell growth and that the ribosome is functional in the absence of either of them.

scholarly journals Recent Advances on the Structure and Function of RNA Acetyltransferase Kre33/NAT10

Cells ◽  
2019 ◽  
Vol 8 (9) ◽  
pp. 1035 ◽  
Author(s):  
Sophie Sleiman ◽  
Francois Dragon
Keyword(s):  
Ribosomal Proteins ◽  
Ribosome Biogenesis ◽  
Ribosomal Subunit ◽  
40S Ribosomal Subunit ◽  
Functional Features ◽  
And Function ◽  
Progeria Syndrome ◽  
Hutchinson Gilford Progeria Syndrome ◽  
Nucleolar Rna

Ribosome biogenesis is one of the most energy demanding processes in the cell. In eukaryotes, the main steps of this process occur in the nucleolus and include pre-ribosomal RNA (pre-rRNA) processing, post-transcriptional modifications, and assembly of many non-ribosomal factors and ribosomal proteins in order to form mature and functional ribosomes. In yeast and humans, the nucleolar RNA acetyltransferase Kre33/NAT10 participates in different maturation events, such as acetylation and processing of 18S rRNA, and assembly of the 40S ribosomal subunit. Here, we review the structural and functional features of Kre33/NAT10 RNA acetyltransferase, and we underscore the importance of this enzyme in ribosome biogenesis, as well as in acetylation of non-ribosomal targets. We also report on the role of human NAT10 in Hutchinson–Gilford progeria syndrome.

Organization of genes encoding the L11, L1, L10, and L12 equivalent ribosomal proteins in eubacteria, archaebacteria, and eucaryotes

1989 ◽  
Vol 35 (1) ◽  
pp. 164-170 ◽  
Author(s):  
Lawrence C. Shimmin ◽  
C. Hunter Newton ◽  
Celia Ramirez ◽  
Janet Yee ◽  
Willa Lee Downing ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Saccharomyces Cerevisiae ◽  
Ribosomal Proteins ◽  
Sulfolobus Solfataricus ◽  
Mrna Translation ◽  
Restriction Fragments ◽  
E Coli ◽  
Transcription Pattern ◽  
Genes Encoding ◽  
Cytoplasmic Ribosomes

Archaebacterial and eucaryotic cytoplasmic ribosomes contain proteins equivalent to the L11, L1, L10, and L12 proteins of the eubacterium Escherichia coli. In E. coli the genes encoding these ribosomal proteins are clustered, cotranscribed, and autogenously regulated at the level of mRNA translation. Genomic restriction fragments encoding the L11e, L1e, L10e, and L12e (equivalent) proteins from two divergent archaebacteria, Halobacterium cutirubrum and Sulfolobus solfataricus, and the L10e and L12e proteins from the eucaryote Saccharomyces cerevisiae have been cloned, sequenced, and analyzed. In the archaebacteria, as in eubacteria, the four genes are clustered and the L11e, L1e, L 10e, and L12e order is maintained. The transcription pattern of the H. cutirubrum cluster is different from the E. coli pattern and the flanking genes on either side of the tetragenic clusters in E. coli, H. cutirubrum, and Sulfolobus solfataricus are all unrelated to each other. In the eucaryote Saccharomyces cerevisiae there is a single L10e gene and four separate L12e genes that are designated L12eIA, L12eIB, L12eIIA, and L12eIIB. These five genes are not closely linked and each is transcribed as a monocistronic mRNA; the L10e, L12eIA, L12eIB, and the L12eIIA genes are contiguous and uninterrupted, whereas the L12eIIB gene is interrupted by a 301 nucleotide long intron located between codons 38 and 39.Key words: archaebacteria, ribosome, Halobacterium, Sulfolobus.

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