Limits...
Hypoxia-induced MIR155 is a potent autophagy inducer by targeting multiple players in the MTOR pathway.

Wan G, Xie W, Liu Z, Xu W, Lao Y, Huang N, Cui K, Liao M, He J, Jiang Y, Yang BB, Xu H, Xu N, Zhang Y - Autophagy (2013)

Bottom Line: MIR155 suppresses target-gene expression by directly interacting with their 3' untranslated regions (UTRs), mutations of the binding sites abolish their MIR155 responsiveness.Furthermore, by downregulating MTOR signaling, MIR155 also attenuates cell proliferation and induces G 1/S cell cycle arrest.Collectively, these data present a new role for MIR155 as a key regulator of autophagy via dysregulation of MTOR pathway.

View Article: PubMed Central - PubMed

Affiliation: School of Life Sciences; Tsinghua University; Beijing, China; Key Lab in Healthy Science and Technology; Division of Life Science; Graduate School at Shenzhen; Tsinghua University; Shenzhen, China.

ABSTRACT
Hypoxia activates autophagy, an evolutionarily conserved cellular catabolic process. Dysfunction in the autophagy pathway has been implicated in an increasing number of human diseases, including cancer. Hypoxia induces upregulation of a specific set of microRNAs (miRNAs) in a variety of cell types. Here, we describe hypoxia-induced MIR155 as a potent inducer of autophagy. Enforced expression of MIR155 increases autophagic activity in human nasopharyngeal cancer and cervical cancer cells. Knocking down endogenous MIR155 inhibits hypoxia-induced autophagy. We demonstrated that MIR155 targets multiple players in MTOR signaling, including RHEB, RICTOR, and RPS6KB2. MIR155 suppresses target-gene expression by directly interacting with their 3' untranslated regions (UTRs), mutations of the binding sites abolish their MIR155 responsiveness. Furthermore, by downregulating MTOR signaling, MIR155 also attenuates cell proliferation and induces G 1/S cell cycle arrest. Collectively, these data present a new role for MIR155 as a key regulator of autophagy via dysregulation of MTOR pathway.

Show MeSH

Related in: MedlinePlus

Figure 5.MIR155 directly targets RHEB, RICTOR and RPS6KB2. (A) Predicted binding sequences between MIR155 and seed matches in RHEB, RICTOR, and RPS6KB2 3′ UTRs. (B) Luciferase reporter vectors were generated by inserting the wild-type or mutated 3′UTR fragments of RHEB, RICTOR, and RPS6KB2 into pRL-TK plasmid. Luciferase reporter assays at 24 h after transfection with wild-type (WT) or mutated (MUT) plasmids, cotransfected with NC or MIR155. Data shown are means ± s.d. of independent experiments, n = 4, **P < 0.01, ***P < 0.001, Student 2-tailed t test.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC4389881&req=5

Figure 5: Figure 5.MIR155 directly targets RHEB, RICTOR and RPS6KB2. (A) Predicted binding sequences between MIR155 and seed matches in RHEB, RICTOR, and RPS6KB2 3′ UTRs. (B) Luciferase reporter vectors were generated by inserting the wild-type or mutated 3′UTR fragments of RHEB, RICTOR, and RPS6KB2 into pRL-TK plasmid. Luciferase reporter assays at 24 h after transfection with wild-type (WT) or mutated (MUT) plasmids, cotransfected with NC or MIR155. Data shown are means ± s.d. of independent experiments, n = 4, **P < 0.01, ***P < 0.001, Student 2-tailed t test.

Mentions: To clarify if MIR155 regulates the expression of target genes via direct interaction with their 3′ UTRs, we used 2 bioinformatics softwares TargetScan and FindTar to predict the potential binding sequences in the 3′ UTRs of target genes (Fig. 5A). Then we designed pRL-TK-luciferase reporter constructs containing either the wild-type (WT) or mutated (MUT) MIR155 binding sequences in the 3′ UTRs of RHEB, RICTOR and RPS6KB2. The pRL-TK-luciferase reporter constructs were cotransfected with either MIR155 or NC control. Luciferase activity was determined at 24 h after transfection. MIR155 significantly reduced luciferase activity when cells transfected with wild-type 3′ UTRs of RHEB, RICTOR and RPS6KB2. By contrast, the inhibitory effect of MIR155 on luciferase activity was completely restored by the mutants (Fig. 5B). Moreover, the luciferase activity was also susceptible to knockdown of endogenous MIR155. Inhibition of MIR155 could specifically relieve the repression of all 3 reporters (Fig. S4).


Hypoxia-induced MIR155 is a potent autophagy inducer by targeting multiple players in the MTOR pathway.

Wan G, Xie W, Liu Z, Xu W, Lao Y, Huang N, Cui K, Liao M, He J, Jiang Y, Yang BB, Xu H, Xu N, Zhang Y - Autophagy (2013)

Figure 5.MIR155 directly targets RHEB, RICTOR and RPS6KB2. (A) Predicted binding sequences between MIR155 and seed matches in RHEB, RICTOR, and RPS6KB2 3′ UTRs. (B) Luciferase reporter vectors were generated by inserting the wild-type or mutated 3′UTR fragments of RHEB, RICTOR, and RPS6KB2 into pRL-TK plasmid. Luciferase reporter assays at 24 h after transfection with wild-type (WT) or mutated (MUT) plasmids, cotransfected with NC or MIR155. Data shown are means ± s.d. of independent experiments, n = 4, **P < 0.01, ***P < 0.001, Student 2-tailed t test.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4389881&req=5

Figure 5: Figure 5.MIR155 directly targets RHEB, RICTOR and RPS6KB2. (A) Predicted binding sequences between MIR155 and seed matches in RHEB, RICTOR, and RPS6KB2 3′ UTRs. (B) Luciferase reporter vectors were generated by inserting the wild-type or mutated 3′UTR fragments of RHEB, RICTOR, and RPS6KB2 into pRL-TK plasmid. Luciferase reporter assays at 24 h after transfection with wild-type (WT) or mutated (MUT) plasmids, cotransfected with NC or MIR155. Data shown are means ± s.d. of independent experiments, n = 4, **P < 0.01, ***P < 0.001, Student 2-tailed t test.
Mentions: To clarify if MIR155 regulates the expression of target genes via direct interaction with their 3′ UTRs, we used 2 bioinformatics softwares TargetScan and FindTar to predict the potential binding sequences in the 3′ UTRs of target genes (Fig. 5A). Then we designed pRL-TK-luciferase reporter constructs containing either the wild-type (WT) or mutated (MUT) MIR155 binding sequences in the 3′ UTRs of RHEB, RICTOR and RPS6KB2. The pRL-TK-luciferase reporter constructs were cotransfected with either MIR155 or NC control. Luciferase activity was determined at 24 h after transfection. MIR155 significantly reduced luciferase activity when cells transfected with wild-type 3′ UTRs of RHEB, RICTOR and RPS6KB2. By contrast, the inhibitory effect of MIR155 on luciferase activity was completely restored by the mutants (Fig. 5B). Moreover, the luciferase activity was also susceptible to knockdown of endogenous MIR155. Inhibition of MIR155 could specifically relieve the repression of all 3 reporters (Fig. S4).

Bottom Line: MIR155 suppresses target-gene expression by directly interacting with their 3' untranslated regions (UTRs), mutations of the binding sites abolish their MIR155 responsiveness.Furthermore, by downregulating MTOR signaling, MIR155 also attenuates cell proliferation and induces G 1/S cell cycle arrest.Collectively, these data present a new role for MIR155 as a key regulator of autophagy via dysregulation of MTOR pathway.

View Article: PubMed Central - PubMed

Affiliation: School of Life Sciences; Tsinghua University; Beijing, China; Key Lab in Healthy Science and Technology; Division of Life Science; Graduate School at Shenzhen; Tsinghua University; Shenzhen, China.

ABSTRACT
Hypoxia activates autophagy, an evolutionarily conserved cellular catabolic process. Dysfunction in the autophagy pathway has been implicated in an increasing number of human diseases, including cancer. Hypoxia induces upregulation of a specific set of microRNAs (miRNAs) in a variety of cell types. Here, we describe hypoxia-induced MIR155 as a potent inducer of autophagy. Enforced expression of MIR155 increases autophagic activity in human nasopharyngeal cancer and cervical cancer cells. Knocking down endogenous MIR155 inhibits hypoxia-induced autophagy. We demonstrated that MIR155 targets multiple players in MTOR signaling, including RHEB, RICTOR, and RPS6KB2. MIR155 suppresses target-gene expression by directly interacting with their 3' untranslated regions (UTRs), mutations of the binding sites abolish their MIR155 responsiveness. Furthermore, by downregulating MTOR signaling, MIR155 also attenuates cell proliferation and induces G 1/S cell cycle arrest. Collectively, these data present a new role for MIR155 as a key regulator of autophagy via dysregulation of MTOR pathway.

Show MeSH
Related in: MedlinePlus