Telomere homeostasis is regulated at multiple levels, including the local chromatin structure of telomeres and subtelomeres. Recent reports demonstrated that a decrease in repressive chromatin marks, such as levels of cytosine methylation in subtelomeric regions, results in telomere elongation in mouse cells. Here we show that a considerable fraction of cytosines is methylated not only in subtelomeric, but also in telomeric DNA of tobacco BY-2 cells. Drug-induced hypomethylation (demonstrated at subtelomeric, telomeric, and global DNA levels) results in activation of telomerase. However, in contrast to mouse cells, the decrease in 5-methylcytosine levels and upregulation of telomerase do not result in any changes of telomere lengths. These results demonstrate the involvement of epigenetic mechanisms in the multilevel process of regulation of telomerase activity in plant cells and, at the same time, they indicate that changes in telomerase activity can be overridden by other factors governing telomere length stability.

• MeSH
• adenin analogy a deriváty   farmakologie   MeSH
• aktivace enzymů účinky léků   MeSH
• cytidin analogy a deriváty   farmakologie   MeSH
• DNA rostlinná chemie   účinky léků   MeSH
• epigeneze genetická MeSH
• genetická transkripce účinky léků   MeSH
• kultivované buňky MeSH
• metylace DNA účinky léků   MeSH
• nukleozomy účinky léků   fyziologie   MeSH
• rostlinné proteiny genetika   metabolismus   MeSH
• tabák cytologie   účinky léků   genetika   metabolismus   MeSH
• telomerasa metabolismus   MeSH
• telomery chemie   účinky léků   metabolismus   MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• 9-(2,3-dihydroxypropyl)adenine MeSH Prohlížeč
• adenin MeSH
• cytidin MeSH
• DNA rostlinná MeSH
• nukleozomy MeSH
• pyrimidin-2-one beta-ribofuranoside MeSH Prohlížeč
• rostlinné proteiny MeSH
• telomerasa MeSH

This paper examines telomeres from an evolutionary perspective. In the monocot plant order Asparagales two evolutionary switch-points in telomere sequence are known. The first occurred when the Arabidopsis-type telomere was replaced by a telomere based on a repeat motif more typical of vertebrates. The replacement is associated with telomerase activity, but the telomerase has low fidelity and this may have implications for the binding of telomeric proteins. At the second evolutionary switch-point, the telomere and its mode of synthesis are replaced by an unknown mechanism. Elsewhere in plants (Sessia, Vestia, Cestrum) and in arthropods, the telomere "typical" of the group is lost. Probably many other groups with "unusual" telomeres will be found. We question whether telomerase is indeed the original end-maintenance system and point to other candidate processes involving t-loops, t-circles, rolling circle replication and recombination. Possible evolutionary outcomes arising from the loss of telomerase activity in alternative lengthening of telomere (ALT) systems are discussed. We propose that elongation of minisatellite repeats using recombination/replication processes initially substitutes for the loss of telomerase function. Then in more established ALT groups, subtelomeric satellite repeats may replace the telomeric minisatellite repeat whilst maintaining the recombination/replication mechanisms for telomere elongation. Thereafter a retrotransposition-based end-maintenance system may become established. The influence of changing sequence motifs on the properties of the telomere cap is discussed. The DNA and protein components of telomeres should be regarded--as with any other chromosome elements--as evolving and co-evolving over time and responding to changes in the genome and to environmental stresses. We describe how telomere dysfunction, resulting in end-to-end chromosome fusions, can have a profound effect on chromosome evolution and perhaps even speciation.

• MeSH
• chromozomy rostlin genetika   metabolismus   MeSH
• fylogeneze MeSH
• genom rostlinný * MeSH
• minisatelitní repetice MeSH
• molekulární evoluce * MeSH
• proteiny vázající telomery fyziologie   MeSH
• repetitivní sekvence nukleových kyselin MeSH
• retroelementy MeSH
• telomery genetika   fyziologie   MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• přehledy MeSH
• srovnávací studie MeSH
• Názvy látek
• proteiny vázající telomery MeSH
• retroelementy MeSH

Telomere-associated regions represent boundaries between the relatively homogeneous telomeres and the subtelomeres, which show much greater heterogeneity in chromatin structure and DNA composition. Although a major fraction of subtelomeres is usually formed by a limited number of highly repeated DNA sequence families, their mutual arrangement, attachment to telomeres and the presence of interspersed unique or low-copy-number sequences make these terminal domains chromosome specific. In this study, we describe the structures of junctions between telomeres and a major subtelomeric repeat of the plant Silene latifolia, X43.1. Our results show that on individual chromosome arms, X43.1 is attached to the telomere either directly at sites corresponding to nucleosome boundaries previously mapped in this sequence, or via other spacer sequences, both previously characterized and newly described ones. Sites of telomere junctions are non-random in all the telomere-associated sequences analysed. These data obtained at the molecular level have been verified using in situ hybridization to metaphase chromosomes and extended DNA fibres.

• MeSH
• chromatin chemie   genetika   MeSH
• chromozom X MeSH
• chromozomy rostlin MeSH
• DNA rostlinná genetika   MeSH
• genetická variace MeSH
• hybridizace in situ fluorescenční MeSH
• koncové repetice genetika   MeSH
• molekulární sekvence - údaje MeSH
• nukleozomy chemie   genetika   MeSH
• sekvence nukleotidů MeSH
• sekvenční homologie nukleových kyselin MeSH
• Silene genetika   MeSH
• tandemové repetitivní sekvence genetika   MeSH
• telomery genetika   MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• chromatin MeSH
• DNA rostlinná MeSH
• nukleozomy MeSH

The manner of packing of the terminal DNA loci into nucleosomes and higher order structures may strongly influence their functional interactions. Besides the structural flexibility of telomeric DNA sequences, conserved features of their chromatin including short nucleosome phasing (157 bp) and nucleosome sliding have been described previously. To gain a complementary knowledge of subtelomeres, we have analysed the chromatin structure of two subtelomeric tandem repeats from the plant Silene latifolia: X43.1 and 15Ssp. X43.1 shows two distinct nucleosome periodicities--157 and 188 bp. Preferred positions of its two nucleosomes have been mapped at both low and high resolution and the experimental results correspond to computer-predicted positions. 15Ssp is a newly-discovered sequence showing a telomere-associated position by PCR and a subtelomeric location by pulsed-field gel electrophoresis and fluorescence in situ hybridisation. Its 159 bp sequence unit shows a tandem arrangement and the presence of micrococcal nuclease-hypersensitive sites when either naked DNA or chromatin is digested. Use of a chemical nuclease results in a regular nucleosome ladder of 157 bp periodicity. Moreover, 15Ssp mononucleosomes show instability and absence of specific positioning, features typical for telomeric chromatin.

• MeSH
• DNA rostlinná chemie   genetika   metabolismus   MeSH
• genetické matrice MeSH
• heterochromatin chemie   genetika   metabolismus   MeSH
• hybridizace in situ fluorescenční MeSH
• konformace nukleové kyseliny * MeSH
• Magnoliopsida genetika   MeSH
• mikrokoková nukleasa metabolismus   MeSH
• molekulární sekvence - údaje MeSH
• nukleozomy chemie   genetika   metabolismus   MeSH
• restrikční mapování MeSH
• sekvence nukleotidů MeSH
• sekvenční analýza DNA MeSH
• Southernův blotting MeSH
• tandemové repetitivní sekvence genetika   MeSH
• telomery genetika   MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• DNA rostlinná MeSH
• heterochromatin MeSH
• mikrokoková nukleasa MeSH
• nukleozomy MeSH

Melandrium album (syn. Silene latifolia) belongs to dioecious plant species possessing heteromorphic sex chromosomes, X and Y. Our previous experiments using in situ nick translation and replication kinetics analysis indicated structural and functional differences between the two X chromosomes in homogametic female cells. Here we show DNA methylation patterns of M. album root tip chromosomes using the indirect immunofluorescence approach with a monoclonal antibody raised against 5-methylcytosine (5-mC). In male cells, a more intensive 5-mC labelling on the shorter arm of the only X chromosome was observed in comparison with the longer X arm. A global hypermethylation of the male Y chromosome was not found, which indicates its prevalent euchromatic character. In female cells, the specific 5-mC pattern of the X chromosome was found on a single X chromosome, whereas the other X displayed an overall higher level of 5-mC labelling. Application of a hypomethylating drug, 5-azacytidine (5-azaC), during seed germination led to a deletion of any specific differences in the 5-mC distribution between the two X chromosomes. Confocal laser scanning microscopy analysis of DNA methylation in interphase nuclei showed hypermethylated domains that were efficiently decondensed and hypomethylated by 5-azaC treatment. The presented data show reproducible differences in the DNA methylation patterns between the two X chromosomes in M. album female somatic cells, which indicate their distinct transcriptional activities as a possible consequence of the negative dosage compensation of X-linked genes.

• MeSH
• azacytidin farmakologie   MeSH
• chromatin metabolismus   MeSH
• chromozom X * MeSH
• chromozom Y * MeSH
• fluorescenční protilátková technika nepřímá MeSH
• konfokální mikroskopie metody   MeSH
• metylace DNA * MeSH
• rostliny genetika   MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• azacytidin MeSH
• chromatin MeSH

A single-strand-specific chemical probe, potassium permanganate (KMnO4), was used to study the sequence-dependent conformation periodicity of tandem multicopy repetitive DNA sequences HRS60 and GRS (Nicotiana Species) at the level of single base pair and dinucleotide step. Local DNA structures, sensitive to KMnO4, revealed periodicity of 182 +/- 2 bp, equal to the length of repeat units. Permanganate-sensitive local structures were mapped to both DNA strands of genomic HRS60 sequences and were found to be linked to d(A)n tracts. These adenine tracts are located in the proximity of the intrinsically curved domains. Distamycin A increased reactivity of the DNA but decreased the specificity of DNA cleavage. Similar conformation periodicity has been detected also in the 'canrep' family of repeats (Brassica species). All studied repetitive sequences are predominantly located in the constitutive heterochromatin. We discuss the role of conformation periodicities in relation to a structural code for nucleosome phasing at tandem arrays of DNA repeats.

• MeSH
• Brassica genetika   MeSH
• distamyciny MeSH
• DNA primery genetika   MeSH
• DNA rostlinná chemie   genetika   MeSH
• jedovaté rostliny MeSH
• konformace nukleové kyseliny * MeSH
• manganistan draselný MeSH
• mapování chromozomů MeSH
• molekulární sekvence - údaje MeSH
• molekulární sondy MeSH
• polymorfismus genetický MeSH
• repetitivní sekvence nukleových kyselin * MeSH
• sekvence nukleotidů MeSH
• tabák genetika   MeSH
• vazebná místa MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• distamyciny MeSH
• DNA primery MeSH
• DNA rostlinná MeSH
• manganistan draselný MeSH
• molekulární sondy MeSH
• stallimycin MeSH Prohlížeč

We have recently shown that hypomethylation of cytosine residues in the HRS60 family of repetitive DNA sequences can be induced with 5-azacytidine (5-azaC) in tobacco tissue cultures. We have also proven that such a DNA methylation status is maintained during the recovery of protoplasts, plant regeneration, and vegetative development. In the present paper we follow meiotic transmission of hypomethylated HRS60 DNA. Plants obtained from seeds treated with 5-azaC were either self pollinated or crossed with a non-treated control in a reciprocal way. Analysis of the methylation status of the HRS60 DNA revealed that these sequences were hypomethylated in the progenies up to the extent found in the parental 5-azaC-treated plant. Since no parent-of-origin effect was observed, we presume that both male and female gametes transmit an artificial methylation imprint to a similar extent. This result is supported by methylcytosine evaluation in the total genomic DNA samples. A temporal analysis of 5-azaC effects on germinating seeds and a phenotypic evaluation of 5-azaC-treated tobacco plants are also presented.

• Publikační typ
• časopisecké články MeSH

We have examined the structure and chromatin organization of telomeres in Nicotiana tabacum. In tobacco the blocks of simple telomeric repeats (TT-TAGGG)n are many times larger than in other plants, e.g., Arabidopsis thaliana or tomato. They are resolved as multiple fragments 60-160 kb in size (in most cases 90-130 kb) on pulsed-field gel electrophoresis (PFGE) of restriction endonuclease-digested DNA. The major subtelomeric repeat of the HRS60 family forms large homogeneous blocks of a basic 180 bp motif having comparable lengths. Micrococcal nuclease (MNase) cleaves tobacco telomeric chromatin into subunits with a short repeat length of 157 +/- 5 bp; the subtelomeric heterochromatin characterized by tandemly repeated sequences of the HRS60 family is cut by MNase with a 180 bp periodicity. The monomeric and dimeric particles of telomeric and subtelomeric chromatin differ in sensitivity to MNase treatment: the telomeric particles are readily digested, producing ladders with a periodicity of 7 bp, while the subtelomeric particles appear to be rather resistant to intranucleosomal cleavage. The results presented show apparent similarities in the organization of telomeric chromatin in higher plants and mammals.

• MeSH
• chromatin genetika   MeSH
• DNA rostlinná analýza   genetika   MeSH
• jedovaté rostliny * MeSH
• repetitivní sekvence nukleových kyselin MeSH
• tabák genetika   MeSH
• telomery genetika   MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• chromatin MeSH
• DNA rostlinná MeSH

Members of a new family of highly repetitive DNA sequences called GRS were isolated from Nicotiana tabacum L. genomic DNA and characterized. Cloned, sequenced monomeric units (180-182 bp) of GRS exhibit properties characteristic of molecules that possess a stable curvature. The GRS family represents about 0.15% of total genomic DNA (10(4) copies per haploid genome) and could be derived from either Nicotiana tomentosiformis or Nicotiana otophora, two possible ancestors of the T genome of the amphidiploid N. tabacum. Sequence homology between the HRS60 (Koukalová et al. 1989) and the GRS family has been estimated to be 57%. In situ hybridization was used to localize GRS on mitotic chromosomes. Hybridization signals were obtained on five pairs of chromosomes at intercalary sites of the longer chromosome arms. The majority of GRS sequences appeared to be organized in tandem arrays and a minority were found to be dispersed through the genome in short clusters, interspersed with other types of DNA repeats, including 25S rDNA sequences. Several loci containing both GRS and HRS60 were also found. Such hybrid loci may indicate intergenomic transfer of the DNA in the amphidiploid N. tabacum. GRS sequences, like HRS60 (Fajkus et al. 1992), were found to specify the location of nucleosomes. The position of the nucleosome core has been mapped with respect to a conserved Mbol site in the GRS sequence and an oligo A/T tract is a major centre of the DNA curvature.

• MeSH
• chromatin MeSH
• DNA rostlinná analýza   genetika   metabolismus   MeSH
• druhová specificita MeSH
• genom rostlinný MeSH
• jedovaté rostliny * MeSH
• klonování DNA MeSH
• metylace MeSH
• molekulární sekvence - údaje MeSH
• nukleozomy MeSH
• polymorfismus délky restrikčních fragmentů MeSH
• repetitivní sekvence nukleových kyselin genetika   MeSH
• sekvence nukleotidů MeSH
• sekvenční analýza DNA MeSH
• sekvenční seřazení MeSH
• tabák genetika   MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• chromatin MeSH
• DNA rostlinná MeSH
• nukleozomy MeSH