Limits...
The heterochromatic chromosome caps in great apes impact telomere metabolism.

Novo C, Arnoult N, Bordes WY, Castro-Vega L, Gibaud A, Dutrillaux B, Bacchetti S, Londoño-Vallejo A - Nucleic Acids Res. (2013)

Bottom Line: In contrast with the limited sequence divergence accumulated after separation of higher primate lineages, marked cytogenetic variation has been associated with the genome evolution in these species.Studying the impact of such structural variations on defined molecular processes can provide valuable insights on how genome structural organization contributes to organismal evolution.In gorilla, on the other hand, a proportion of the subtelomeric heterochromatic caps present in most chromosome arms are associated with large blocks of telomere-like sequences that follow a replication program different from that of bona fide telomeres.

View Article: PubMed Central - PubMed

Affiliation: Telomeres and Cancer laboratory, 'Equipe Labellisée Ligue contre le Cancer', UMR3244, Institut Curie, 26 rue d'Ulm, 75248 Paris, France.

ABSTRACT
In contrast with the limited sequence divergence accumulated after separation of higher primate lineages, marked cytogenetic variation has been associated with the genome evolution in these species. Studying the impact of such structural variations on defined molecular processes can provide valuable insights on how genome structural organization contributes to organismal evolution. Here, we show that telomeres on chromosome arms carrying subtelomeric heterochromatic caps in the chimpanzee, which are completely absent in humans, replicate later than telomeres on chromosome arms without caps. In gorilla, on the other hand, a proportion of the subtelomeric heterochromatic caps present in most chromosome arms are associated with large blocks of telomere-like sequences that follow a replication program different from that of bona fide telomeres. Strikingly, telomere-containing RNA accumulates extrachromosomally in gorilla mitotic cells, suggesting that at least some aspects of telomere-containing RNA biogenesis have diverged in gorilla, perhaps in concert with the evolution of heterochromatic caps in this species.

Show MeSH

Related in: MedlinePlus

TERRA accumulates in metaphases of siXRN2-treated gorilla cells. Q-FISH analyses show an accumulation of extrachromosomal telomeric signals (A) and an increase in the mean intensity of total telomere signals (B) revealed by the C-rich probe in siXRN2-treated cells. This increase is eliminated by an RNase treatment of the slides, indicating that this increase is due to TERRA accumulation. Numbers above bars indicate P-values (student t-test). On the other hand, the Q-FISH analyses also revealed the presence of telomeric doublets (sister chromatids bearing two telomeric signals, yellow arrows) in control cells, as well as in siXRN2-treated cells (C). In the latter, however, doublets were significantly more abundant, suggesting that many of these doublets were the consequence of TERRA accumulation (D). This conclusion is also supported by the fact that an RNase I treatment of the slides leads to a significant reduction of the doublets (D). At least 600 chromosomes were considered per condition. Numbers above bars indicate P-values (student t-test). N.S., no significant difference.
© Copyright Policy - creative-commons
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC3643582&req=5

gkt169-F5: TERRA accumulates in metaphases of siXRN2-treated gorilla cells. Q-FISH analyses show an accumulation of extrachromosomal telomeric signals (A) and an increase in the mean intensity of total telomere signals (B) revealed by the C-rich probe in siXRN2-treated cells. This increase is eliminated by an RNase treatment of the slides, indicating that this increase is due to TERRA accumulation. Numbers above bars indicate P-values (student t-test). On the other hand, the Q-FISH analyses also revealed the presence of telomeric doublets (sister chromatids bearing two telomeric signals, yellow arrows) in control cells, as well as in siXRN2-treated cells (C). In the latter, however, doublets were significantly more abundant, suggesting that many of these doublets were the consequence of TERRA accumulation (D). This conclusion is also supported by the fact that an RNase I treatment of the slides leads to a significant reduction of the doublets (D). At least 600 chromosomes were considered per condition. Numbers above bars indicate P-values (student t-test). N.S., no significant difference.

Mentions: It has been shown in the budding yeast that the RNA exonuclease Rat1P controls the levels of yeast TERRA by inducing its degradation (32). Although the role of the Rat1P-homologue XRN2 (5′–3′ exoribonuclease 2) in controlling TERRA levels remains to be demonstrated in higher primates, we tested whether XRN2 was involved in TERRA regulation in gorilla cells. Using an siRNA approach, we achieved a partial expression knockdown of the gorilla XRN2 gene (60% as determined by RT–PCR, Figure 4A) after 48 h of treatment. Strikingly, this decrease in the expression of XRN2 was associated with an increase in the detection of TERRA molecules by northern blot (Figure 4B). Remarkably, an increase in extrachromosomal TERRA molecules was also detected during mitosis, leading to a significant increase in the total fluorescence intensity associated with metaphases from cells treated with siXRN2 when compared with cells treated with an irrelevant siRNA (Figure 5A and B). This increase completely disappeared when metaphase preparations were treated with RNase (Figure 5A and B). Although most TERRA seemed to accumulate extrachromosomally, it is possible that TERRA also accumulated in association with chromosome ends, as suggested both by the fact that the proportion of sister chromatids bearing telomeric doublets was increased in siXRN2-treated cells and by the fact that this increase was lost on RNase treatment (Figure 5C and D). At the same time, as there was not detectable difference in the overall intensity of telomeres after RNase treatment (Figure 5D), telomere replication was likely not affected.Figure 4.


The heterochromatic chromosome caps in great apes impact telomere metabolism.

Novo C, Arnoult N, Bordes WY, Castro-Vega L, Gibaud A, Dutrillaux B, Bacchetti S, Londoño-Vallejo A - Nucleic Acids Res. (2013)

TERRA accumulates in metaphases of siXRN2-treated gorilla cells. Q-FISH analyses show an accumulation of extrachromosomal telomeric signals (A) and an increase in the mean intensity of total telomere signals (B) revealed by the C-rich probe in siXRN2-treated cells. This increase is eliminated by an RNase treatment of the slides, indicating that this increase is due to TERRA accumulation. Numbers above bars indicate P-values (student t-test). On the other hand, the Q-FISH analyses also revealed the presence of telomeric doublets (sister chromatids bearing two telomeric signals, yellow arrows) in control cells, as well as in siXRN2-treated cells (C). In the latter, however, doublets were significantly more abundant, suggesting that many of these doublets were the consequence of TERRA accumulation (D). This conclusion is also supported by the fact that an RNase I treatment of the slides leads to a significant reduction of the doublets (D). At least 600 chromosomes were considered per condition. Numbers above bars indicate P-values (student t-test). N.S., no significant difference.
© Copyright Policy - creative-commons
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3643582&req=5

gkt169-F5: TERRA accumulates in metaphases of siXRN2-treated gorilla cells. Q-FISH analyses show an accumulation of extrachromosomal telomeric signals (A) and an increase in the mean intensity of total telomere signals (B) revealed by the C-rich probe in siXRN2-treated cells. This increase is eliminated by an RNase treatment of the slides, indicating that this increase is due to TERRA accumulation. Numbers above bars indicate P-values (student t-test). On the other hand, the Q-FISH analyses also revealed the presence of telomeric doublets (sister chromatids bearing two telomeric signals, yellow arrows) in control cells, as well as in siXRN2-treated cells (C). In the latter, however, doublets were significantly more abundant, suggesting that many of these doublets were the consequence of TERRA accumulation (D). This conclusion is also supported by the fact that an RNase I treatment of the slides leads to a significant reduction of the doublets (D). At least 600 chromosomes were considered per condition. Numbers above bars indicate P-values (student t-test). N.S., no significant difference.
Mentions: It has been shown in the budding yeast that the RNA exonuclease Rat1P controls the levels of yeast TERRA by inducing its degradation (32). Although the role of the Rat1P-homologue XRN2 (5′–3′ exoribonuclease 2) in controlling TERRA levels remains to be demonstrated in higher primates, we tested whether XRN2 was involved in TERRA regulation in gorilla cells. Using an siRNA approach, we achieved a partial expression knockdown of the gorilla XRN2 gene (60% as determined by RT–PCR, Figure 4A) after 48 h of treatment. Strikingly, this decrease in the expression of XRN2 was associated with an increase in the detection of TERRA molecules by northern blot (Figure 4B). Remarkably, an increase in extrachromosomal TERRA molecules was also detected during mitosis, leading to a significant increase in the total fluorescence intensity associated with metaphases from cells treated with siXRN2 when compared with cells treated with an irrelevant siRNA (Figure 5A and B). This increase completely disappeared when metaphase preparations were treated with RNase (Figure 5A and B). Although most TERRA seemed to accumulate extrachromosomally, it is possible that TERRA also accumulated in association with chromosome ends, as suggested both by the fact that the proportion of sister chromatids bearing telomeric doublets was increased in siXRN2-treated cells and by the fact that this increase was lost on RNase treatment (Figure 5C and D). At the same time, as there was not detectable difference in the overall intensity of telomeres after RNase treatment (Figure 5D), telomere replication was likely not affected.Figure 4.

Bottom Line: In contrast with the limited sequence divergence accumulated after separation of higher primate lineages, marked cytogenetic variation has been associated with the genome evolution in these species.Studying the impact of such structural variations on defined molecular processes can provide valuable insights on how genome structural organization contributes to organismal evolution.In gorilla, on the other hand, a proportion of the subtelomeric heterochromatic caps present in most chromosome arms are associated with large blocks of telomere-like sequences that follow a replication program different from that of bona fide telomeres.

View Article: PubMed Central - PubMed

Affiliation: Telomeres and Cancer laboratory, 'Equipe Labellisée Ligue contre le Cancer', UMR3244, Institut Curie, 26 rue d'Ulm, 75248 Paris, France.

ABSTRACT
In contrast with the limited sequence divergence accumulated after separation of higher primate lineages, marked cytogenetic variation has been associated with the genome evolution in these species. Studying the impact of such structural variations on defined molecular processes can provide valuable insights on how genome structural organization contributes to organismal evolution. Here, we show that telomeres on chromosome arms carrying subtelomeric heterochromatic caps in the chimpanzee, which are completely absent in humans, replicate later than telomeres on chromosome arms without caps. In gorilla, on the other hand, a proportion of the subtelomeric heterochromatic caps present in most chromosome arms are associated with large blocks of telomere-like sequences that follow a replication program different from that of bona fide telomeres. Strikingly, telomere-containing RNA accumulates extrachromosomally in gorilla mitotic cells, suggesting that at least some aspects of telomere-containing RNA biogenesis have diverged in gorilla, perhaps in concert with the evolution of heterochromatic caps in this species.

Show MeSH
Related in: MedlinePlus