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MicroRNA-130a associates with ribosomal protein L11 to suppress c-Myc expression in response to UV irradiation.

Li Y, Challagundla KB, Sun XX, Zhang Q, Dai MS - Oncotarget (2015)

Bottom Line: Yet, mechanism underlying UV-induced c-Myc reduction is not completely understood.Overexpression of miR-130a promotes the Ago2 binding to c-myc mRNA, significantly reduces the levels of both c-Myc protein and mRNA and inhibits cell proliferation.Inhibiting miR-130a significantly suppresses UV-mediated c-Myc reduction.

View Article: PubMed Central - PubMed

Affiliation: Departments of Molecular & Medical Genetics, School of Medicine and the OHSU Knight Cancer Institute, Oregon Health & Science University, Portland, OR 97239, USA.

ABSTRACT
The oncoprotein c-Myc is essential for cell growth and proliferation while its deregulated overexpression is associated with most human cancers. Thus tightly regulated levels and activity of c-Myc are critical for maintaining normal cell homeostasis. c-Myc is down-regulated in response to several types of stress, including UV-induced DNA damage. Yet, mechanism underlying UV-induced c-Myc reduction is not completely understood. Here we report that L11 promotes miR-130a targeting of c-myc mRNA to repress c-Myc expression in response to UV irradiation. miR-130a targets the 3'-untranslated region (UTR) of c-myc mRNA. Overexpression of miR-130a promotes the Ago2 binding to c-myc mRNA, significantly reduces the levels of both c-Myc protein and mRNA and inhibits cell proliferation. UV treatment markedly promotes the binding of L11 to miR-130a, c-myc mRNA as well as Ago2 in cells. Inhibiting miR-130a significantly suppresses UV-mediated c-Myc reduction. We further show that L11 is relocalized from the nucleolus to the cytoplasm where it associates with c-myc mRNA upon UV treatment. Together, these results reveal a novel mechanism underlying c-Myc down-regulation in response to UV-mediated DNA damage, wherein L11 promotes miR-130a-loaded miRISC to target c-myc mRNA.

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UV irradiation promotes L11 interaction with miR-130a and c-myc mRNA in the cytoplasm(A) UV treatment releases L11 from the nucleolus into the nucleoplasm and the cytoplasm. U2OS cells treated with or without 40 J/m2 UV for 6 hours were subjected to isolation of cytoplasm (Cyto), nucleoplasm (Np), and the nucleolus (No) fractions, followed by IB detection of indicated proteins. Tubulin, SP1, and nucleolin (C23) were used as cytoplasm, nucleoplasm and nucleolar markers, respectively. (B–C) UV treatment increases L11 binding to miR-130a and c-myc mRNA in the cytoplasm. The cytoplasmic and the nuclear (Nuc) fractions isolated from U2OS cells treated with or without 40 J/m2 UV for 6 hours were immunoprecipitated with anti-L11 antibodies or control rabbit IgG, followed by RT-qPCR detection of c-myc mRNA (B) and miR-130a (C).
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Figure 7: UV irradiation promotes L11 interaction with miR-130a and c-myc mRNA in the cytoplasm(A) UV treatment releases L11 from the nucleolus into the nucleoplasm and the cytoplasm. U2OS cells treated with or without 40 J/m2 UV for 6 hours were subjected to isolation of cytoplasm (Cyto), nucleoplasm (Np), and the nucleolus (No) fractions, followed by IB detection of indicated proteins. Tubulin, SP1, and nucleolin (C23) were used as cytoplasm, nucleoplasm and nucleolar markers, respectively. (B–C) UV treatment increases L11 binding to miR-130a and c-myc mRNA in the cytoplasm. The cytoplasmic and the nuclear (Nuc) fractions isolated from U2OS cells treated with or without 40 J/m2 UV for 6 hours were immunoprecipitated with anti-L11 antibodies or control rabbit IgG, followed by RT-qPCR detection of c-myc mRNA (B) and miR-130a (C).

Mentions: To test how L11 targets c-myc mRNA in response to UV treatment, we examined whether UV treatment could increase L11 levels in the cytoplasm. U2OS cells treated with or without UV were fractionated into the cytoplasm, nucleoplasm and nucleolar fractions, followed by IB. As shown in Fig. 7A, UV treatment significantly increased the levels of L11 in both the nucleoplasm and the cytoplasm, whereas the nucleolar L11 was reduced by UV treatment. This is consistent with previous report showing that UV damage causes nucleolar disruption [42]. RNA-IP assays using the cytoplasmic and nucleoplasmic lysates with control or anti-L11 antibodies showed that UV treatment significantly increased the L11 binding to both c-myc mRNA (Fig. 7B) and miR-130a (Fig. 7C) in the cytoplasm, but not in the nucleoplasm. These results indicate that in response to UV damage, L11 is released form the nucleolus to the cytoplasm where it recruits miR-130a/miRISC to the c-myc 3′-UTR, leading to c-myc mRNA decay.


MicroRNA-130a associates with ribosomal protein L11 to suppress c-Myc expression in response to UV irradiation.

Li Y, Challagundla KB, Sun XX, Zhang Q, Dai MS - Oncotarget (2015)

UV irradiation promotes L11 interaction with miR-130a and c-myc mRNA in the cytoplasm(A) UV treatment releases L11 from the nucleolus into the nucleoplasm and the cytoplasm. U2OS cells treated with or without 40 J/m2 UV for 6 hours were subjected to isolation of cytoplasm (Cyto), nucleoplasm (Np), and the nucleolus (No) fractions, followed by IB detection of indicated proteins. Tubulin, SP1, and nucleolin (C23) were used as cytoplasm, nucleoplasm and nucleolar markers, respectively. (B–C) UV treatment increases L11 binding to miR-130a and c-myc mRNA in the cytoplasm. The cytoplasmic and the nuclear (Nuc) fractions isolated from U2OS cells treated with or without 40 J/m2 UV for 6 hours were immunoprecipitated with anti-L11 antibodies or control rabbit IgG, followed by RT-qPCR detection of c-myc mRNA (B) and miR-130a (C).
© Copyright Policy - open-access
Related In: Results  -  Collection

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Figure 7: UV irradiation promotes L11 interaction with miR-130a and c-myc mRNA in the cytoplasm(A) UV treatment releases L11 from the nucleolus into the nucleoplasm and the cytoplasm. U2OS cells treated with or without 40 J/m2 UV for 6 hours were subjected to isolation of cytoplasm (Cyto), nucleoplasm (Np), and the nucleolus (No) fractions, followed by IB detection of indicated proteins. Tubulin, SP1, and nucleolin (C23) were used as cytoplasm, nucleoplasm and nucleolar markers, respectively. (B–C) UV treatment increases L11 binding to miR-130a and c-myc mRNA in the cytoplasm. The cytoplasmic and the nuclear (Nuc) fractions isolated from U2OS cells treated with or without 40 J/m2 UV for 6 hours were immunoprecipitated with anti-L11 antibodies or control rabbit IgG, followed by RT-qPCR detection of c-myc mRNA (B) and miR-130a (C).
Mentions: To test how L11 targets c-myc mRNA in response to UV treatment, we examined whether UV treatment could increase L11 levels in the cytoplasm. U2OS cells treated with or without UV were fractionated into the cytoplasm, nucleoplasm and nucleolar fractions, followed by IB. As shown in Fig. 7A, UV treatment significantly increased the levels of L11 in both the nucleoplasm and the cytoplasm, whereas the nucleolar L11 was reduced by UV treatment. This is consistent with previous report showing that UV damage causes nucleolar disruption [42]. RNA-IP assays using the cytoplasmic and nucleoplasmic lysates with control or anti-L11 antibodies showed that UV treatment significantly increased the L11 binding to both c-myc mRNA (Fig. 7B) and miR-130a (Fig. 7C) in the cytoplasm, but not in the nucleoplasm. These results indicate that in response to UV damage, L11 is released form the nucleolus to the cytoplasm where it recruits miR-130a/miRISC to the c-myc 3′-UTR, leading to c-myc mRNA decay.

Bottom Line: Yet, mechanism underlying UV-induced c-Myc reduction is not completely understood.Overexpression of miR-130a promotes the Ago2 binding to c-myc mRNA, significantly reduces the levels of both c-Myc protein and mRNA and inhibits cell proliferation.Inhibiting miR-130a significantly suppresses UV-mediated c-Myc reduction.

View Article: PubMed Central - PubMed

Affiliation: Departments of Molecular & Medical Genetics, School of Medicine and the OHSU Knight Cancer Institute, Oregon Health & Science University, Portland, OR 97239, USA.

ABSTRACT
The oncoprotein c-Myc is essential for cell growth and proliferation while its deregulated overexpression is associated with most human cancers. Thus tightly regulated levels and activity of c-Myc are critical for maintaining normal cell homeostasis. c-Myc is down-regulated in response to several types of stress, including UV-induced DNA damage. Yet, mechanism underlying UV-induced c-Myc reduction is not completely understood. Here we report that L11 promotes miR-130a targeting of c-myc mRNA to repress c-Myc expression in response to UV irradiation. miR-130a targets the 3'-untranslated region (UTR) of c-myc mRNA. Overexpression of miR-130a promotes the Ago2 binding to c-myc mRNA, significantly reduces the levels of both c-Myc protein and mRNA and inhibits cell proliferation. UV treatment markedly promotes the binding of L11 to miR-130a, c-myc mRNA as well as Ago2 in cells. Inhibiting miR-130a significantly suppresses UV-mediated c-Myc reduction. We further show that L11 is relocalized from the nucleolus to the cytoplasm where it associates with c-myc mRNA upon UV treatment. Together, these results reveal a novel mechanism underlying c-Myc down-regulation in response to UV-mediated DNA damage, wherein L11 promotes miR-130a-loaded miRISC to target c-myc mRNA.

Show MeSH
Related in: MedlinePlus