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The deacetylase Sir2 from the yeast Clavispora lusitaniae lacks the evolutionarily conserved capacity to generate subtelomeric heterochromatin.

Froyd CA, Kapoor S, Dietrich F, Rusche LN - PLoS Genet. (2013)

Bottom Line: However, we found no evidence that ClHst1 associates with subtelomeric regions or impacts gene expression directly.After subsequent species diversification, the SIR2 paralog was apparently lost in the C. lusitaniae lineage.Thus, C. lusitaniae presents an opportunity to discover how subtelomeric chromatin can be reconfigured.

View Article: PubMed Central - PubMed

Affiliation: Biochemistry Department, Duke University, Durham, North Carolina, United States of America.

ABSTRACT
Deacetylases of the Sir2 or sirtuin family are thought to regulate life cycle progression and life span in response to nutrient availability. This family has undergone successive rounds of duplication and diversification, enabling the enzymes to perform a wide variety of biological functions. Two evolutionarily conserved functions of yeast Sir2 proteins are the generation of repressive chromatin in subtelomeric domains and the suppression of unbalanced recombination within the tandem rDNA array. Here, we describe the function of the Sir2 ortholog ClHst1 in the yeast Clavispora lusitaniae, an occasional opportunistic pathogen. ClHst1 was localized to the non-transcribed spacer regions of the rDNA repeats and deacetylated histones at these loci, indicating that, like other Sir2 proteins, ClHst1 modulates chromatin structure at the rDNA repeats. However, we found no evidence that ClHst1 associates with subtelomeric regions or impacts gene expression directly. This surprising observation highlights the plasticity of sirtuin function. Related yeast species, including Candida albicans, possess an additional Sir2 family member. Thus, it is likely that the ancestral Candida SIR2/HST1 gene was duplicated and subfunctionalized, such that HST1 retained the capacity to regulate rDNA whereas SIR2 had other functions, perhaps including the generation of subtelomeric chromatin. After subsequent species diversification, the SIR2 paralog was apparently lost in the C. lusitaniae lineage. Thus, C. lusitaniae presents an opportunity to discover how subtelomeric chromatin can be reconfigured.

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Related in: MedlinePlus

Telomere repeats are located at ends of chromosomes.(A) Features at the ends of five supercontigs are illustrated. Telomere repeats are represented by gray boxes. Restriction sites are marked with their distance from the telomere repeat, and the distance between sites is indicated in italics. Probes used for Southern analysis are shown as black rectangles. (B) Digested genomic DNA (LRY2826) was separated electrophoretically, transferred to membranes, and hybridized with the indicated probes located near telomere repeat sequences.
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pgen-1003935-g004: Telomere repeats are located at ends of chromosomes.(A) Features at the ends of five supercontigs are illustrated. Telomere repeats are represented by gray boxes. Restriction sites are marked with their distance from the telomere repeat, and the distance between sites is indicated in italics. Probes used for Southern analysis are shown as black rectangles. (B) Digested genomic DNA (LRY2826) was separated electrophoretically, transferred to membranes, and hybridized with the indicated probes located near telomere repeat sequences.

Mentions: To confirm that the identified telomere repeat sequences are indeed located at the ends of chromosomes, Southern blot analysis was performed. Genomic DNA was digested separately with three restriction enzymes that cut 0.5–2.5 kb from each telomere repeat sequence (Figure 4A). The cut DNA was separated electrophoretically, transferred to membranes, and analyzed using probes located just inside the telomere repeat. If the identified sequences are indeed at the ends of the chromosomes, the three digestions should yield bands whose sizes differ by the distances between the restriction sites. In contrast, if the repeats are actually internal sequences and additional unknown sequence occurs beyond the repeats, the sizes of the restriction products will be inconsistent with the prediction. For telomere repeats 2L, 4R, 7L, and 7R, the observed restriction fragments were consistent with the repeat sequence being at the end of the chromosome (Figure 4B). In all four cases, the telomere repeat length can be deduced to be 500–600 bp. Putative telomere 3R was not examined because it is associated with an rDNA array (Figure 1A). For contig 5L, multiple bands were observed, some of which differed in length by the expected amounts. However, these restriction fragments were longer than expected, suggesting that there is approximately 500 bp of additional unknown sequence between the restriction sites and the chromosome end. Nevertheless, four bona fide telomeres were identified.


The deacetylase Sir2 from the yeast Clavispora lusitaniae lacks the evolutionarily conserved capacity to generate subtelomeric heterochromatin.

Froyd CA, Kapoor S, Dietrich F, Rusche LN - PLoS Genet. (2013)

Telomere repeats are located at ends of chromosomes.(A) Features at the ends of five supercontigs are illustrated. Telomere repeats are represented by gray boxes. Restriction sites are marked with their distance from the telomere repeat, and the distance between sites is indicated in italics. Probes used for Southern analysis are shown as black rectangles. (B) Digested genomic DNA (LRY2826) was separated electrophoretically, transferred to membranes, and hybridized with the indicated probes located near telomere repeat sequences.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3814328&req=5

pgen-1003935-g004: Telomere repeats are located at ends of chromosomes.(A) Features at the ends of five supercontigs are illustrated. Telomere repeats are represented by gray boxes. Restriction sites are marked with their distance from the telomere repeat, and the distance between sites is indicated in italics. Probes used for Southern analysis are shown as black rectangles. (B) Digested genomic DNA (LRY2826) was separated electrophoretically, transferred to membranes, and hybridized with the indicated probes located near telomere repeat sequences.
Mentions: To confirm that the identified telomere repeat sequences are indeed located at the ends of chromosomes, Southern blot analysis was performed. Genomic DNA was digested separately with three restriction enzymes that cut 0.5–2.5 kb from each telomere repeat sequence (Figure 4A). The cut DNA was separated electrophoretically, transferred to membranes, and analyzed using probes located just inside the telomere repeat. If the identified sequences are indeed at the ends of the chromosomes, the three digestions should yield bands whose sizes differ by the distances between the restriction sites. In contrast, if the repeats are actually internal sequences and additional unknown sequence occurs beyond the repeats, the sizes of the restriction products will be inconsistent with the prediction. For telomere repeats 2L, 4R, 7L, and 7R, the observed restriction fragments were consistent with the repeat sequence being at the end of the chromosome (Figure 4B). In all four cases, the telomere repeat length can be deduced to be 500–600 bp. Putative telomere 3R was not examined because it is associated with an rDNA array (Figure 1A). For contig 5L, multiple bands were observed, some of which differed in length by the expected amounts. However, these restriction fragments were longer than expected, suggesting that there is approximately 500 bp of additional unknown sequence between the restriction sites and the chromosome end. Nevertheless, four bona fide telomeres were identified.

Bottom Line: However, we found no evidence that ClHst1 associates with subtelomeric regions or impacts gene expression directly.After subsequent species diversification, the SIR2 paralog was apparently lost in the C. lusitaniae lineage.Thus, C. lusitaniae presents an opportunity to discover how subtelomeric chromatin can be reconfigured.

View Article: PubMed Central - PubMed

Affiliation: Biochemistry Department, Duke University, Durham, North Carolina, United States of America.

ABSTRACT
Deacetylases of the Sir2 or sirtuin family are thought to regulate life cycle progression and life span in response to nutrient availability. This family has undergone successive rounds of duplication and diversification, enabling the enzymes to perform a wide variety of biological functions. Two evolutionarily conserved functions of yeast Sir2 proteins are the generation of repressive chromatin in subtelomeric domains and the suppression of unbalanced recombination within the tandem rDNA array. Here, we describe the function of the Sir2 ortholog ClHst1 in the yeast Clavispora lusitaniae, an occasional opportunistic pathogen. ClHst1 was localized to the non-transcribed spacer regions of the rDNA repeats and deacetylated histones at these loci, indicating that, like other Sir2 proteins, ClHst1 modulates chromatin structure at the rDNA repeats. However, we found no evidence that ClHst1 associates with subtelomeric regions or impacts gene expression directly. This surprising observation highlights the plasticity of sirtuin function. Related yeast species, including Candida albicans, possess an additional Sir2 family member. Thus, it is likely that the ancestral Candida SIR2/HST1 gene was duplicated and subfunctionalized, such that HST1 retained the capacity to regulate rDNA whereas SIR2 had other functions, perhaps including the generation of subtelomeric chromatin. After subsequent species diversification, the SIR2 paralog was apparently lost in the C. lusitaniae lineage. Thus, C. lusitaniae presents an opportunity to discover how subtelomeric chromatin can be reconfigured.

Show MeSH
Related in: MedlinePlus