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Decreased Dicer expression elicits DNA damage and up-regulation of MICA and MICB.

Tang KF, Ren H, Cao J, Zeng GL, Xie J, Chen M, Wang L, He CX - J. Cell Biol. (2008)

Bottom Line: We reveal that knockdown of Dicer elicits DNA damage.Therefore we conclude that up-regulation of MICA and MICB is the result of DNA damage response activation caused by Dicer knockdown.Our results suggest that RNAi is indirectly linked to the human innate immune system via the DNA damage pathway.

View Article: PubMed Central - PubMed

Affiliation: Key Laboratory of Molecular Biology for Infectious Diseases of the State Ministry of Education, The Second Affiliated Hospital of Chongqing Medical University, Chongqing 400010, People's Republic of China. tang_kaifu@yahoo.com.cn

ABSTRACT
RNA interference (RNAi) acts constitutively to silence the innate immune response, and innate immunity genes are misregulated in Dicer-deficient Caenorhabditis elegans. Here, we show that inhibition of Dicer expression by RNAi in human cells up-regulates major histocompatibility complex class I-related molecules A and B (MICA and MICB). MICA and MICB are innate immune system ligands for the NKG2D receptor expressed by natural killer cells and activated CD8(+)T cells. We reveal that knockdown of Dicer elicits DNA damage. Up-regulation of MICA and MICB by Dicer knockdown is prevented by pharmacologic or genetic inhibition of DNA damage pathway components, including ataxia telangiectasia mutated (ATM) kinase, ATM- and Rad3-related kinase, or checkpoint kinase 1. Therefore we conclude that up-regulation of MICA and MICB is the result of DNA damage response activation caused by Dicer knockdown. Our results suggest that RNAi is indirectly linked to the human innate immune system via the DNA damage pathway.

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Inhibition of DNA damage response prevents MICA and MICB up-regulation in Dicer knockdown cells. Dicer knockdown-induced MICA and MICB up-regulation was prevented by incubation with caffeine (C) or staurosporine (S) and by cotransfection with ATR siRNA (R), ATM siRNA (M), or Chk1 siRNA (K). MICA and MICB mRNA levels were determined by real time RT-PCR, and were normalized to these in mock-transfected cells, which are defined as 1. Data represent means ± SD from three independent experiments.
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fig3: Inhibition of DNA damage response prevents MICA and MICB up-regulation in Dicer knockdown cells. Dicer knockdown-induced MICA and MICB up-regulation was prevented by incubation with caffeine (C) or staurosporine (S) and by cotransfection with ATR siRNA (R), ATM siRNA (M), or Chk1 siRNA (K). MICA and MICB mRNA levels were determined by real time RT-PCR, and were normalized to these in mock-transfected cells, which are defined as 1. Data represent means ± SD from three independent experiments.

Mentions: DNA damage–induced up-regulation of NKG2D ligands can be prevented by pharmacologic or genetic inhibition of ATR, ATM, or Chk1 (Gasser et al., 2005). The roles of ATR, ATM, and Chk1 in Dicer knockdown–induced MICA and MICB up-regulation were investigated. Up-regulation of MICA and MICB in response to Dicer knockdown was blocked by caffeine, an inhibitor of ATR and ATM, and by staurosporine, an inhibitor of Chk1 (Figs. 3 and S3 A, available at http://www.jcb.org/cgi/content/full/jcb.200801169/DC1). As a more specific test, expression of ATM, ATR, and Chk1 was repressed using siRNAs (Fig. S3, E and F). Up-regulation of MICA and MICB induced by Dicer knockdown was inhibited by cotransfection of Dicer siRNAs with ATR siRNA (Figs. 3 and S3 B). In contrast, cotransfection with control siRNA did not significantly affect Dicer knockdown–induced up-regulation of MICA and MICB (Fig. 3). Similarly, inhibition of ATM and Chk1 expression by siRNAs also blocked MICA and MICB up-regulation in response to Dicer knockdown (Figs. 3 and S3, C and D). In addition, we demonstrated that DNA damage progressed in parallel with the kinetics of up-regulation of MICA and MICB in Dicer knockdown cells (Fig. 4). These data suggest that up-regulation of MICA and MICB is a consequence of DNA damage response activation. Compared with ATR suppression, ATM suppression had larger effects on MICA and MICB expression (Figs. 3 and S3, B and C), which suggests that MICA and MICB up-regulation is predominantly in response to DSBs.


Decreased Dicer expression elicits DNA damage and up-regulation of MICA and MICB.

Tang KF, Ren H, Cao J, Zeng GL, Xie J, Chen M, Wang L, He CX - J. Cell Biol. (2008)

Inhibition of DNA damage response prevents MICA and MICB up-regulation in Dicer knockdown cells. Dicer knockdown-induced MICA and MICB up-regulation was prevented by incubation with caffeine (C) or staurosporine (S) and by cotransfection with ATR siRNA (R), ATM siRNA (M), or Chk1 siRNA (K). MICA and MICB mRNA levels were determined by real time RT-PCR, and were normalized to these in mock-transfected cells, which are defined as 1. Data represent means ± SD from three independent experiments.
© Copyright Policy
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC2483517&req=5

fig3: Inhibition of DNA damage response prevents MICA and MICB up-regulation in Dicer knockdown cells. Dicer knockdown-induced MICA and MICB up-regulation was prevented by incubation with caffeine (C) or staurosporine (S) and by cotransfection with ATR siRNA (R), ATM siRNA (M), or Chk1 siRNA (K). MICA and MICB mRNA levels were determined by real time RT-PCR, and were normalized to these in mock-transfected cells, which are defined as 1. Data represent means ± SD from three independent experiments.
Mentions: DNA damage–induced up-regulation of NKG2D ligands can be prevented by pharmacologic or genetic inhibition of ATR, ATM, or Chk1 (Gasser et al., 2005). The roles of ATR, ATM, and Chk1 in Dicer knockdown–induced MICA and MICB up-regulation were investigated. Up-regulation of MICA and MICB in response to Dicer knockdown was blocked by caffeine, an inhibitor of ATR and ATM, and by staurosporine, an inhibitor of Chk1 (Figs. 3 and S3 A, available at http://www.jcb.org/cgi/content/full/jcb.200801169/DC1). As a more specific test, expression of ATM, ATR, and Chk1 was repressed using siRNAs (Fig. S3, E and F). Up-regulation of MICA and MICB induced by Dicer knockdown was inhibited by cotransfection of Dicer siRNAs with ATR siRNA (Figs. 3 and S3 B). In contrast, cotransfection with control siRNA did not significantly affect Dicer knockdown–induced up-regulation of MICA and MICB (Fig. 3). Similarly, inhibition of ATM and Chk1 expression by siRNAs also blocked MICA and MICB up-regulation in response to Dicer knockdown (Figs. 3 and S3, C and D). In addition, we demonstrated that DNA damage progressed in parallel with the kinetics of up-regulation of MICA and MICB in Dicer knockdown cells (Fig. 4). These data suggest that up-regulation of MICA and MICB is a consequence of DNA damage response activation. Compared with ATR suppression, ATM suppression had larger effects on MICA and MICB expression (Figs. 3 and S3, B and C), which suggests that MICA and MICB up-regulation is predominantly in response to DSBs.

Bottom Line: We reveal that knockdown of Dicer elicits DNA damage.Therefore we conclude that up-regulation of MICA and MICB is the result of DNA damage response activation caused by Dicer knockdown.Our results suggest that RNAi is indirectly linked to the human innate immune system via the DNA damage pathway.

View Article: PubMed Central - PubMed

Affiliation: Key Laboratory of Molecular Biology for Infectious Diseases of the State Ministry of Education, The Second Affiliated Hospital of Chongqing Medical University, Chongqing 400010, People's Republic of China. tang_kaifu@yahoo.com.cn

ABSTRACT
RNA interference (RNAi) acts constitutively to silence the innate immune response, and innate immunity genes are misregulated in Dicer-deficient Caenorhabditis elegans. Here, we show that inhibition of Dicer expression by RNAi in human cells up-regulates major histocompatibility complex class I-related molecules A and B (MICA and MICB). MICA and MICB are innate immune system ligands for the NKG2D receptor expressed by natural killer cells and activated CD8(+)T cells. We reveal that knockdown of Dicer elicits DNA damage. Up-regulation of MICA and MICB by Dicer knockdown is prevented by pharmacologic or genetic inhibition of DNA damage pathway components, including ataxia telangiectasia mutated (ATM) kinase, ATM- and Rad3-related kinase, or checkpoint kinase 1. Therefore we conclude that up-regulation of MICA and MICB is the result of DNA damage response activation caused by Dicer knockdown. Our results suggest that RNAi is indirectly linked to the human innate immune system via the DNA damage pathway.

Show MeSH
Related in: MedlinePlus