Dysfunction of chromatin assembly factor 1 in FASCIATA mutants (fas) of Arabidopsis thaliana results in progressive loss of telomeric DNA. Although replicative telomere shortening is typically associated with incomplete resynthesis of their ends by telomerase, no change in telomerase activity could be detected in vitro in extracts from fas mutants. Besides a possible telomerase malfunction, the telomere shortening in fas mutants could presumably be due to problems with conventional replication of telomeres. To distinguish between the possible contribution of suboptimal function of telomerase in fas mutants under in vivo conditions and problems in conventional telomere replication, we crossed fas and tert (telomerase reverse transcriptase) knockout mutants and analyzed telomere shortening in segregated fas mutants, tert mutants, and double fas tert mutants in parallel. We demonstrate that fas tert knockouts show greater replicative telomere shortening than that observed even in the complete absence of telomerase (tert mutants). While the effect of tert and fas mutations on telomere lengths in double mutants is additive, manifestations of telomere dysfunction in double fas tert mutants (frequency of anaphase bridges, onset of chromosome end fusions, and common involvement of 45S rDNA in chromosome fusion sites) are similar to those in tert mutants. We conclude that in addition to possible impairment of telomerase action, a further mechanism contributes to telomere shortening in fas mutants.

• MeSH
• Arabidopsis enzymology   genetics   metabolism   MeSH
• Chromosomes, Plant genetics   metabolism   MeSH
• Chromatin Assembly Factor-1 genetics   metabolism   MeSH
• Mutation * MeSH
• Arabidopsis Proteins genetics   metabolism   MeSH
• RNA Splicing Factors MeSH
• Telomerase genetics   metabolism   MeSH
• Telomere genetics   metabolism   MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Names of Substances
• At2g20020 protein, Arabidopsis MeSH Browser
• Chromatin Assembly Factor-1 MeSH
• FAS protein, Arabidopsis MeSH Browser
• Arabidopsis Proteins MeSH
• RNA Splicing Factors MeSH
• Telomerase MeSH
• TERT protein, Arabidopsis MeSH Browser

BACKGROUND: Telomeres, as elaborate nucleo-protein complexes, ensure chromosomal stability. When impaired, the ends of linear chromosomes can be recognised by cellular repair mechanisms as double-strand DNA breaks and can be healed by non-homologous-end-joining activities to produce dicentric chromosomes. During cell divisions, particularly during anaphase, dicentrics can break, thus producing naked chromosome tips susceptible to additional unwanted chromosome fusion. Many telomere-building protein complexes are associated with telomeres to ensure their proper capping function. It has been found however, that a number of repair complexes also contribute to telomere stability. RESULTS: We used Arabidopsis thaliana to study the possible functions of the DNA repair subunit, NBS1, in telomere homeostasis using knockout nbs1 mutants. The results showed that although NBS1-deficient plants were viable, lacked any sign of developmental aberration and produced fertile seeds through many generations upon self-fertilisation, plants also missing the functional telomerase (double mutants), rapidly, within three generations, displayed severe developmental defects. Cytogenetic inspection of cycling somatic cells revealed a very early onset of massive genome instability. Molecular methods used for examining the length of telomeres in double homozygous mutants detected much faster telomere shortening than in plants deficient in telomerase gene alone. CONCLUSIONS: Our findings suggest that NBS1 acts in concert with telomerase and plays a profound role in plant telomere renewal.

• MeSH
• Anaphase MeSH
• Arabidopsis cytology   enzymology   genetics   growth & development   MeSH
• Chromosomal Instability MeSH
• Chromosomes, Plant genetics   metabolism   MeSH
• Cytogenetic Analysis MeSH
• DNA-Binding Proteins genetics   metabolism   MeSH
• Telomere Homeostasis MeSH
• MRE11 Homologue Protein MeSH
• In Situ Hybridization, Fluorescence MeSH
• Nuclear Proteins genetics   metabolism   MeSH
• Germination MeSH
• Flowers cytology   genetics   metabolism   MeSH
• Protein Interaction Mapping MeSH
• Meiosis MeSH
• DNA Repair MeSH
• Cell Cycle Proteins genetics   metabolism   MeSH
• Arabidopsis Proteins genetics   metabolism   MeSH
• Plant Cells enzymology   metabolism   MeSH
• Self-Fertilization MeSH
• Seeds genetics   growth & development   metabolism   MeSH
• Telomerase genetics   metabolism   MeSH
• Telomere genetics   metabolism   MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Names of Substances
• DNA-Binding Proteins MeSH
• MRE11 Homologue Protein MeSH
• Nuclear Proteins MeSH
• Mre11 protein, Arabidopsis MeSH Browser
• Cell Cycle Proteins MeSH
• Arabidopsis Proteins MeSH
• rad50 protein, Arabidopsis MeSH Browser
• Telomerase MeSH
• TERT protein, Arabidopsis MeSH Browser

Telomeres in many eukaryotes are maintained by telomerase in whose absence telomere shortening occurs. However, telomerase-deficient Arabidopsis thaliana mutants (Attert (-/-)) show extremely low rates of telomere shortening per plant generation (250-500 bp), which does not correspond to the expected outcome of replicative telomere shortening resulting from ca. 1,000 meristem cell divisions per seed-to-seed generation. To investigate the influence of the number of cell divisions per seed-to-seed generation, Attert (-/-) mutant plants were propagated from seeds coming either from the lower-most or the upper-most siliques (L- and U-plants) and the length of their telomeres were followed over several generations. The rate of telomere shortening was faster in U-plants, than in L-plants, as would be expected from their higher number of cell divisions per generation. However, this trend was observed only in telomeres whose initial length is relatively high and the differences decreased with progressive general telomere shortening over generations. But in generation 4, the L-plants frequently show a net telomere elongation, while the U-plants fail to do so. We propose that this is due to the activation of alternative telomere lengthening (ALT), a process which is activated in early embryonic development in both U- and L-plants, but is overridden in U-plants due to their higher number of cell divisions per generation. These data demonstrate what so far has only been speculated, that in the absence of telomerase, the number of cell divisions within one generation influences the control of telomere lengths. These results also reveal a fast and efficient activation of ALT mechanism(s) in response to the loss of telomerase activity and imply that ALT is probably involved also in normal plant development.

• MeSH
• Arabidopsis enzymology   genetics   growth & development   MeSH
• Cell Division genetics   physiology   MeSH
• Chromosomes, Plant genetics   MeSH
• Mutation * MeSH
• Seeds enzymology   genetics   growth & development   MeSH
• Telomerase genetics   metabolism   MeSH
• Telomere genetics   metabolism   MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Names of Substances
• Telomerase 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
• Chromosomes, Plant genetics   metabolism   MeSH
• Phylogeny MeSH
• Genome, Plant * MeSH
• Minisatellite Repeats MeSH
• Evolution, Molecular * MeSH
• Telomere-Binding Proteins physiology   MeSH
• Repetitive Sequences, Nucleic Acid MeSH
• Retroelements MeSH
• Telomere genetics   physiology   MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Review MeSH
• Comparative Study MeSH
• Names of Substances
• Telomere-Binding Proteins MeSH
• Retroelements MeSH

Silene latifolia is a key plant model in the study of sex determination and sex chromosome evolution. Current studies have been based on genetic mapping of the sequences linked to sex chromosomes with analysis of their characters and relative positions on the X and Y chromosomes. Until recently, very few DNA sequences have been physically mapped to the sex chromosomes of S. latifolia. We have carried out multicolor fluorescent in situ hybridization (FISH) analysis of S. latifolia chromosomes based on the presence and intensity of FISH signals on individual chromosomes. We have generated new markers by constructing and screening a sample bacterial artificial chromosome (BAC) library for appropriate FISH probes. Five newly isolated BAC clones yielded discrete signals on the chromosomes: two were specific for one autosome pair and three hybridized preferentially to the sex chromosomes. We present the FISH hybridization patterns of these five BAC inserts together with previously described repetitive sequences (X-43.1, 25S rDNA and 5S rDNA) and use them to analyze the S. latifolia karyotype. The autosomes of S. latifolia are difficult to distinguish based on their relative arm lengths. Using one BAC insert and the three repetitive sequences, we have constructed a standard FISH karyotype that can be used to distinguish all autosome pairs. We also analyze the hybridization patterns of these sequences on the sex chromosomes and discuss the utility of the karyotype mapping strategy presented to study sex chromosome evolution and Y chromosome degeneration.

• MeSH
• Chromosomes, Plant genetics   MeSH
• Genetic Markers genetics   MeSH
• In Situ Hybridization, Fluorescence MeSH
• Karyotyping MeSH
• Chromosome Mapping * MeSH
• Sex Chromosomes genetics   MeSH
• Repetitive Sequences, Nucleic Acid genetics   MeSH
• Silene genetics   MeSH
• Blotting, Southern MeSH
• Chromosomes, Artificial, Bacterial MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Comparative Study MeSH
• Names of Substances
• Genetic Markers MeSH