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DNA double-strand breaks activate a multi-functional genetic program in developing lymphocytes.

Bredemeyer AL, Helmink BA, Innes CL, Calderon B, McGinnis LM, Mahowald GK, Gapud EJ, Walker LM, Collins JB, Weaver BK, Mandik-Nayak L, Schreiber RD, Allen PM, May MJ, Paules RS, Bassing CH, Sleckman BP - Nature (2008)

Bottom Line: DNA double-strand breaks are generated by genotoxic agents and by cellular endonucleases as intermediates of several important physiological processes.The cellular response to genotoxic DNA breaks includes the activation of transcriptional programs known primarily to regulate cell-cycle checkpoints and cell survival.Thus, physiological DNA double-strand breaks provide cues that can regulate cell-type-specific processes not directly involved in maintaining the integrity of the genome, and genotoxic DNA breaks could disrupt normal cellular functions by corrupting these processes.

View Article: PubMed Central - PubMed

Affiliation: Department of Pathology and Immunology, Washington University School of Medicine, St. Louis, Missouri 63110, USA.

ABSTRACT
DNA double-strand breaks are generated by genotoxic agents and by cellular endonucleases as intermediates of several important physiological processes. The cellular response to genotoxic DNA breaks includes the activation of transcriptional programs known primarily to regulate cell-cycle checkpoints and cell survival. DNA double-strand breaks are generated in all developing lymphocytes during the assembly of antigen receptor genes, a process that is essential for normal lymphocyte development. Here we show that in murine lymphocytes these physiological DNA breaks activate a broad transcriptional program. This program transcends the canonical DNA double-strand break response and includes many genes that regulate diverse cellular processes important for lymphocyte development. Moreover, the expression of several of these genes is regulated similarly in response to genotoxic DNA damage. Thus, physiological DNA double-strand breaks provide cues that can regulate cell-type-specific processes not directly involved in maintaining the integrity of the genome, and genotoxic DNA breaks could disrupt normal cellular functions by corrupting these processes.

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Genotoxic DSBs promote changes in expression of lymphocyte-specific genes(a) RT-PCR analysis of gene expression in Rag2−/− abl pre-B cells 2 hours after receiving 0 Gy (white bars) or 4 Gy of IR in the presence (grey bars) or absence (red bars) of the Atm inhibitor KU-55933 (iAtm). All cells were treated with STI571 for 24 hours prior to IR. Results are the mean and standard deviation from two experiments. P values were calculated using a one-tailed t-test. (b) Flow cytometric analysis of CD69 expression on STI571-treated Rag2−/− abl pre-B cells treated with IR as described in (a) or 2 hours after being treated with 5µM etoposide (Etp). Results are representative of three experiments. (c) Flow cytometric analysis of B220 and CD69 expression on bone marrow cells from Rag1−/−:IgHtg mice 2 hours after irradiation (0.5 Gy) or etoposide treatment (5µM) in the presence or absence of the Atm inhibitor KU-55933. Results are representative of two experiments.
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Figure 4: Genotoxic DSBs promote changes in expression of lymphocyte-specific genes(a) RT-PCR analysis of gene expression in Rag2−/− abl pre-B cells 2 hours after receiving 0 Gy (white bars) or 4 Gy of IR in the presence (grey bars) or absence (red bars) of the Atm inhibitor KU-55933 (iAtm). All cells were treated with STI571 for 24 hours prior to IR. Results are the mean and standard deviation from two experiments. P values were calculated using a one-tailed t-test. (b) Flow cytometric analysis of CD69 expression on STI571-treated Rag2−/− abl pre-B cells treated with IR as described in (a) or 2 hours after being treated with 5µM etoposide (Etp). Results are representative of three experiments. (c) Flow cytometric analysis of B220 and CD69 expression on bone marrow cells from Rag1−/−:IgHtg mice 2 hours after irradiation (0.5 Gy) or etoposide treatment (5µM) in the presence or absence of the Atm inhibitor KU-55933. Results are representative of two experiments.

Mentions: As physiologic Rag DSBs activate a broad genetic program, we reasoned that genotoxic DSBs may also activate components of this program in developing lymphocytes. Indeed, treatment of STI571-treated Rag2−/− abl pre-B cells with either ionizing radiation (IR) or etoposide, which both generate DNA DSBs, led to the Atm-dependent induction of many genes that are also induced by Rag DSBs (Fig. 4a and b, Supplementary Fig. 18). Furthermore, treatment of bone marrow from Rag1−/−:IgHtg mice with either ionizing radiation or etoposide leads to a B-cell-specific (B220+), Atm-dependent induction of CD69 expression (Fig. 4c). These findings demonstrate that, in developing B cells, genotoxic DSBs can induce lymphocyte-specific gene expression changes that are normally induced by Rag DSBs.


DNA double-strand breaks activate a multi-functional genetic program in developing lymphocytes.

Bredemeyer AL, Helmink BA, Innes CL, Calderon B, McGinnis LM, Mahowald GK, Gapud EJ, Walker LM, Collins JB, Weaver BK, Mandik-Nayak L, Schreiber RD, Allen PM, May MJ, Paules RS, Bassing CH, Sleckman BP - Nature (2008)

Genotoxic DSBs promote changes in expression of lymphocyte-specific genes(a) RT-PCR analysis of gene expression in Rag2−/− abl pre-B cells 2 hours after receiving 0 Gy (white bars) or 4 Gy of IR in the presence (grey bars) or absence (red bars) of the Atm inhibitor KU-55933 (iAtm). All cells were treated with STI571 for 24 hours prior to IR. Results are the mean and standard deviation from two experiments. P values were calculated using a one-tailed t-test. (b) Flow cytometric analysis of CD69 expression on STI571-treated Rag2−/− abl pre-B cells treated with IR as described in (a) or 2 hours after being treated with 5µM etoposide (Etp). Results are representative of three experiments. (c) Flow cytometric analysis of B220 and CD69 expression on bone marrow cells from Rag1−/−:IgHtg mice 2 hours after irradiation (0.5 Gy) or etoposide treatment (5µM) in the presence or absence of the Atm inhibitor KU-55933. Results are representative of two experiments.
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Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2605662&req=5

Figure 4: Genotoxic DSBs promote changes in expression of lymphocyte-specific genes(a) RT-PCR analysis of gene expression in Rag2−/− abl pre-B cells 2 hours after receiving 0 Gy (white bars) or 4 Gy of IR in the presence (grey bars) or absence (red bars) of the Atm inhibitor KU-55933 (iAtm). All cells were treated with STI571 for 24 hours prior to IR. Results are the mean and standard deviation from two experiments. P values were calculated using a one-tailed t-test. (b) Flow cytometric analysis of CD69 expression on STI571-treated Rag2−/− abl pre-B cells treated with IR as described in (a) or 2 hours after being treated with 5µM etoposide (Etp). Results are representative of three experiments. (c) Flow cytometric analysis of B220 and CD69 expression on bone marrow cells from Rag1−/−:IgHtg mice 2 hours after irradiation (0.5 Gy) or etoposide treatment (5µM) in the presence or absence of the Atm inhibitor KU-55933. Results are representative of two experiments.
Mentions: As physiologic Rag DSBs activate a broad genetic program, we reasoned that genotoxic DSBs may also activate components of this program in developing lymphocytes. Indeed, treatment of STI571-treated Rag2−/− abl pre-B cells with either ionizing radiation (IR) or etoposide, which both generate DNA DSBs, led to the Atm-dependent induction of many genes that are also induced by Rag DSBs (Fig. 4a and b, Supplementary Fig. 18). Furthermore, treatment of bone marrow from Rag1−/−:IgHtg mice with either ionizing radiation or etoposide leads to a B-cell-specific (B220+), Atm-dependent induction of CD69 expression (Fig. 4c). These findings demonstrate that, in developing B cells, genotoxic DSBs can induce lymphocyte-specific gene expression changes that are normally induced by Rag DSBs.

Bottom Line: DNA double-strand breaks are generated by genotoxic agents and by cellular endonucleases as intermediates of several important physiological processes.The cellular response to genotoxic DNA breaks includes the activation of transcriptional programs known primarily to regulate cell-cycle checkpoints and cell survival.Thus, physiological DNA double-strand breaks provide cues that can regulate cell-type-specific processes not directly involved in maintaining the integrity of the genome, and genotoxic DNA breaks could disrupt normal cellular functions by corrupting these processes.

View Article: PubMed Central - PubMed

Affiliation: Department of Pathology and Immunology, Washington University School of Medicine, St. Louis, Missouri 63110, USA.

ABSTRACT
DNA double-strand breaks are generated by genotoxic agents and by cellular endonucleases as intermediates of several important physiological processes. The cellular response to genotoxic DNA breaks includes the activation of transcriptional programs known primarily to regulate cell-cycle checkpoints and cell survival. DNA double-strand breaks are generated in all developing lymphocytes during the assembly of antigen receptor genes, a process that is essential for normal lymphocyte development. Here we show that in murine lymphocytes these physiological DNA breaks activate a broad transcriptional program. This program transcends the canonical DNA double-strand break response and includes many genes that regulate diverse cellular processes important for lymphocyte development. Moreover, the expression of several of these genes is regulated similarly in response to genotoxic DNA damage. Thus, physiological DNA double-strand breaks provide cues that can regulate cell-type-specific processes not directly involved in maintaining the integrity of the genome, and genotoxic DNA breaks could disrupt normal cellular functions by corrupting these processes.

Show MeSH
Related in: MedlinePlus