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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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L11 is involved in UV-induced c-Myc downregulation(A–D) UV irradiation decreases c-Myc levels. U2OS cells were exposed to different dosages of UV (A) (B) or 40 J/m2 UV for different times (C) (D). The cells were assayed for the expression of c-Myc protein by IB (A) (C) and c-myc mRNA by RT-qPCR (B) (D). (E) MG132 treatment partially rescued the c-Myc reduction by UV treatment. U2OS cells treated with 40 J/m2 UV were cultured in the presence or absence of 40 μM MG132 for 6 hours followed by IB. (F) UV treatment does not reduce the levels of HuR, eIF4G and ribosomal protein L5 (RPL5). U2OS cells were treated with different dosages of UV for 6 hours and assayed by IB. (G–H) Knockdown of L11 abolished the c-Myc reduction by UV treatment. U2OS cells transfected with scrambled or L11 siRNA were treated with 40 J/m2 UV for 6 hours. The cells were subjected to IB detection of c-Myc protein (G) and RT-qPCR detection of c-myc mRNA (H). *p < 0.01, compared to scrambled RNA transfected cells.
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Figure 5: L11 is involved in UV-induced c-Myc downregulation(A–D) UV irradiation decreases c-Myc levels. U2OS cells were exposed to different dosages of UV (A) (B) or 40 J/m2 UV for different times (C) (D). The cells were assayed for the expression of c-Myc protein by IB (A) (C) and c-myc mRNA by RT-qPCR (B) (D). (E) MG132 treatment partially rescued the c-Myc reduction by UV treatment. U2OS cells treated with 40 J/m2 UV were cultured in the presence or absence of 40 μM MG132 for 6 hours followed by IB. (F) UV treatment does not reduce the levels of HuR, eIF4G and ribosomal protein L5 (RPL5). U2OS cells were treated with different dosages of UV for 6 hours and assayed by IB. (G–H) Knockdown of L11 abolished the c-Myc reduction by UV treatment. U2OS cells transfected with scrambled or L11 siRNA were treated with 40 J/m2 UV for 6 hours. The cells were subjected to IB detection of c-Myc protein (G) and RT-qPCR detection of c-myc mRNA (H). *p < 0.01, compared to scrambled RNA transfected cells.

Mentions: To determine the physiological relevance of the L11-miR-130a regulation of c-Myc, we asked whether L11 recruits miR-130a to target c-Myc in response to stress. It has recently been shown that c-Myc protein is reduced by treatment of cells with UV irradiation [29] and DNA damaging agents [30], although the underlying mechanism is not completely understood. In agreement with these studies, we observed that c-Myc protein is reduced by UV treatment in U2OS cells (Fig. 5A and 5C). Interestingly, c-myc mRNA was also significantly reduced by UV treatment in a dose- and time-dependent manner (Figs. 5B and 5D), indicating that c-Myc is regulated at mRNA levels as well in response to UV-induced DNA damage. Also consistent with the previous study (30), UV-mediated c-Myc reduction was partially rescued by the treatment with the proteasome inhibitor MG132 (Fig. 5E). Thus, UV treatment leads to both degradation of existing c-Myc protein and the reduction of c-myc mRNA. Of note, the c-Myc reduction is not a general consequence of cellular response to UV irradiation, as the levels of several other tested proteins, including HuR, eIF4G, and ribosomal protein L5 (RPL5), were not decreased following UV treatment (Fig. 5F).


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)

L11 is involved in UV-induced c-Myc downregulation(A–D) UV irradiation decreases c-Myc levels. U2OS cells were exposed to different dosages of UV (A) (B) or 40 J/m2 UV for different times (C) (D). The cells were assayed for the expression of c-Myc protein by IB (A) (C) and c-myc mRNA by RT-qPCR (B) (D). (E) MG132 treatment partially rescued the c-Myc reduction by UV treatment. U2OS cells treated with 40 J/m2 UV were cultured in the presence or absence of 40 μM MG132 for 6 hours followed by IB. (F) UV treatment does not reduce the levels of HuR, eIF4G and ribosomal protein L5 (RPL5). U2OS cells were treated with different dosages of UV for 6 hours and assayed by IB. (G–H) Knockdown of L11 abolished the c-Myc reduction by UV treatment. U2OS cells transfected with scrambled or L11 siRNA were treated with 40 J/m2 UV for 6 hours. The cells were subjected to IB detection of c-Myc protein (G) and RT-qPCR detection of c-myc mRNA (H). *p < 0.01, compared to scrambled RNA transfected cells.
© Copyright Policy - open-access
Related In: Results  -  Collection

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Figure 5: L11 is involved in UV-induced c-Myc downregulation(A–D) UV irradiation decreases c-Myc levels. U2OS cells were exposed to different dosages of UV (A) (B) or 40 J/m2 UV for different times (C) (D). The cells were assayed for the expression of c-Myc protein by IB (A) (C) and c-myc mRNA by RT-qPCR (B) (D). (E) MG132 treatment partially rescued the c-Myc reduction by UV treatment. U2OS cells treated with 40 J/m2 UV were cultured in the presence or absence of 40 μM MG132 for 6 hours followed by IB. (F) UV treatment does not reduce the levels of HuR, eIF4G and ribosomal protein L5 (RPL5). U2OS cells were treated with different dosages of UV for 6 hours and assayed by IB. (G–H) Knockdown of L11 abolished the c-Myc reduction by UV treatment. U2OS cells transfected with scrambled or L11 siRNA were treated with 40 J/m2 UV for 6 hours. The cells were subjected to IB detection of c-Myc protein (G) and RT-qPCR detection of c-myc mRNA (H). *p < 0.01, compared to scrambled RNA transfected cells.
Mentions: To determine the physiological relevance of the L11-miR-130a regulation of c-Myc, we asked whether L11 recruits miR-130a to target c-Myc in response to stress. It has recently been shown that c-Myc protein is reduced by treatment of cells with UV irradiation [29] and DNA damaging agents [30], although the underlying mechanism is not completely understood. In agreement with these studies, we observed that c-Myc protein is reduced by UV treatment in U2OS cells (Fig. 5A and 5C). Interestingly, c-myc mRNA was also significantly reduced by UV treatment in a dose- and time-dependent manner (Figs. 5B and 5D), indicating that c-Myc is regulated at mRNA levels as well in response to UV-induced DNA damage. Also consistent with the previous study (30), UV-mediated c-Myc reduction was partially rescued by the treatment with the proteasome inhibitor MG132 (Fig. 5E). Thus, UV treatment leads to both degradation of existing c-Myc protein and the reduction of c-myc mRNA. Of note, the c-Myc reduction is not a general consequence of cellular response to UV irradiation, as the levels of several other tested proteins, including HuR, eIF4G, and ribosomal protein L5 (RPL5), were not decreased following UV treatment (Fig. 5F).

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