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Global DNA Methylation patterns on marsupial and devil facial tumour chromosomes.

Ingles ED, Deakin JE - Mol Cytogenet (2015)

Bottom Line: In males, the X chromosome was hypermethylated as was one X in females.Similarly, telomeric regions on DFTD chromosomes and regions corresponding to material from one of the two X chromosomes were hypermethylated.No difference in global methylation in samples of the same strain taken in different years was observed.

View Article: PubMed Central - PubMed

Affiliation: Institute for Applied Ecology, University of Canberra, Canberra, ACT 2601 Australia.

ABSTRACT

Background: Despite DNA methylation being one of the most widely studied epigenetic modifications in eukaryotes, only a few studies have examined the global methylation status of marsupial chromosomes. The emergence of devil facial tumour disease (DFTD), a clonally transmissible cancer spreading through the Tasmanian devil population, makes it a particularly pertinent time to determine the methylation status of marsupial and devil facial tumour chromosomes. DNA methylation perturbations are known to play a role in genome instability in human tumours. One of the interesting features of the devil facial tumour is its remarkable karyotypic stability over time as only four strains with minor karyotypic differences having been reported. The cytogenetic monitoring of devil facial tumour (DFT) samples collected over an eight year period and detailed molecular cytogenetic analysis performed on the different DFT strains enables chromosome rearrangements to be correlated with methylation status as the tumour evolves.

Results: We used immunofluorescent staining with an antibody to 5-methylcytosine on metaphase chromosomes prepared from fibroblast cells of three distantly related marsupials, including the Tasmanian devil, as well as DFTD chromosomes prepared from samples collected from different years and representing different karyotypic strains. Staining of chromosomes from male and female marsupial cell lines indicate species-specific differences in global methylation patterns but with the most intense staining regions corresponding to telomeric and/or centromeric regions of autosomes. In males, the X chromosome was hypermethylated as was one X in females. Similarly, telomeric regions on DFTD chromosomes and regions corresponding to material from one of the two X chromosomes were hypermethylated. No difference in global methylation in samples of the same strain taken in different years was observed.

Conclusions: The methylation patterns on DFTD chromosomes suggests that the hypermethylated active X was shattered in the formation of the tumour chromosomes, with atypical areas of methylation on DFTD chromosomes corresponding to locations of X chromosome material from the shattered X. The incredibly stable broad methylation patterns observed between strains and over time may reflect the overall genomic stability of the devil facial tumour.

No MeSH data available.


Related in: MedlinePlus

Methylation status of metaphase chromosomes from a tammar wallaby, b devil and c opossum female fibroblast cells. Images for DAPI and 5-methylcytosine are shown separately as well as merged. X chromosomes are indicated
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Fig3: Methylation status of metaphase chromosomes from a tammar wallaby, b devil and c opossum female fibroblast cells. Images for DAPI and 5-methylcytosine are shown separately as well as merged. X chromosomes are indicated

Mentions: All tammar wallaby autosomes had intense staining for the anti-5-methylcytosine antibody in the pericentric regions and more moderate staining along the rest of the chromosome (Fig. 3a). This was consistent with the pattern previously described using a nick translation technique [3]. The telomeric/subtelomeric regions of chromosomes 1 and 3 were consistently stained, although less intensely than that in the pericentric region. Devil fibroblasts displayed strong staining for 5-methylcytosine in telomeric regions, and faint levels of staining on remaining autosomal regions (Fig. 3b). There were no detectable differences in intensity or extent of methylation in telomeric regions of homologous chromosomes. Therefore, the intensity of methylation staining using the immunofluorescence technique does not appear to correspond to the telomere length dimorphism observed in this species [6]. The opossum also had strong staining of telomeric regions on autosomes, and a lack of methylation in centromeric regions. The rest of the autosomal regions had more moderate levels of methylation (Fig. 3c). Comparisons of methylation patterns between species for autosomes with the same gene content demonstrated a methylation profile independent of gene content. This is indicated in Fig. 4 where a comparison of devil chromosome 4 with its homologue in the tammar wallaby (chromosome 2) and opossum (chromosome 2) shows that the conserved region on the distal region of the long arm of each of these chromosomes has a different methylation pattern. However, most intense methylation staining was apparent in heterochromatic regions (centromeres or telomeres).Fig. 3


Global DNA Methylation patterns on marsupial and devil facial tumour chromosomes.

Ingles ED, Deakin JE - Mol Cytogenet (2015)

Methylation status of metaphase chromosomes from a tammar wallaby, b devil and c opossum female fibroblast cells. Images for DAPI and 5-methylcytosine are shown separately as well as merged. X chromosomes are indicated
© Copyright Policy - OpenAccess
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4591559&req=5

Fig3: Methylation status of metaphase chromosomes from a tammar wallaby, b devil and c opossum female fibroblast cells. Images for DAPI and 5-methylcytosine are shown separately as well as merged. X chromosomes are indicated
Mentions: All tammar wallaby autosomes had intense staining for the anti-5-methylcytosine antibody in the pericentric regions and more moderate staining along the rest of the chromosome (Fig. 3a). This was consistent with the pattern previously described using a nick translation technique [3]. The telomeric/subtelomeric regions of chromosomes 1 and 3 were consistently stained, although less intensely than that in the pericentric region. Devil fibroblasts displayed strong staining for 5-methylcytosine in telomeric regions, and faint levels of staining on remaining autosomal regions (Fig. 3b). There were no detectable differences in intensity or extent of methylation in telomeric regions of homologous chromosomes. Therefore, the intensity of methylation staining using the immunofluorescence technique does not appear to correspond to the telomere length dimorphism observed in this species [6]. The opossum also had strong staining of telomeric regions on autosomes, and a lack of methylation in centromeric regions. The rest of the autosomal regions had more moderate levels of methylation (Fig. 3c). Comparisons of methylation patterns between species for autosomes with the same gene content demonstrated a methylation profile independent of gene content. This is indicated in Fig. 4 where a comparison of devil chromosome 4 with its homologue in the tammar wallaby (chromosome 2) and opossum (chromosome 2) shows that the conserved region on the distal region of the long arm of each of these chromosomes has a different methylation pattern. However, most intense methylation staining was apparent in heterochromatic regions (centromeres or telomeres).Fig. 3

Bottom Line: In males, the X chromosome was hypermethylated as was one X in females.Similarly, telomeric regions on DFTD chromosomes and regions corresponding to material from one of the two X chromosomes were hypermethylated.No difference in global methylation in samples of the same strain taken in different years was observed.

View Article: PubMed Central - PubMed

Affiliation: Institute for Applied Ecology, University of Canberra, Canberra, ACT 2601 Australia.

ABSTRACT

Background: Despite DNA methylation being one of the most widely studied epigenetic modifications in eukaryotes, only a few studies have examined the global methylation status of marsupial chromosomes. The emergence of devil facial tumour disease (DFTD), a clonally transmissible cancer spreading through the Tasmanian devil population, makes it a particularly pertinent time to determine the methylation status of marsupial and devil facial tumour chromosomes. DNA methylation perturbations are known to play a role in genome instability in human tumours. One of the interesting features of the devil facial tumour is its remarkable karyotypic stability over time as only four strains with minor karyotypic differences having been reported. The cytogenetic monitoring of devil facial tumour (DFT) samples collected over an eight year period and detailed molecular cytogenetic analysis performed on the different DFT strains enables chromosome rearrangements to be correlated with methylation status as the tumour evolves.

Results: We used immunofluorescent staining with an antibody to 5-methylcytosine on metaphase chromosomes prepared from fibroblast cells of three distantly related marsupials, including the Tasmanian devil, as well as DFTD chromosomes prepared from samples collected from different years and representing different karyotypic strains. Staining of chromosomes from male and female marsupial cell lines indicate species-specific differences in global methylation patterns but with the most intense staining regions corresponding to telomeric and/or centromeric regions of autosomes. In males, the X chromosome was hypermethylated as was one X in females. Similarly, telomeric regions on DFTD chromosomes and regions corresponding to material from one of the two X chromosomes were hypermethylated. No difference in global methylation in samples of the same strain taken in different years was observed.

Conclusions: The methylation patterns on DFTD chromosomes suggests that the hypermethylated active X was shattered in the formation of the tumour chromosomes, with atypical areas of methylation on DFTD chromosomes corresponding to locations of X chromosome material from the shattered X. The incredibly stable broad methylation patterns observed between strains and over time may reflect the overall genomic stability of the devil facial tumour.

No MeSH data available.


Related in: MedlinePlus