<i>MIR93</i> (<i>microRNA -93</i>) regulates tumorigenicity and therapy response of glioblastoma by targeting autophagy

Macroautophagy/autophagy is a natural intracellular process that maintains cellular homeostasis and protects cells from death under stress conditions. Autophagy sustains tumor survival and growth when induced by common cancer treatments, including IR and cytotoxic chemotherapy, thereby contributing to therapeutic resistance of tumors. In this study, we report that the expression of MIR93, noted in two clinically relevant tumor subtypes of GBM, influenced GSC phenotype as well as tumor response to therapy through its effects on autophagy. Our mechanistic studies revealed that MIR93 regulated autophagic activities in GSCs through simultaneous inhibition of multiple autophagy regulators, including BECN1/Beclin 1, ATG5, ATG4B, and SQSTM1/p62. Moreover, two first-line treatments for GBM, IR and temozolomide (TMZ), as well as rapamycin (Rap), the prototypic MTOR inhibitor, decreased MIR93 expression that, in turn, stimulated autophagic processes in GSCs. Inhibition of autophagy by ectopic MIR93 expression, or via autophagy inhibitors NSC (an ATG4B inhibitor) and CQ, enhanced the activity of IR and TMZ against GSCs. Collectively, our findings reveal a key role for MIR93 in the regulation of autophagy and suggest a combination treatment strategy involving the inhibition of autophagy while administering cytotoxic therapy. Abbreviations: ACTB: actin beta; ATG4B: autophagy related 4B cysteine peptidase; ATG5: autophagy related 5; BECN1: beclin 1; CL: classical; CQ: chloroquine diphosphate; CSCs: cancer stem cells; GBM: glioblastoma; GSCs: glioma stem-like cells; HEK: human embryonic kidney; IB: immunoblotting; IF: immunofluorescent staining; IR: irradiation; MAP1LC3/LC3: microtubule associated protein 1 light chain 3; MES: mesenchymal; MIR93: microRNA 93; MIRC: a control miRNA; miRNA/miR: microRNA; MTOR: mechanistic target of rapamycin kinase; NSC: NSC185085; PN: proneural; qRT-PCR: quantitative reverse transcription-polymerase chain reaction; Rap: rapamycin; SQSTM1/p62: sequestosome 1; TCGA: the cancer genome atlas; TMZ: temozolomide; WT: wild type; ZIP93: lentiviral miRZIP targeting MIR93; ZIPC: lentiviral miRZip targeting control miRNA</p

Supplementary Table S4 from SRSF3-Regulated RNA Alternative Splicing Promotes Glioblastoma Tumorigenicity by Affecting Multiple Cellular Processes

List of SRSF3-affected and -correlated AS events in GSCs</p

Supplementary Table S2 from SRSF3-Regulated RNA Alternative Splicing Promotes Glioblastoma Tumorigenicity by Affecting Multiple Cellular Processes

List of SRSF3-affected AS events in GSCs</p

Supplementary Data from SRSF3-Regulated RNA Alternative Splicing Promotes Glioblastoma Tumorigenicity by Affecting Multiple Cellular Processes

This files contains supplemental figures related to Figures 1-3, 4, and 6, and four supplemental tables, and supplemental materials and methods.</p

Supplementary Figure 5 from Identification of miR-133b and RB1CC1 as Independent Predictors for Biochemical Recurrence and Potential Therapeutic Targets for Prostate Cancer

PDF file - 234KB, Identifying putative targets of miR-133b.</p

Supplementary Figure 1 from Identification of miR-133b and RB1CC1 as Independent Predictors for Biochemical Recurrence and Potential Therapeutic Targets for Prostate Cancer

PDF file - 167KB, The expression of PSA and KLK2 in DHT treated LNCaP cells and the transfection efficiency of cell proliferation in prostate cancer cells.</p

Supplementary Table S1 from SRSF3-Regulated RNA Alternative Splicing Promotes Glioblastoma Tumorigenicity by Affecting Multiple Cellular Processes

Primers and gRNA sequences</p

Supplementary Table S3 from SRSF3-Regulated RNA Alternative Splicing Promotes Glioblastoma Tumorigenicity by Affecting Multiple Cellular Processes

List of differentially expressed genes after SRSF3-KO in GSCs</p

Supplementary Materials and Methods, Figure Legends, Tables 1 - 9 from Identification of miR-133b and RB1CC1 as Independent Predictors for Biochemical Recurrence and Potential Therapeutic Targets for Prostate Cancer

PDF file - 183KB, Table S1. Correlation between miR-133b expression and RB1CC1 protein level and clinicopathologic features in prostate cancer patients after radical prostatectomy (RP). Table S2. 5-year biochemical recurrence (BCR)-free survival rates and mean BCR-free time for negative and positive miR-133b expression (a) and RB1CC1 expression (b) groups of prostate cancer patients after RP. Table S3. Sequences of synthetic oligonucleotides. Table S4. Primers used for real-time PCR amplification. Table S5. Locations and primer pairs for candidate AREs of miR-133b and negative control (a region of the DNA adjacent to miR-133b gene without a putative ARE). Table S6. Primers used for positive and negative DNA controls of ChIP assay. Table S7. Primers used for cloning the wild-type and mutation of 3'-UTR of RB1CC1 gene into pGL3-promoter Luciferase vector downstream of the Luciferase gene. Table S8. Pearson's rank correlation coefficient analysis. Table S9. Spearman's rank correlation coefficient analysis.</p

Supplementary Figure 4 from Identification of miR-133b and RB1CC1 as Independent Predictors for Biochemical Recurrence and Potential Therapeutic Targets for Prostate Cancer

PDF file - 458KB, Quantitative analyses of activated caspase and caspase-3 in LNCaP and PC-3 cells and the transfection efficiency in these two cell lines.</p