Limits...
SATB1 Mediates Long-Range Chromatin Interactions: A Dual Regulator of Anti-Apoptotic BCL2 and Pro-Apoptotic NOXA Genes.

Yang Y, Wang Z, Sun L, Shao L, Yang N, Yu D, Zhang X, Han X, Sun Y - PLoS ONE (2015)

Bottom Line: Decreased SATB1 levels switched the mbr-BCL2 loop to mbr-NOXA loop, and thus changed expression of these two genes.The SATB1-mediated dynamic switch of the chromatin loop structures was essential for the cooperative expression of the BCL2 and NOXA genes in apoptosis.Notably, the role of SATB1 was specific, since inhibition of SATB1 degradation by caspase-6 inhibitor or caspase-6-resistant SATB1 mutant reversed expression of BCL-2 and NOXA in response to apoptotic stimulation.

View Article: PubMed Central - PubMed

Affiliation: Key Laboratory of Human Functional Genomics of Jiangsu Province, Nanjing medical University, Nanjing, PR China; Department of Cell Biology, Nanjing Medical University, Nanjing, PR China.

ABSTRACT
Aberrant expression of special AT-rich binding protein 1 (SATB1), a global genomic organizer, has been associated with various cancers, which raises the question of how higher-order chromatin structure contributes to carcinogenesis. Disruption of apoptosis is one of the hallmarks of cancer. We previously demonstrated that SATB1 mediated specific long-range chromosomal interactions between the mbr enhancer located within 3'-UTR of the BCL2 gene and the promoter to regulate BCL2 expression during early apoptosis. In the present study, we used chromosome conformation capture (3C) assays and molecular analyses to further investigate the function of the SATB1-mediated higher-order chromatin structure in co-regulation of the anti-apoptotic BCL2 gene and the pro-apoptotic NOXA gene located 3.4Mb downstream on Chromosome 18. We demonstrated that the mbr enhancer spatially juxtaposed the promoters of BCL2 and NOXA genes through SATB1-mediated chromatin-loop in Jurkat cells. Decreased SATB1 levels switched the mbr-BCL2 loop to mbr-NOXA loop, and thus changed expression of these two genes. The SATB1-mediated dynamic switch of the chromatin loop structures was essential for the cooperative expression of the BCL2 and NOXA genes in apoptosis. Notably, the role of SATB1 was specific, since inhibition of SATB1 degradation by caspase-6 inhibitor or caspase-6-resistant SATB1 mutant reversed expression of BCL-2 and NOXA in response to apoptotic stimulation. This study reveals the critical role of SATB1-organized higher-order chromatin structure in regulating the dynamic equilibrium of apoptosis-controlling genes with antagonistic functions and suggests that aberrant SATB1 expression might contribute to cancer development by disrupting the co-regulated genes in apoptosis pathways.

No MeSH data available.


Related in: MedlinePlus

ChIP analysis of SATB1 binding on the NOXA promoter region and the mbr element in vivo.Diagram shows the relative positions of three SATB1 binding sites(N-SBS1、N-SBS2、N-SBS3) in NOXA promoter region and mbr(A). The results showed that NOXA-SBS1 sequence located 2.4kb relative to the translational start site were specifically immunoprecipitated with anti-SATB1 (B, indicating that SATB1 binds to this sequence in vivo. The SATB1 binding to SBS2 was very weak (C. No SATB1 binding to SBS3 was detected (D. As confirmed earlier, SATB1 bound to the mbr (E). The input represented 1% of total DNA used in immunoprecipitation. Non-specific IgG was used as a negative control. The PCR products were visualized by ethidium bromide staining of a 1.5% agarose gel.
© Copyright Policy
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC4589335&req=5

pone.0139170.g001: ChIP analysis of SATB1 binding on the NOXA promoter region and the mbr element in vivo.Diagram shows the relative positions of three SATB1 binding sites(N-SBS1、N-SBS2、N-SBS3) in NOXA promoter region and mbr(A). The results showed that NOXA-SBS1 sequence located 2.4kb relative to the translational start site were specifically immunoprecipitated with anti-SATB1 (B, indicating that SATB1 binds to this sequence in vivo. The SATB1 binding to SBS2 was very weak (C. No SATB1 binding to SBS3 was detected (D. As confirmed earlier, SATB1 bound to the mbr (E). The input represented 1% of total DNA used in immunoprecipitation. Non-specific IgG was used as a negative control. The PCR products were visualized by ethidium bromide staining of a 1.5% agarose gel.

Mentions: We have demonstrated that the SATB1-mediated long-range chromatin interaction between the mbr enhancer and BCL2 promoter is critical to regulating BCL2 expression in the cellular apoptotic response [25]. To investigate whether SATB1-mediated long-range chromatin interaction is involved in co-regulation of anti-apoptotic and pro-apoptotic genes, we used Genomatix Software (Genomatix Software, http://www.genomatix.de/index.html) to analyze the promoter regions of Bcl2 family members for the binding sites of SATB1). The NOXA gene located 3.4Mb downstream of the BCL2 gene was of particular interest. NOXA has 3 sequences that are possible SATB1 binding sites. These sequences are NOXA-SBS1 to NOXA-SBS3, and are located within the 3.5kb region upstream of the NOXA transcription start site. The illustrations of three SATB1 binding sites in NOXA promoter have been shown in Fig 1. We used ChIP assays with the anti-SATB1 antibody to test SATB1 binding on the NOXA promoter. NOXA-SBS1 sequence located 2.4kb relative to the translational start site were found to be specifically immunoprecipitated with anti-SATB1 (Fig 2), indicating that SATB1 binds to this sequence in vivo. The precipitated products were weak for SBS2. No specific precipitate was detected for SBS3 (Fig 2). As expected and confirmed in our previous work, SATB1 also bound to the mbr enhancer (Fig 2). Given that SATB1 is a critical factor in regulating long-range interactions between the mbr and BCL2 promoter [25], the binding of SATB1 to the NOXA promoter strongly suggested that SATB1 not only mediated an interaction between the mbr and the BCL2 promoter, but also was involved in the mbr-NOXA chromatin interaction.


SATB1 Mediates Long-Range Chromatin Interactions: A Dual Regulator of Anti-Apoptotic BCL2 and Pro-Apoptotic NOXA Genes.

Yang Y, Wang Z, Sun L, Shao L, Yang N, Yu D, Zhang X, Han X, Sun Y - PLoS ONE (2015)

ChIP analysis of SATB1 binding on the NOXA promoter region and the mbr element in vivo.Diagram shows the relative positions of three SATB1 binding sites(N-SBS1、N-SBS2、N-SBS3) in NOXA promoter region and mbr(A). The results showed that NOXA-SBS1 sequence located 2.4kb relative to the translational start site were specifically immunoprecipitated with anti-SATB1 (B, indicating that SATB1 binds to this sequence in vivo. The SATB1 binding to SBS2 was very weak (C. No SATB1 binding to SBS3 was detected (D. As confirmed earlier, SATB1 bound to the mbr (E). The input represented 1% of total DNA used in immunoprecipitation. Non-specific IgG was used as a negative control. The PCR products were visualized by ethidium bromide staining of a 1.5% agarose gel.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4589335&req=5

pone.0139170.g001: ChIP analysis of SATB1 binding on the NOXA promoter region and the mbr element in vivo.Diagram shows the relative positions of three SATB1 binding sites(N-SBS1、N-SBS2、N-SBS3) in NOXA promoter region and mbr(A). The results showed that NOXA-SBS1 sequence located 2.4kb relative to the translational start site were specifically immunoprecipitated with anti-SATB1 (B, indicating that SATB1 binds to this sequence in vivo. The SATB1 binding to SBS2 was very weak (C. No SATB1 binding to SBS3 was detected (D. As confirmed earlier, SATB1 bound to the mbr (E). The input represented 1% of total DNA used in immunoprecipitation. Non-specific IgG was used as a negative control. The PCR products were visualized by ethidium bromide staining of a 1.5% agarose gel.
Mentions: We have demonstrated that the SATB1-mediated long-range chromatin interaction between the mbr enhancer and BCL2 promoter is critical to regulating BCL2 expression in the cellular apoptotic response [25]. To investigate whether SATB1-mediated long-range chromatin interaction is involved in co-regulation of anti-apoptotic and pro-apoptotic genes, we used Genomatix Software (Genomatix Software, http://www.genomatix.de/index.html) to analyze the promoter regions of Bcl2 family members for the binding sites of SATB1). The NOXA gene located 3.4Mb downstream of the BCL2 gene was of particular interest. NOXA has 3 sequences that are possible SATB1 binding sites. These sequences are NOXA-SBS1 to NOXA-SBS3, and are located within the 3.5kb region upstream of the NOXA transcription start site. The illustrations of three SATB1 binding sites in NOXA promoter have been shown in Fig 1. We used ChIP assays with the anti-SATB1 antibody to test SATB1 binding on the NOXA promoter. NOXA-SBS1 sequence located 2.4kb relative to the translational start site were found to be specifically immunoprecipitated with anti-SATB1 (Fig 2), indicating that SATB1 binds to this sequence in vivo. The precipitated products were weak for SBS2. No specific precipitate was detected for SBS3 (Fig 2). As expected and confirmed in our previous work, SATB1 also bound to the mbr enhancer (Fig 2). Given that SATB1 is a critical factor in regulating long-range interactions between the mbr and BCL2 promoter [25], the binding of SATB1 to the NOXA promoter strongly suggested that SATB1 not only mediated an interaction between the mbr and the BCL2 promoter, but also was involved in the mbr-NOXA chromatin interaction.

Bottom Line: Decreased SATB1 levels switched the mbr-BCL2 loop to mbr-NOXA loop, and thus changed expression of these two genes.The SATB1-mediated dynamic switch of the chromatin loop structures was essential for the cooperative expression of the BCL2 and NOXA genes in apoptosis.Notably, the role of SATB1 was specific, since inhibition of SATB1 degradation by caspase-6 inhibitor or caspase-6-resistant SATB1 mutant reversed expression of BCL-2 and NOXA in response to apoptotic stimulation.

View Article: PubMed Central - PubMed

Affiliation: Key Laboratory of Human Functional Genomics of Jiangsu Province, Nanjing medical University, Nanjing, PR China; Department of Cell Biology, Nanjing Medical University, Nanjing, PR China.

ABSTRACT
Aberrant expression of special AT-rich binding protein 1 (SATB1), a global genomic organizer, has been associated with various cancers, which raises the question of how higher-order chromatin structure contributes to carcinogenesis. Disruption of apoptosis is one of the hallmarks of cancer. We previously demonstrated that SATB1 mediated specific long-range chromosomal interactions between the mbr enhancer located within 3'-UTR of the BCL2 gene and the promoter to regulate BCL2 expression during early apoptosis. In the present study, we used chromosome conformation capture (3C) assays and molecular analyses to further investigate the function of the SATB1-mediated higher-order chromatin structure in co-regulation of the anti-apoptotic BCL2 gene and the pro-apoptotic NOXA gene located 3.4Mb downstream on Chromosome 18. We demonstrated that the mbr enhancer spatially juxtaposed the promoters of BCL2 and NOXA genes through SATB1-mediated chromatin-loop in Jurkat cells. Decreased SATB1 levels switched the mbr-BCL2 loop to mbr-NOXA loop, and thus changed expression of these two genes. The SATB1-mediated dynamic switch of the chromatin loop structures was essential for the cooperative expression of the BCL2 and NOXA genes in apoptosis. Notably, the role of SATB1 was specific, since inhibition of SATB1 degradation by caspase-6 inhibitor or caspase-6-resistant SATB1 mutant reversed expression of BCL-2 and NOXA in response to apoptotic stimulation. This study reveals the critical role of SATB1-organized higher-order chromatin structure in regulating the dynamic equilibrium of apoptosis-controlling genes with antagonistic functions and suggests that aberrant SATB1 expression might contribute to cancer development by disrupting the co-regulated genes in apoptosis pathways.

No MeSH data available.


Related in: MedlinePlus