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Sub1 and Maf1, two effectors of RNA polymerase III, are involved in the yeast quiescence cycle.

Acker J, Nguyen NT, Vandamme M, Tavenet A, Briand-Suleau A, Conesa C - PLoS ONE (2014)

Bottom Line: Sub1 and Maf1 exert an opposite effect on RNA polymerase III transcription interfering with different steps of the transcription cycle.First, cells lacking Sub1 need more time than wild type to exit from resting and this lag in re-proliferation is correlated with a delay in transcriptional reactivation.On the other hand, we show that maf1Δ cells are long-lived mutant suggesting a connection between Pol III transcription and yeast longevity.

View Article: PubMed Central - PubMed

Affiliation: iBiTec-S CEA, FRE3377, Gif-sur-Yvette, France; CNRS, FRE3377, Gif-sur-Yvette, France; Université Paris-Sud, FRE3377, Gif-sur-Yvette, France.

ABSTRACT
Sub1 and Maf1 exert an opposite effect on RNA polymerase III transcription interfering with different steps of the transcription cycle. In this study, we present evidence that Sub1 and Maf1 also exhibit an opposite role on yeast chronological life span. First, cells lacking Sub1 need more time than wild type to exit from resting and this lag in re-proliferation is correlated with a delay in transcriptional reactivation. Second, our data show that the capacity of the cells to properly establish a quiescent state is impaired in the absence of Sub1 resulting in a premature death that is dependent on the Ras/PKA and Tor1/Sch9 signalling pathways. On the other hand, we show that maf1Δ cells are long-lived mutant suggesting a connection between Pol III transcription and yeast longevity.

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Reactivation of transcription upon exit from quiescence.(A) Day4 cultures of the indicated strains grown in SD4x were re-inoculated into fresh medium. RNA synthesis by Pol III (5S rRNA, tRNAs) or Pol I (25S, 18S, 5.8S rRNAs) was monitored over time by metabolic labelling with 3H-uracil (3H). For 25S and 18S rRNAs, a shorter exposure (SE) is shown. Total RNA loaded was visualized by staining with ethidium bromide (EtBr). (B) The expression of the 35S rRNA gene and of RPS6A, ACT1 or IMD2 genes, three of Sub1 genomic targets, in the indicated strains grown in YPD, at day5 (S) and after 1 to 6 h post-inoculation into fresh media was determined by real-time PCR. Expression levels were normalized to those of the 5S rRNA gene. Standard deviations were calculated from two independent preparations of cDNA. As expected from the repressive role of Sub1 on IMD2 gene expression, a higher expression level of IMD2 gene was detected in the absence of Sub1.
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pone-0114587-g003: Reactivation of transcription upon exit from quiescence.(A) Day4 cultures of the indicated strains grown in SD4x were re-inoculated into fresh medium. RNA synthesis by Pol III (5S rRNA, tRNAs) or Pol I (25S, 18S, 5.8S rRNAs) was monitored over time by metabolic labelling with 3H-uracil (3H). For 25S and 18S rRNAs, a shorter exposure (SE) is shown. Total RNA loaded was visualized by staining with ethidium bromide (EtBr). (B) The expression of the 35S rRNA gene and of RPS6A, ACT1 or IMD2 genes, three of Sub1 genomic targets, in the indicated strains grown in YPD, at day5 (S) and after 1 to 6 h post-inoculation into fresh media was determined by real-time PCR. Expression levels were normalized to those of the 5S rRNA gene. Standard deviations were calculated from two independent preparations of cDNA. As expected from the repressive role of Sub1 on IMD2 gene expression, a higher expression level of IMD2 gene was detected in the absence of Sub1.

Mentions: To obtain a global view of the reactivation of Pol III transcription, we next performed RNA metabolic labelling experiments that also allowed the visualization of Pol I transcription. Newly synthetized RNAs were monitored over time after the release of resting cells into fresh SD4x medium containing 3H-uracil (Fig. 3A). Although transcription is reduced but not totally abolished in stationary phase, no labelled signals could be observed in resting state probably due to the loss of uracil uptake [17]. In wild type, sub1Δ or maf1Δ strains, the reactivation of 5S rRNA or tRNAs transcription occurred within the 2 first hours post-inoculation with optimal levels of labelled signals observed at 2–3 h. The overall reactivation of Pol III transcription seemed to occur to a lesser extent in sub1Δ and maf1Δ cells as compared to wild type. At 6–8 h post inoculation, as also observed at an early stage in the growth cycle for Pol I transcription [18], Pol III transcription decreased in these three strains. In sharp contrast, the appearance of neo-synthetized Pol III transcripts was strongly delayed in the sub1Δmaf1Δ strain, with optimal levels of labelled signals reached only after 6–8 h post-inoculation, in good correlation with a longer delay in exiting quiescence (Fig. 1).


Sub1 and Maf1, two effectors of RNA polymerase III, are involved in the yeast quiescence cycle.

Acker J, Nguyen NT, Vandamme M, Tavenet A, Briand-Suleau A, Conesa C - PLoS ONE (2014)

Reactivation of transcription upon exit from quiescence.(A) Day4 cultures of the indicated strains grown in SD4x were re-inoculated into fresh medium. RNA synthesis by Pol III (5S rRNA, tRNAs) or Pol I (25S, 18S, 5.8S rRNAs) was monitored over time by metabolic labelling with 3H-uracil (3H). For 25S and 18S rRNAs, a shorter exposure (SE) is shown. Total RNA loaded was visualized by staining with ethidium bromide (EtBr). (B) The expression of the 35S rRNA gene and of RPS6A, ACT1 or IMD2 genes, three of Sub1 genomic targets, in the indicated strains grown in YPD, at day5 (S) and after 1 to 6 h post-inoculation into fresh media was determined by real-time PCR. Expression levels were normalized to those of the 5S rRNA gene. Standard deviations were calculated from two independent preparations of cDNA. As expected from the repressive role of Sub1 on IMD2 gene expression, a higher expression level of IMD2 gene was detected in the absence of Sub1.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4273968&req=5

pone-0114587-g003: Reactivation of transcription upon exit from quiescence.(A) Day4 cultures of the indicated strains grown in SD4x were re-inoculated into fresh medium. RNA synthesis by Pol III (5S rRNA, tRNAs) or Pol I (25S, 18S, 5.8S rRNAs) was monitored over time by metabolic labelling with 3H-uracil (3H). For 25S and 18S rRNAs, a shorter exposure (SE) is shown. Total RNA loaded was visualized by staining with ethidium bromide (EtBr). (B) The expression of the 35S rRNA gene and of RPS6A, ACT1 or IMD2 genes, three of Sub1 genomic targets, in the indicated strains grown in YPD, at day5 (S) and after 1 to 6 h post-inoculation into fresh media was determined by real-time PCR. Expression levels were normalized to those of the 5S rRNA gene. Standard deviations were calculated from two independent preparations of cDNA. As expected from the repressive role of Sub1 on IMD2 gene expression, a higher expression level of IMD2 gene was detected in the absence of Sub1.
Mentions: To obtain a global view of the reactivation of Pol III transcription, we next performed RNA metabolic labelling experiments that also allowed the visualization of Pol I transcription. Newly synthetized RNAs were monitored over time after the release of resting cells into fresh SD4x medium containing 3H-uracil (Fig. 3A). Although transcription is reduced but not totally abolished in stationary phase, no labelled signals could be observed in resting state probably due to the loss of uracil uptake [17]. In wild type, sub1Δ or maf1Δ strains, the reactivation of 5S rRNA or tRNAs transcription occurred within the 2 first hours post-inoculation with optimal levels of labelled signals observed at 2–3 h. The overall reactivation of Pol III transcription seemed to occur to a lesser extent in sub1Δ and maf1Δ cells as compared to wild type. At 6–8 h post inoculation, as also observed at an early stage in the growth cycle for Pol I transcription [18], Pol III transcription decreased in these three strains. In sharp contrast, the appearance of neo-synthetized Pol III transcripts was strongly delayed in the sub1Δmaf1Δ strain, with optimal levels of labelled signals reached only after 6–8 h post-inoculation, in good correlation with a longer delay in exiting quiescence (Fig. 1).

Bottom Line: Sub1 and Maf1 exert an opposite effect on RNA polymerase III transcription interfering with different steps of the transcription cycle.First, cells lacking Sub1 need more time than wild type to exit from resting and this lag in re-proliferation is correlated with a delay in transcriptional reactivation.On the other hand, we show that maf1Δ cells are long-lived mutant suggesting a connection between Pol III transcription and yeast longevity.

View Article: PubMed Central - PubMed

Affiliation: iBiTec-S CEA, FRE3377, Gif-sur-Yvette, France; CNRS, FRE3377, Gif-sur-Yvette, France; Université Paris-Sud, FRE3377, Gif-sur-Yvette, France.

ABSTRACT
Sub1 and Maf1 exert an opposite effect on RNA polymerase III transcription interfering with different steps of the transcription cycle. In this study, we present evidence that Sub1 and Maf1 also exhibit an opposite role on yeast chronological life span. First, cells lacking Sub1 need more time than wild type to exit from resting and this lag in re-proliferation is correlated with a delay in transcriptional reactivation. Second, our data show that the capacity of the cells to properly establish a quiescent state is impaired in the absence of Sub1 resulting in a premature death that is dependent on the Ras/PKA and Tor1/Sch9 signalling pathways. On the other hand, we show that maf1Δ cells are long-lived mutant suggesting a connection between Pol III transcription and yeast longevity.

Show MeSH
Related in: MedlinePlus