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Repressed synthesis of ribosomal proteins generates protein-specific cell cycle and morphological phenotypes.

Thapa M, Bommakanti A, Shamsuzzaman M, Gregory B, Samsel L, Zengel JM, Lindahl L - Mol. Biol. Cell (2013)

Bottom Line: We found that repression of nine 60S r-protein genes results in arrest in the G2/M phase, whereas repression of nine other 60S and 22 40S r-protein genes causes arrest in the G1 phase.Finally, repression of most r-protein genes results in changed sites of bud formation.Strikingly, the r-proteins whose repression generates similar effects on cell cycle progression cluster in the ribosome physical structure, suggesting that different topological areas of the precursor and/or mature ribosome are mechanistically connected to separate aspects of the cell cycle.

View Article: PubMed Central - PubMed

Affiliation: Department of Biological Sciences, University of Maryland, Baltimore County, Baltimore, MD 21250 Flow Cytometry Core Facility, National Heart, Lung and Blood Institute, National Institutes of Health, Bethesda, MD 20892.

ABSTRACT
The biogenesis of ribosomes is coordinated with cell growth and proliferation. Distortion of the coordinated synthesis of ribosomal components affects not only ribosome formation, but also cell fate. However, the connection between ribosome biogenesis and cell fate is not well understood. To establish a model system for inquiries into these processes, we systematically analyzed cell cycle progression, cell morphology, and bud site selection after repression of 54 individual ribosomal protein (r-protein) genes in Saccharomyces cerevisiae. We found that repression of nine 60S r-protein genes results in arrest in the G2/M phase, whereas repression of nine other 60S and 22 40S r-protein genes causes arrest in the G1 phase. Furthermore, bud morphology changes after repression of some r-protein genes. For example, very elongated buds form after repression of seven 60S r-protein genes. These genes overlap with, but are not identical to, those causing the G2/M cell cycle phenotype. Finally, repression of most r-protein genes results in changed sites of bud formation. Strikingly, the r-proteins whose repression generates similar effects on cell cycle progression cluster in the ribosome physical structure, suggesting that different topological areas of the precursor and/or mature ribosome are mechanistically connected to separate aspects of the cell cycle.

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Grouping of proteins on the model for the S. cerevisiae ribosome. (A, B) The 60S subunit proteins (rRNA omitted) viewed from the solvent side (A) or the interface side (B). (C, D) The 60S proteins (rRNA included) viewed from the solvent side (C) or interface side (D). (E, F) The 80S ribosome viewed from the solvent side of the 60S subunit (E) or the solvent side of the 40S subunit (F). (G, H) The 80S ribosome viewed from the P-stalk side (G) or the L1 stalk side (H). Proteins are color coded according to the phenotype generated in response to repression of their synthesis. Red, 60S proteins with G2/M phenotype; yellow, 60S proteins with G1 phenotype; magenta, 40S proteins with G1 phenotype; turquoise, 60S proteins with no phenotype; green, 40S proteins with no phenotype; gray, 25S, 5.8S, and 5S rRNA; blue, 18S rRNA. Models were rendered with Chimera using Protein Data Base files 3U5B, 3U5C, 3U5D, and 3U5E. Phenotypes generated upon repression of the synthesis of specific proteins are listed in Figure 1, D and F; see also Figure 1, B and C.
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Figure 5: Grouping of proteins on the model for the S. cerevisiae ribosome. (A, B) The 60S subunit proteins (rRNA omitted) viewed from the solvent side (A) or the interface side (B). (C, D) The 60S proteins (rRNA included) viewed from the solvent side (C) or interface side (D). (E, F) The 80S ribosome viewed from the solvent side of the 60S subunit (E) or the solvent side of the 40S subunit (F). (G, H) The 80S ribosome viewed from the P-stalk side (G) or the L1 stalk side (H). Proteins are color coded according to the phenotype generated in response to repression of their synthesis. Red, 60S proteins with G2/M phenotype; yellow, 60S proteins with G1 phenotype; magenta, 40S proteins with G1 phenotype; turquoise, 60S proteins with no phenotype; green, 40S proteins with no phenotype; gray, 25S, 5.8S, and 5S rRNA; blue, 18S rRNA. Models were rendered with Chimera using Protein Data Base files 3U5B, 3U5C, 3U5D, and 3U5E. Phenotypes generated upon repression of the synthesis of specific proteins are listed in Figure 1, D and F; see also Figure 1, B and C.

Mentions: In contrast to the incomplete overlap in the classifications just discussed, there is an intriguing correlation between the cell cycle phenotypes and the locations in the ribosome of the proteins whose reduced synthesis induces these phenotypes (Figure 5). Proteins linked to the G2/M response all map to the solvent side of the 60S subunit. On the other hand, proteins whose cessation of synthesis generates the G1 response are either 40S proteins or 60S proteins on the interface side, forming a ring around the 25S rRNA domain that connects to the 40S subunit. Of interest, the same subunit specificity may also apply to the few ribosomal assembly factors that have been tested for effects on the cell cycle. Cessation of the synthesis of a number of proteins from the small subunit processome results in G1 arrest (Bernstein and Baserga, 2004; Bernstein et al., 2007), whereas repression of the synthesis of the 60S-specific factor Nop15 or Rrp14 generates accumulation of cells during the late part of the cell cycle (Oeffinger and Tollervey, 2003; Oeffinger et al., 2007; Yamada et al., 2007). The differential effects of disrupting the 40S and 60S pathways may also apply to mammalian cells. Conditional deletion of the RPS6 gene in liver cells prevents the transition from G1 to S phase (Thomas et al., 2000; Fumagalli et al., 2009), but combined disruption of the synthesis of one protein from each subunit activates a G2/M checkpoint (Fumagalli et al., 2012) that is akin to our observations in yeast.


Repressed synthesis of ribosomal proteins generates protein-specific cell cycle and morphological phenotypes.

Thapa M, Bommakanti A, Shamsuzzaman M, Gregory B, Samsel L, Zengel JM, Lindahl L - Mol. Biol. Cell (2013)

Grouping of proteins on the model for the S. cerevisiae ribosome. (A, B) The 60S subunit proteins (rRNA omitted) viewed from the solvent side (A) or the interface side (B). (C, D) The 60S proteins (rRNA included) viewed from the solvent side (C) or interface side (D). (E, F) The 80S ribosome viewed from the solvent side of the 60S subunit (E) or the solvent side of the 40S subunit (F). (G, H) The 80S ribosome viewed from the P-stalk side (G) or the L1 stalk side (H). Proteins are color coded according to the phenotype generated in response to repression of their synthesis. Red, 60S proteins with G2/M phenotype; yellow, 60S proteins with G1 phenotype; magenta, 40S proteins with G1 phenotype; turquoise, 60S proteins with no phenotype; green, 40S proteins with no phenotype; gray, 25S, 5.8S, and 5S rRNA; blue, 18S rRNA. Models were rendered with Chimera using Protein Data Base files 3U5B, 3U5C, 3U5D, and 3U5E. Phenotypes generated upon repression of the synthesis of specific proteins are listed in Figure 1, D and F; see also Figure 1, B and C.
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Related In: Results  -  Collection

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Figure 5: Grouping of proteins on the model for the S. cerevisiae ribosome. (A, B) The 60S subunit proteins (rRNA omitted) viewed from the solvent side (A) or the interface side (B). (C, D) The 60S proteins (rRNA included) viewed from the solvent side (C) or interface side (D). (E, F) The 80S ribosome viewed from the solvent side of the 60S subunit (E) or the solvent side of the 40S subunit (F). (G, H) The 80S ribosome viewed from the P-stalk side (G) or the L1 stalk side (H). Proteins are color coded according to the phenotype generated in response to repression of their synthesis. Red, 60S proteins with G2/M phenotype; yellow, 60S proteins with G1 phenotype; magenta, 40S proteins with G1 phenotype; turquoise, 60S proteins with no phenotype; green, 40S proteins with no phenotype; gray, 25S, 5.8S, and 5S rRNA; blue, 18S rRNA. Models were rendered with Chimera using Protein Data Base files 3U5B, 3U5C, 3U5D, and 3U5E. Phenotypes generated upon repression of the synthesis of specific proteins are listed in Figure 1, D and F; see also Figure 1, B and C.
Mentions: In contrast to the incomplete overlap in the classifications just discussed, there is an intriguing correlation between the cell cycle phenotypes and the locations in the ribosome of the proteins whose reduced synthesis induces these phenotypes (Figure 5). Proteins linked to the G2/M response all map to the solvent side of the 60S subunit. On the other hand, proteins whose cessation of synthesis generates the G1 response are either 40S proteins or 60S proteins on the interface side, forming a ring around the 25S rRNA domain that connects to the 40S subunit. Of interest, the same subunit specificity may also apply to the few ribosomal assembly factors that have been tested for effects on the cell cycle. Cessation of the synthesis of a number of proteins from the small subunit processome results in G1 arrest (Bernstein and Baserga, 2004; Bernstein et al., 2007), whereas repression of the synthesis of the 60S-specific factor Nop15 or Rrp14 generates accumulation of cells during the late part of the cell cycle (Oeffinger and Tollervey, 2003; Oeffinger et al., 2007; Yamada et al., 2007). The differential effects of disrupting the 40S and 60S pathways may also apply to mammalian cells. Conditional deletion of the RPS6 gene in liver cells prevents the transition from G1 to S phase (Thomas et al., 2000; Fumagalli et al., 2009), but combined disruption of the synthesis of one protein from each subunit activates a G2/M checkpoint (Fumagalli et al., 2012) that is akin to our observations in yeast.

Bottom Line: We found that repression of nine 60S r-protein genes results in arrest in the G2/M phase, whereas repression of nine other 60S and 22 40S r-protein genes causes arrest in the G1 phase.Finally, repression of most r-protein genes results in changed sites of bud formation.Strikingly, the r-proteins whose repression generates similar effects on cell cycle progression cluster in the ribosome physical structure, suggesting that different topological areas of the precursor and/or mature ribosome are mechanistically connected to separate aspects of the cell cycle.

View Article: PubMed Central - PubMed

Affiliation: Department of Biological Sciences, University of Maryland, Baltimore County, Baltimore, MD 21250 Flow Cytometry Core Facility, National Heart, Lung and Blood Institute, National Institutes of Health, Bethesda, MD 20892.

ABSTRACT
The biogenesis of ribosomes is coordinated with cell growth and proliferation. Distortion of the coordinated synthesis of ribosomal components affects not only ribosome formation, but also cell fate. However, the connection between ribosome biogenesis and cell fate is not well understood. To establish a model system for inquiries into these processes, we systematically analyzed cell cycle progression, cell morphology, and bud site selection after repression of 54 individual ribosomal protein (r-protein) genes in Saccharomyces cerevisiae. We found that repression of nine 60S r-protein genes results in arrest in the G2/M phase, whereas repression of nine other 60S and 22 40S r-protein genes causes arrest in the G1 phase. Furthermore, bud morphology changes after repression of some r-protein genes. For example, very elongated buds form after repression of seven 60S r-protein genes. These genes overlap with, but are not identical to, those causing the G2/M cell cycle phenotype. Finally, repression of most r-protein genes results in changed sites of bud formation. Strikingly, the r-proteins whose repression generates similar effects on cell cycle progression cluster in the ribosome physical structure, suggesting that different topological areas of the precursor and/or mature ribosome are mechanistically connected to separate aspects of the cell cycle.

Show MeSH
Related in: MedlinePlus