Shufeng Li1, Qian Qiang2, Haitao Shan2, Minke Shi3, Guangming Gan2, Fang Ma2, Baojun Chen4. 1. Key Laboratory of Developmental Genes and Human Disease in Ministry of Education, Department of Biochemistry and Molecular Biology, Medical School of Southeast University, Nanjing 210009, China. Electronic address: shufengli@seu.edu.cn. 2. Key Laboratory of Developmental Genes and Human Disease in Ministry of Education, Department of Biochemistry and Molecular Biology, Medical School of Southeast University, Nanjing 210009, China. 3. Department of Thoracic and Cardiovascular Surgery, the Affiliated Drum Tower Hospital of Nanjing University Medical School, Nanjing, China. 4. Department of Thoracic and Cardiovascular Surgery, the Affiliated Drum Tower Hospital of Nanjing University Medical School, Nanjing, China. Electronic address: cbaoj_608@126.com.
Abstract
AIMS: RB1CC1/FIP200 was essential for autophagosome formation. Therefore, RB1CC1/FIP200 cellular levels are critical for the activation of the autophagy pathways. Following the screen of miRNAs affecting RB1CC1/FIP200 level and rapamycin-induced autophagy, we discovered miR-20a and miR-20b could regulate autophagy by targeting RB1CC1/FIP200. MAIN METHODS: Inhibitory effect of miR-20a and 20b on basal and rapamycin-stimulated autophagy was demonstrated using various autophagic tests including GFP-LC3 puncta analysis, LC3II/LC3I gel shift and TEM observation. KEY FINDINGS: We discovered RB1CC1/FIP200 as cellular targets of miR-20a and miR-20b. Upon miR-20a and miR-20b overexpression, both mRNA and protein levels of RB1CC1/FIP200 decreased. miR-20a and miR-20b target sequences present in the 3' UTR of RB1CC1/FIP200 mRNAs and introduction of mutations abolished the miR-20a and miR-20b responsiveness. In MCF7 and MDA-MB-231 breast cancer cells, miR-20a and miR-20b over-expression attenuated basal and rapamycin-induced autophagy; while suppression of miR-20a or miR-20b by specific antagomir showed normal rapamycin-induced autophagic activity. SIGNIFICANCE: To our knowledge, this is the first study showing the significance of miR-20a and miR-20b regulating autophagy by targeting RB1CC1/FIP200.
AIMS: RB1CC1/FIP200 was essential for autophagosome formation. Therefore, RB1CC1/FIP200 cellular levels are critical for the activation of the autophagy pathways. Following the screen of miRNAs affecting RB1CC1/FIP200 level and rapamycin-induced autophagy, we discovered miR-20a and miR-20b could regulate autophagy by targeting RB1CC1/FIP200. MAIN METHODS: Inhibitory effect of miR-20a and 20b on basal and rapamycin-stimulated autophagy was demonstrated using various autophagic tests including GFP-LC3 puncta analysis, LC3II/LC3I gel shift and TEM observation. KEY FINDINGS: We discovered RB1CC1/FIP200 as cellular targets of miR-20a and miR-20b. Upon miR-20a and miR-20b overexpression, both mRNA and protein levels of RB1CC1/FIP200 decreased. miR-20a and miR-20b target sequences present in the 3' UTR of RB1CC1/FIP200 mRNAs and introduction of mutations abolished the miR-20a and miR-20b responsiveness. In MCF7 and MDA-MB-231 breast cancer cells, miR-20a and miR-20b over-expression attenuated basal and rapamycin-induced autophagy; while suppression of miR-20a or miR-20b by specific antagomir showed normal rapamycin-induced autophagic activity. SIGNIFICANCE: To our knowledge, this is the first study showing the significance of miR-20a and miR-20b regulating autophagy by targeting RB1CC1/FIP200.
Authors: Nooshin Hashemi-Sadraei; Gaëlle M Müller-Greven; Fadi W Abdul-Karim; Ilya Ulasov; Erinn Downs-Kelly; Monica E Burgett; Adam Lauko; Maha A Qadan; Robert J Weil; Manmeet S Ahluwalia; Lingling Du; Richard A Prayson; Samuel T Chao; Thomas G Budd; Jill Barnholtz-Sloan; Amy S Nowacki; Ruth A Keri; Candece L Gladson Journal: J Neurooncol Date: 2018-08-09 Impact factor: 4.130