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Bookmarking promoters in mitotic chromatin: poly(ADP-ribose)polymerase-1 as an epigenetic mark.

Lodhi N, Kossenkov AV, Tulin AV - Nucleic Acids Res. (2014)

Bottom Line: Epigenetics are the heritable changes in gene expression or cellular phenotype caused by mechanisms other than changes in the underlying DNA sequence.After mitosis, it is thought that bookmarking transcription factors remain at promoters, regulating which genes become active and which remain silent.Herein, we demonstrate that poly(ADP-ribose)polymerase-1 (PARP-1) is a genome-wide epigenetic memory mark in mitotic chromatin, and we further show that the presence of PARP-1 is absolutely crucial for reactivation of transcription after mitosis.

View Article: PubMed Central - PubMed

Affiliation: Fox Chase Cancer Center, Philadelphia, PA, 19111 USA.

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PARP-1 binding sites co-occur with NFATC2 binding consensus. (A) Consensus binding motif found in proximity to PARP-1 binding sites. The most significant motifs associated with PARP-1 binding sites in mitotic and asynchronized cells were compared to known NFATC2 binding motif. (B) NFATC2 remains associated with chromatin during mitosis. Equal amounts of total chromatin proteins extracted from asynchronous and mitosis-arrested cells were analyzed by PAGE and western blot using anti-NFATC2, anti-PARP-1, anti-RNAP (RNA polymerase II), anti-H3 and anti-phospho-Ser10-H3 (marker of mitotic chromatin) antibodies. CF, chromatin fraction; N-CF, non-chromatin fraction. (C) NFATC2 binds to TGGAAA sequence in three randomly picked PARP-1-positive loci (OR51B2, ZSCAN4, RRAS2) and did not bind to a PARP-1-negative locus (TMCC2). Blue bars indicate NFATC2 binding in mitosis, red—in interphase. (D) The locations of predicted NFATC2 binding (blue line) sites relative to PARP-1 binding sites in mitosis (M) and interphase (A) within OR51B2, ZSCAN4, RRAS2 and TMCC2 loci. Arrows indicate positions of oligonucleotide primers around NFATC2 consensus sequence, which were used in the ChIP assay. Position of PARP-1 binding signal is shown in green for interphase and in red for mitosis. The distance between the PARP-1 binding site and NFATC2 consensus sequence is shown above each figure. Red arrow indicates the location and direction of transcription within each locus.
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Figure 4: PARP-1 binding sites co-occur with NFATC2 binding consensus. (A) Consensus binding motif found in proximity to PARP-1 binding sites. The most significant motifs associated with PARP-1 binding sites in mitotic and asynchronized cells were compared to known NFATC2 binding motif. (B) NFATC2 remains associated with chromatin during mitosis. Equal amounts of total chromatin proteins extracted from asynchronous and mitosis-arrested cells were analyzed by PAGE and western blot using anti-NFATC2, anti-PARP-1, anti-RNAP (RNA polymerase II), anti-H3 and anti-phospho-Ser10-H3 (marker of mitotic chromatin) antibodies. CF, chromatin fraction; N-CF, non-chromatin fraction. (C) NFATC2 binds to TGGAAA sequence in three randomly picked PARP-1-positive loci (OR51B2, ZSCAN4, RRAS2) and did not bind to a PARP-1-negative locus (TMCC2). Blue bars indicate NFATC2 binding in mitosis, red—in interphase. (D) The locations of predicted NFATC2 binding (blue line) sites relative to PARP-1 binding sites in mitosis (M) and interphase (A) within OR51B2, ZSCAN4, RRAS2 and TMCC2 loci. Arrows indicate positions of oligonucleotide primers around NFATC2 consensus sequence, which were used in the ChIP assay. Position of PARP-1 binding signal is shown in green for interphase and in red for mitosis. The distance between the PARP-1 binding site and NFATC2 consensus sequence is shown above each figure. Red arrow indicates the location and direction of transcription within each locus.

Mentions: To address which specific genes are marked by PARP-1 during mitosis, we performed consensus sequence comparison for the mitotic and interphase sites of PARP-1 occupancy. This analysis revealed GGAAAGG consensus sequences in proximity to approximately 65% of all sites of PARP-1 localization in mitosis and interphase (Figure 4A and Supplementary Figure S6). This sequence is similar to the known binding sites of transcriptional factor NFATC2 (51). NFATC2 serves as a transcriptional factor controlling inducible expression of interleukin gene family (52,53). Recent research has shown that transcription factors from NFAT family act as regulators of oncogenic transformation (54,55). NFATC2 remains bound to chromatin during interphase, as does PARP-1 (Figure 4B). We determined NFATC2 occupancy at a randomly selected group of PARP-1-positive peaks. Indeed, three tested PARP-1-positive loci OR51B2, ZSCAN4 and RRAS2 demonstrated significant occupancy by NFATC2, while the PARP-1 negative locus TMCC2 showed no NFATC2 binding in either mitosis or interphase (Figure 4C–D). Genome-wide co-occurrence of PARP-1 and NFATC2 binding sites suggests that genes marked by PARP-1 may share a common mechanism of regulation by NFATC2. This hypothesis is supported by earlier reports that NFATC2 directly interacts with PARP-1 and PARP-1 mediated poly(ADP-ribosyl)ation (56,57).


Bookmarking promoters in mitotic chromatin: poly(ADP-ribose)polymerase-1 as an epigenetic mark.

Lodhi N, Kossenkov AV, Tulin AV - Nucleic Acids Res. (2014)

PARP-1 binding sites co-occur with NFATC2 binding consensus. (A) Consensus binding motif found in proximity to PARP-1 binding sites. The most significant motifs associated with PARP-1 binding sites in mitotic and asynchronized cells were compared to known NFATC2 binding motif. (B) NFATC2 remains associated with chromatin during mitosis. Equal amounts of total chromatin proteins extracted from asynchronous and mitosis-arrested cells were analyzed by PAGE and western blot using anti-NFATC2, anti-PARP-1, anti-RNAP (RNA polymerase II), anti-H3 and anti-phospho-Ser10-H3 (marker of mitotic chromatin) antibodies. CF, chromatin fraction; N-CF, non-chromatin fraction. (C) NFATC2 binds to TGGAAA sequence in three randomly picked PARP-1-positive loci (OR51B2, ZSCAN4, RRAS2) and did not bind to a PARP-1-negative locus (TMCC2). Blue bars indicate NFATC2 binding in mitosis, red—in interphase. (D) The locations of predicted NFATC2 binding (blue line) sites relative to PARP-1 binding sites in mitosis (M) and interphase (A) within OR51B2, ZSCAN4, RRAS2 and TMCC2 loci. Arrows indicate positions of oligonucleotide primers around NFATC2 consensus sequence, which were used in the ChIP assay. Position of PARP-1 binding signal is shown in green for interphase and in red for mitosis. The distance between the PARP-1 binding site and NFATC2 consensus sequence is shown above each figure. Red arrow indicates the location and direction of transcription within each locus.
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Figure 4: PARP-1 binding sites co-occur with NFATC2 binding consensus. (A) Consensus binding motif found in proximity to PARP-1 binding sites. The most significant motifs associated with PARP-1 binding sites in mitotic and asynchronized cells were compared to known NFATC2 binding motif. (B) NFATC2 remains associated with chromatin during mitosis. Equal amounts of total chromatin proteins extracted from asynchronous and mitosis-arrested cells were analyzed by PAGE and western blot using anti-NFATC2, anti-PARP-1, anti-RNAP (RNA polymerase II), anti-H3 and anti-phospho-Ser10-H3 (marker of mitotic chromatin) antibodies. CF, chromatin fraction; N-CF, non-chromatin fraction. (C) NFATC2 binds to TGGAAA sequence in three randomly picked PARP-1-positive loci (OR51B2, ZSCAN4, RRAS2) and did not bind to a PARP-1-negative locus (TMCC2). Blue bars indicate NFATC2 binding in mitosis, red—in interphase. (D) The locations of predicted NFATC2 binding (blue line) sites relative to PARP-1 binding sites in mitosis (M) and interphase (A) within OR51B2, ZSCAN4, RRAS2 and TMCC2 loci. Arrows indicate positions of oligonucleotide primers around NFATC2 consensus sequence, which were used in the ChIP assay. Position of PARP-1 binding signal is shown in green for interphase and in red for mitosis. The distance between the PARP-1 binding site and NFATC2 consensus sequence is shown above each figure. Red arrow indicates the location and direction of transcription within each locus.
Mentions: To address which specific genes are marked by PARP-1 during mitosis, we performed consensus sequence comparison for the mitotic and interphase sites of PARP-1 occupancy. This analysis revealed GGAAAGG consensus sequences in proximity to approximately 65% of all sites of PARP-1 localization in mitosis and interphase (Figure 4A and Supplementary Figure S6). This sequence is similar to the known binding sites of transcriptional factor NFATC2 (51). NFATC2 serves as a transcriptional factor controlling inducible expression of interleukin gene family (52,53). Recent research has shown that transcription factors from NFAT family act as regulators of oncogenic transformation (54,55). NFATC2 remains bound to chromatin during interphase, as does PARP-1 (Figure 4B). We determined NFATC2 occupancy at a randomly selected group of PARP-1-positive peaks. Indeed, three tested PARP-1-positive loci OR51B2, ZSCAN4 and RRAS2 demonstrated significant occupancy by NFATC2, while the PARP-1 negative locus TMCC2 showed no NFATC2 binding in either mitosis or interphase (Figure 4C–D). Genome-wide co-occurrence of PARP-1 and NFATC2 binding sites suggests that genes marked by PARP-1 may share a common mechanism of regulation by NFATC2. This hypothesis is supported by earlier reports that NFATC2 directly interacts with PARP-1 and PARP-1 mediated poly(ADP-ribosyl)ation (56,57).

Bottom Line: Epigenetics are the heritable changes in gene expression or cellular phenotype caused by mechanisms other than changes in the underlying DNA sequence.After mitosis, it is thought that bookmarking transcription factors remain at promoters, regulating which genes become active and which remain silent.Herein, we demonstrate that poly(ADP-ribose)polymerase-1 (PARP-1) is a genome-wide epigenetic memory mark in mitotic chromatin, and we further show that the presence of PARP-1 is absolutely crucial for reactivation of transcription after mitosis.

View Article: PubMed Central - PubMed

Affiliation: Fox Chase Cancer Center, Philadelphia, PA, 19111 USA.

Show MeSH
Related in: MedlinePlus