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Chromatin features of plant telomeric sequences at terminal vs. internal positions.

Majerová E, Mandáková T, Vu GT, Fajkus J, Lysak MA, Fojtová M - Front Plant Sci (2014)

Bottom Line: Chromatin of genuine telomeres displayed heterochromatic as well as euchromatic marks, while ITRs were just heterochromatic.Telomeric transcripts TERRA (G-rich) and ARRET (C-rich) were identified in both plants and their levels varied among tissues with a maximum in blossoms.Plants with substantially different proportions of internally and terminally located telomeric repeats are instrumental in clarifying the chromatin status of telomeric repeats at distinct chromosome locations.

View Article: PubMed Central - PubMed

Affiliation: Mendel Centre for Plant Genomics and Proteomics, Central European Institute of Technology and Faculty of Science, Masaryk University Brno, Czech Republic.

ABSTRACT
Epigenetic mechanisms are involved in regulation of crucial cellular processes in eukaryotic organisms. Data on the epigenetic features of plant telomeres and their epigenetic regulation were published mostly for Arabidopsis thaliana, in which the presence of interstitial telomeric repeats (ITRs) may interfere with genuine telomeres in most analyses. Here, we studied the epigenetic landscape and transcription of telomeres and ITRs in Nicotiana tabacum with long telomeres and no detectable ITRs, and in Ballantinia antipoda with large blocks of pericentromeric ITRs and relatively short telomeres. Chromatin of genuine telomeres displayed heterochromatic as well as euchromatic marks, while ITRs were just heterochromatic. Methylated cytosines were present at telomeres and ITRs, but showed a bias with more methylation toward distal telomere positions and different blocks of B. antipoda ITRs methylated to different levels. Telomeric transcripts TERRA (G-rich) and ARRET (C-rich) were identified in both plants and their levels varied among tissues with a maximum in blossoms. Plants with substantially different proportions of internally and terminally located telomeric repeats are instrumental in clarifying the chromatin status of telomeric repeats at distinct chromosome locations.

No MeSH data available.


Related in: MedlinePlus

Relative cytosine methylation in telomeres of Nicotiana species. (A) After Bal31 treatment of N. tabacum high molecular weight DNA, TRF analysis was used to control the efficiency of digestion. A loss of the telomere-specific hybridization signal was observed in the course of Bal31 treatment. After 15 min of Bal31 digestion loss of the signal is evident but telomere erosion is not clear, whereas after 45- and 90-min treatments, telomeres were efficiently degraded. Time of Bal31 digestion is given above the lanes. M – DNA size marker. (B) Dot-blot analysis of Bal31-digested DNA from N. tabacum and N. tomentosiformis after treatment with sodium bisulfite. Samples were loaded onto a membrane and hybridized with radioactively labeled probes to detect the total signal of telomeres (loading probe pltel-C complementary to the telomeric G-strand) and the portion of methylated telomeres (DEGENER probe). Time of the Bal31 digestion in minutes is given above the membranes. +, positive hybridization control (tobacco DNA without the bisulfite treatment); −, negative control (DNA from the pUC19 plasmid). (C) Relative density of methylated cytosines along telomeres, calculated as the DEGENER/loading hybridization signals ratio. The ratio in Bal31 non-treated samples was arbitrarily taken as 1. Six independent experiments were performed.
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Figure 3: Relative cytosine methylation in telomeres of Nicotiana species. (A) After Bal31 treatment of N. tabacum high molecular weight DNA, TRF analysis was used to control the efficiency of digestion. A loss of the telomere-specific hybridization signal was observed in the course of Bal31 treatment. After 15 min of Bal31 digestion loss of the signal is evident but telomere erosion is not clear, whereas after 45- and 90-min treatments, telomeres were efficiently degraded. Time of Bal31 digestion is given above the lanes. M – DNA size marker. (B) Dot-blot analysis of Bal31-digested DNA from N. tabacum and N. tomentosiformis after treatment with sodium bisulfite. Samples were loaded onto a membrane and hybridized with radioactively labeled probes to detect the total signal of telomeres (loading probe pltel-C complementary to the telomeric G-strand) and the portion of methylated telomeres (DEGENER probe). Time of the Bal31 digestion in minutes is given above the membranes. +, positive hybridization control (tobacco DNA without the bisulfite treatment); −, negative control (DNA from the pUC19 plasmid). (C) Relative density of methylated cytosines along telomeres, calculated as the DEGENER/loading hybridization signals ratio. The ratio in Bal31 non-treated samples was arbitrarily taken as 1. Six independent experiments were performed.

Mentions: Since telomeres in Nicotiana species are relatively long (20–160 kb in N. tabacum and 20—50 kb in N. tomentosiformis Fajkus et al., 1995a; Kovarik et al., 1996) we studied the 5-methylcytosines (5mC) distribution along these telomere repeat tracts. For this purpose, high molecular weight DNA was progressively digested with Bal31 exonuclease. Efficient degradation of telomeres in Bal31-digested samples was checked by (i) TRF analysis that showed loss of the telomere-specific hybridization signal (Figure 3A) and by (ii) hybridization using the loading pltel-C probe. Similarly as in TRF analysis, the telomere-specific signal in Bal31-digested samples was markedly reduced (Figure 3B).


Chromatin features of plant telomeric sequences at terminal vs. internal positions.

Majerová E, Mandáková T, Vu GT, Fajkus J, Lysak MA, Fojtová M - Front Plant Sci (2014)

Relative cytosine methylation in telomeres of Nicotiana species. (A) After Bal31 treatment of N. tabacum high molecular weight DNA, TRF analysis was used to control the efficiency of digestion. A loss of the telomere-specific hybridization signal was observed in the course of Bal31 treatment. After 15 min of Bal31 digestion loss of the signal is evident but telomere erosion is not clear, whereas after 45- and 90-min treatments, telomeres were efficiently degraded. Time of Bal31 digestion is given above the lanes. M – DNA size marker. (B) Dot-blot analysis of Bal31-digested DNA from N. tabacum and N. tomentosiformis after treatment with sodium bisulfite. Samples were loaded onto a membrane and hybridized with radioactively labeled probes to detect the total signal of telomeres (loading probe pltel-C complementary to the telomeric G-strand) and the portion of methylated telomeres (DEGENER probe). Time of the Bal31 digestion in minutes is given above the membranes. +, positive hybridization control (tobacco DNA without the bisulfite treatment); −, negative control (DNA from the pUC19 plasmid). (C) Relative density of methylated cytosines along telomeres, calculated as the DEGENER/loading hybridization signals ratio. The ratio in Bal31 non-treated samples was arbitrarily taken as 1. Six independent experiments were performed.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4219495&req=5

Figure 3: Relative cytosine methylation in telomeres of Nicotiana species. (A) After Bal31 treatment of N. tabacum high molecular weight DNA, TRF analysis was used to control the efficiency of digestion. A loss of the telomere-specific hybridization signal was observed in the course of Bal31 treatment. After 15 min of Bal31 digestion loss of the signal is evident but telomere erosion is not clear, whereas after 45- and 90-min treatments, telomeres were efficiently degraded. Time of Bal31 digestion is given above the lanes. M – DNA size marker. (B) Dot-blot analysis of Bal31-digested DNA from N. tabacum and N. tomentosiformis after treatment with sodium bisulfite. Samples were loaded onto a membrane and hybridized with radioactively labeled probes to detect the total signal of telomeres (loading probe pltel-C complementary to the telomeric G-strand) and the portion of methylated telomeres (DEGENER probe). Time of the Bal31 digestion in minutes is given above the membranes. +, positive hybridization control (tobacco DNA without the bisulfite treatment); −, negative control (DNA from the pUC19 plasmid). (C) Relative density of methylated cytosines along telomeres, calculated as the DEGENER/loading hybridization signals ratio. The ratio in Bal31 non-treated samples was arbitrarily taken as 1. Six independent experiments were performed.
Mentions: Since telomeres in Nicotiana species are relatively long (20–160 kb in N. tabacum and 20—50 kb in N. tomentosiformis Fajkus et al., 1995a; Kovarik et al., 1996) we studied the 5-methylcytosines (5mC) distribution along these telomere repeat tracts. For this purpose, high molecular weight DNA was progressively digested with Bal31 exonuclease. Efficient degradation of telomeres in Bal31-digested samples was checked by (i) TRF analysis that showed loss of the telomere-specific hybridization signal (Figure 3A) and by (ii) hybridization using the loading pltel-C probe. Similarly as in TRF analysis, the telomere-specific signal in Bal31-digested samples was markedly reduced (Figure 3B).

Bottom Line: Chromatin of genuine telomeres displayed heterochromatic as well as euchromatic marks, while ITRs were just heterochromatic.Telomeric transcripts TERRA (G-rich) and ARRET (C-rich) were identified in both plants and their levels varied among tissues with a maximum in blossoms.Plants with substantially different proportions of internally and terminally located telomeric repeats are instrumental in clarifying the chromatin status of telomeric repeats at distinct chromosome locations.

View Article: PubMed Central - PubMed

Affiliation: Mendel Centre for Plant Genomics and Proteomics, Central European Institute of Technology and Faculty of Science, Masaryk University Brno, Czech Republic.

ABSTRACT
Epigenetic mechanisms are involved in regulation of crucial cellular processes in eukaryotic organisms. Data on the epigenetic features of plant telomeres and their epigenetic regulation were published mostly for Arabidopsis thaliana, in which the presence of interstitial telomeric repeats (ITRs) may interfere with genuine telomeres in most analyses. Here, we studied the epigenetic landscape and transcription of telomeres and ITRs in Nicotiana tabacum with long telomeres and no detectable ITRs, and in Ballantinia antipoda with large blocks of pericentromeric ITRs and relatively short telomeres. Chromatin of genuine telomeres displayed heterochromatic as well as euchromatic marks, while ITRs were just heterochromatic. Methylated cytosines were present at telomeres and ITRs, but showed a bias with more methylation toward distal telomere positions and different blocks of B. antipoda ITRs methylated to different levels. Telomeric transcripts TERRA (G-rich) and ARRET (C-rich) were identified in both plants and their levels varied among tissues with a maximum in blossoms. Plants with substantially different proportions of internally and terminally located telomeric repeats are instrumental in clarifying the chromatin status of telomeric repeats at distinct chromosome locations.

No MeSH data available.


Related in: MedlinePlus