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Arabidopsis thaliana telomeric DNA-binding protein 1 is required for telomere length homeostasis and its Myb-extension domain stabilizes plant telomeric DNA binding.

Hwang MG, Cho MH - Nucleic Acids Res. (2007)

Bottom Line: Here, we demonstrated that lack of AtTBP1 results in a deregulation of telomere length control, with mutant telomeres expanding steadily by the fourth generation.DNA-binding studies with mutant AtTBP1 proteins showed that the Myb-extension domain of AtTBP1 is required for binding to plant telomeric DNA.Our results suggest that AtTBP1 is involved in the telomere length mechanism in A. thaliana and that the Myb-extension domain of AtTBP1 may stabilize plant telomeric DNA binding.

View Article: PubMed Central - PubMed

Affiliation: Department of Biology, Yonsei University, Seoul 120-749, Republic of Korea.

ABSTRACT
Telomeres are specific protein-DNA complexes that protect the ends of eukaryotic chromosomes from fusion and degradation and are maintained by a specialized mechanism exerted by telomerase and telomere-binding proteins (TBPs), which are evolutionarily conserved. AtTBP1 is an Arabidopsis thaliana protein that binds plant telomeric DNA in vitro. Here, we demonstrated that lack of AtTBP1 results in a deregulation of telomere length control, with mutant telomeres expanding steadily by the fourth generation. DNA-binding studies with mutant AtTBP1 proteins showed that the Myb-extension domain of AtTBP1 is required for binding to plant telomeric DNA. Our results suggest that AtTBP1 is involved in the telomere length mechanism in A. thaliana and that the Myb-extension domain of AtTBP1 may stabilize plant telomeric DNA binding.

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Telomere maintenance in AtTBP1-deficient plants among different generations. (A) Telomere dynamics in AtTBP1−/− A. thaliana. Genomic DNA isolated from a pool of seedlings of wild type or mutant, at each generation, were digested with Tru9I. Telomeric lengths were determined by Southern hybridization with a telomeric probe. (B) TRF analysis of individual plants from each generation of mutant populations. The gDNAs were extracted from rosette leaves of individual plants and were digested with Tru9I. (C) Single telomere analysis, using a telomere-associated probe derived from the Ch2R. The gDNAs in (C) are the identical samples used in (B), except that these were digested with PvuII and SpeI. The arrowhead indicates undigested gDNAs. The asterisks indicate interstitial telomeric DNA.
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Figure 2: Telomere maintenance in AtTBP1-deficient plants among different generations. (A) Telomere dynamics in AtTBP1−/− A. thaliana. Genomic DNA isolated from a pool of seedlings of wild type or mutant, at each generation, were digested with Tru9I. Telomeric lengths were determined by Southern hybridization with a telomeric probe. (B) TRF analysis of individual plants from each generation of mutant populations. The gDNAs were extracted from rosette leaves of individual plants and were digested with Tru9I. (C) Single telomere analysis, using a telomere-associated probe derived from the Ch2R. The gDNAs in (C) are the identical samples used in (B), except that these were digested with PvuII and SpeI. The arrowhead indicates undigested gDNAs. The asterisks indicate interstitial telomeric DNA.

Mentions: As AtTBP1 mutant plants were visibly indistinguishable from wild-type plants, we next examined the role of AtTBP1 in telomere length homeostasis by determining telomere length in mutant plants with TRF analysis. A recent study suggested that the Ws ecotype of A. thaliana displays a bimodal size distribution, with one group bearing shorter telomeres (2–5 kb) and the other bearing longer telomeres (4–9 kb) (24). In our experiments, the wild-type telomeres ranged in size from 2.5 to 4.5 kb and were the same size as the telomeres in the short group (Figure 2A, lane 2). This short telomere size of wild-type plants was stably inherited for at least four successive generations (data not shown). In sharp contrast, TRF analysis of G2 to G4 mutant populations showed that telomeres in AtTBP1 homozygous mutants increased steadily to 10 kb after four generations (Figure 2A, lanes 3–5). Thus, telomeres in AtTBP1 mutants extended to over twice the size of those in wild-type plants in four generations.Figure 2.


Arabidopsis thaliana telomeric DNA-binding protein 1 is required for telomere length homeostasis and its Myb-extension domain stabilizes plant telomeric DNA binding.

Hwang MG, Cho MH - Nucleic Acids Res. (2007)

Telomere maintenance in AtTBP1-deficient plants among different generations. (A) Telomere dynamics in AtTBP1−/− A. thaliana. Genomic DNA isolated from a pool of seedlings of wild type or mutant, at each generation, were digested with Tru9I. Telomeric lengths were determined by Southern hybridization with a telomeric probe. (B) TRF analysis of individual plants from each generation of mutant populations. The gDNAs were extracted from rosette leaves of individual plants and were digested with Tru9I. (C) Single telomere analysis, using a telomere-associated probe derived from the Ch2R. The gDNAs in (C) are the identical samples used in (B), except that these were digested with PvuII and SpeI. The arrowhead indicates undigested gDNAs. The asterisks indicate interstitial telomeric DNA.
© Copyright Policy - openaccess
Related In: Results  -  Collection

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Figure 2: Telomere maintenance in AtTBP1-deficient plants among different generations. (A) Telomere dynamics in AtTBP1−/− A. thaliana. Genomic DNA isolated from a pool of seedlings of wild type or mutant, at each generation, were digested with Tru9I. Telomeric lengths were determined by Southern hybridization with a telomeric probe. (B) TRF analysis of individual plants from each generation of mutant populations. The gDNAs were extracted from rosette leaves of individual plants and were digested with Tru9I. (C) Single telomere analysis, using a telomere-associated probe derived from the Ch2R. The gDNAs in (C) are the identical samples used in (B), except that these were digested with PvuII and SpeI. The arrowhead indicates undigested gDNAs. The asterisks indicate interstitial telomeric DNA.
Mentions: As AtTBP1 mutant plants were visibly indistinguishable from wild-type plants, we next examined the role of AtTBP1 in telomere length homeostasis by determining telomere length in mutant plants with TRF analysis. A recent study suggested that the Ws ecotype of A. thaliana displays a bimodal size distribution, with one group bearing shorter telomeres (2–5 kb) and the other bearing longer telomeres (4–9 kb) (24). In our experiments, the wild-type telomeres ranged in size from 2.5 to 4.5 kb and were the same size as the telomeres in the short group (Figure 2A, lane 2). This short telomere size of wild-type plants was stably inherited for at least four successive generations (data not shown). In sharp contrast, TRF analysis of G2 to G4 mutant populations showed that telomeres in AtTBP1 homozygous mutants increased steadily to 10 kb after four generations (Figure 2A, lanes 3–5). Thus, telomeres in AtTBP1 mutants extended to over twice the size of those in wild-type plants in four generations.Figure 2.

Bottom Line: Here, we demonstrated that lack of AtTBP1 results in a deregulation of telomere length control, with mutant telomeres expanding steadily by the fourth generation.DNA-binding studies with mutant AtTBP1 proteins showed that the Myb-extension domain of AtTBP1 is required for binding to plant telomeric DNA.Our results suggest that AtTBP1 is involved in the telomere length mechanism in A. thaliana and that the Myb-extension domain of AtTBP1 may stabilize plant telomeric DNA binding.

View Article: PubMed Central - PubMed

Affiliation: Department of Biology, Yonsei University, Seoul 120-749, Republic of Korea.

ABSTRACT
Telomeres are specific protein-DNA complexes that protect the ends of eukaryotic chromosomes from fusion and degradation and are maintained by a specialized mechanism exerted by telomerase and telomere-binding proteins (TBPs), which are evolutionarily conserved. AtTBP1 is an Arabidopsis thaliana protein that binds plant telomeric DNA in vitro. Here, we demonstrated that lack of AtTBP1 results in a deregulation of telomere length control, with mutant telomeres expanding steadily by the fourth generation. DNA-binding studies with mutant AtTBP1 proteins showed that the Myb-extension domain of AtTBP1 is required for binding to plant telomeric DNA. Our results suggest that AtTBP1 is involved in the telomere length mechanism in A. thaliana and that the Myb-extension domain of AtTBP1 may stabilize plant telomeric DNA binding.

Show MeSH