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Telomere length homeostasis and telomere position effect on a linear human artificial chromosome are dictated by the genetic background.

Weuts A, Voet T, Verbeeck J, Lambrechts N, Wirix E, Schoonjans L, Danloy S, Marynen P, Froyen G - Nucleic Acids Res. (2012)

Bottom Line: Telomere position effect (TPE) is the influence of telomeres on subtelomeric epigenetic marks and gene expression.We could show consistent genetic background-dependent adaptation of telomere length and telomere-associated de novo subtelomeric DNA methylation in mouse ES-R1 cells as well as in mice.We thus provide a new tool for functional telomere studies and provide strong evidence that telomere length, subtelomeric chromatin marks and expression of subtelomeric genes are genetic background dependent.

View Article: PubMed Central - PubMed

Affiliation: Human Genome Laboratory, VIB Center for the Biology of Disease, Leuven, Belgium.

ABSTRACT
Telomere position effect (TPE) is the influence of telomeres on subtelomeric epigenetic marks and gene expression. Previous studies suggested that TPE depends on genetic background. As these analyses were performed on different chromosomes, cell types and species, it remains unclear whether TPE represents a chromosome-rather than genetic background-specific regulation. We describe the development of a Linear Human Artificial Chromosome (L-HAC) as a new tool for telomere studies. The L-HAC was generated through the Cre-loxP-mediated addition of telomere ends to an existing circular HAC (C-HAC). As it can be transferred to genetically distinct cell lines and animal models the L-HAC enables the study of TPE in an unprecedented manner. The HAC was relocated to four telomerase-positive cell lines via microcell-mediated chromosome transfer and subsequently to mice via blastocyst injection of L-HAC(+)-ES-cells. We could show consistent genetic background-dependent adaptation of telomere length and telomere-associated de novo subtelomeric DNA methylation in mouse ES-R1 cells as well as in mice. Expression of the subtelomeric neomycin gene was inversely correlated with telomere length and subtelomeric methylation. We thus provide a new tool for functional telomere studies and provide strong evidence that telomere length, subtelomeric chromatin marks and expression of subtelomeric genes are genetic background dependent.

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Overview of the transfer of the L-HAC to four different genetic backgrounds. The circular HAC originally resided in the E10B1 cells. Linearization was acquired via the co-transfection of two linear constructs pSP73-TEL08-BLAS-loxP and pSP73-TEL08-5′HPRT1-loxP with the CRE-expression plasmid pOG231. Putative L-HACs were transferred to a BALB/c 3T3/puro cell line by MMCT and subsequently to the DT40-CRE cell line, where they were structurally characterized (Supplementary Data). In addition they were relocated from the BALB/c cells to an hprt−/− CHL cell line and next to the ES-R1 cell line. The latter was applied to generate transchromosomal mice. As such the linear HAC is present in four unique genetic backgrounds (gray rectangles) that can be used to study TPE and (sub-) telomeric alteration at a single telomeric end. Selection criteria are displayed.
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gks926-F3: Overview of the transfer of the L-HAC to four different genetic backgrounds. The circular HAC originally resided in the E10B1 cells. Linearization was acquired via the co-transfection of two linear constructs pSP73-TEL08-BLAS-loxP and pSP73-TEL08-5′HPRT1-loxP with the CRE-expression plasmid pOG231. Putative L-HACs were transferred to a BALB/c 3T3/puro cell line by MMCT and subsequently to the DT40-CRE cell line, where they were structurally characterized (Supplementary Data). In addition they were relocated from the BALB/c cells to an hprt−/− CHL cell line and next to the ES-R1 cell line. The latter was applied to generate transchromosomal mice. As such the linear HAC is present in four unique genetic backgrounds (gray rectangles) that can be used to study TPE and (sub-) telomeric alteration at a single telomeric end. Selection criteria are displayed.

Mentions: To study the function of the artificial telomeres as well as the epigenetic alteration of its (sub-) telomere in different genetic backgrounds, we transferred the L-HAC from the mouse BALB/c genetic background to a chicken DT40-CRE cell line, hamster hprt−/− CHL cells and the mouse embryonic stem cell line ES-R1 by MMCT according to the scheme presented in Figure 3.Figure 3.


Telomere length homeostasis and telomere position effect on a linear human artificial chromosome are dictated by the genetic background.

Weuts A, Voet T, Verbeeck J, Lambrechts N, Wirix E, Schoonjans L, Danloy S, Marynen P, Froyen G - Nucleic Acids Res. (2012)

Overview of the transfer of the L-HAC to four different genetic backgrounds. The circular HAC originally resided in the E10B1 cells. Linearization was acquired via the co-transfection of two linear constructs pSP73-TEL08-BLAS-loxP and pSP73-TEL08-5′HPRT1-loxP with the CRE-expression plasmid pOG231. Putative L-HACs were transferred to a BALB/c 3T3/puro cell line by MMCT and subsequently to the DT40-CRE cell line, where they were structurally characterized (Supplementary Data). In addition they were relocated from the BALB/c cells to an hprt−/− CHL cell line and next to the ES-R1 cell line. The latter was applied to generate transchromosomal mice. As such the linear HAC is present in four unique genetic backgrounds (gray rectangles) that can be used to study TPE and (sub-) telomeric alteration at a single telomeric end. Selection criteria are displayed.
© Copyright Policy - creative-commons
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC3526267&req=5

gks926-F3: Overview of the transfer of the L-HAC to four different genetic backgrounds. The circular HAC originally resided in the E10B1 cells. Linearization was acquired via the co-transfection of two linear constructs pSP73-TEL08-BLAS-loxP and pSP73-TEL08-5′HPRT1-loxP with the CRE-expression plasmid pOG231. Putative L-HACs were transferred to a BALB/c 3T3/puro cell line by MMCT and subsequently to the DT40-CRE cell line, where they were structurally characterized (Supplementary Data). In addition they were relocated from the BALB/c cells to an hprt−/− CHL cell line and next to the ES-R1 cell line. The latter was applied to generate transchromosomal mice. As such the linear HAC is present in four unique genetic backgrounds (gray rectangles) that can be used to study TPE and (sub-) telomeric alteration at a single telomeric end. Selection criteria are displayed.
Mentions: To study the function of the artificial telomeres as well as the epigenetic alteration of its (sub-) telomere in different genetic backgrounds, we transferred the L-HAC from the mouse BALB/c genetic background to a chicken DT40-CRE cell line, hamster hprt−/− CHL cells and the mouse embryonic stem cell line ES-R1 by MMCT according to the scheme presented in Figure 3.Figure 3.

Bottom Line: Telomere position effect (TPE) is the influence of telomeres on subtelomeric epigenetic marks and gene expression.We could show consistent genetic background-dependent adaptation of telomere length and telomere-associated de novo subtelomeric DNA methylation in mouse ES-R1 cells as well as in mice.We thus provide a new tool for functional telomere studies and provide strong evidence that telomere length, subtelomeric chromatin marks and expression of subtelomeric genes are genetic background dependent.

View Article: PubMed Central - PubMed

Affiliation: Human Genome Laboratory, VIB Center for the Biology of Disease, Leuven, Belgium.

ABSTRACT
Telomere position effect (TPE) is the influence of telomeres on subtelomeric epigenetic marks and gene expression. Previous studies suggested that TPE depends on genetic background. As these analyses were performed on different chromosomes, cell types and species, it remains unclear whether TPE represents a chromosome-rather than genetic background-specific regulation. We describe the development of a Linear Human Artificial Chromosome (L-HAC) as a new tool for telomere studies. The L-HAC was generated through the Cre-loxP-mediated addition of telomere ends to an existing circular HAC (C-HAC). As it can be transferred to genetically distinct cell lines and animal models the L-HAC enables the study of TPE in an unprecedented manner. The HAC was relocated to four telomerase-positive cell lines via microcell-mediated chromosome transfer and subsequently to mice via blastocyst injection of L-HAC(+)-ES-cells. We could show consistent genetic background-dependent adaptation of telomere length and telomere-associated de novo subtelomeric DNA methylation in mouse ES-R1 cells as well as in mice. Expression of the subtelomeric neomycin gene was inversely correlated with telomere length and subtelomeric methylation. We thus provide a new tool for functional telomere studies and provide strong evidence that telomere length, subtelomeric chromatin marks and expression of subtelomeric genes are genetic background dependent.

Show MeSH
Related in: MedlinePlus