MicroRNAs (miRs) are a class of small non-coding RNAs that function as mediators of gene expression. Dysregulations of miRs have been implicated in the development and progression of glioma. In the present study, we investigated the role of miR-133b in mediating the proliferation and invasion of glioma cells, and the potential mechanism. Real-time RT-PCR results showed that miR-133b expression was significantly decreased in glioma tissues compared with normal brain tissues. Luciferase reporter assay further identified silent information regulator 1 (Sirt1) as a novel direct target of miR-133b in glioma U87 cells. Overexpression of miR-133b suppressed Sirt1 expression and reduced the proliferation and invasion of U87 cells, which could be partly rescued by forced expression of Sirt1. In addition, the Sirt1 mRNA level was significantly higher in glioma tissues than in normal brain tissues, and was inversely correlated with miR-133b level in glioma tissues. In summary, our study sheds light on the regulatory mechanism of miR-133b in glioma growth and metastasis via direct mediation of Sirt1 expression, and suggests that Sirt1 may serve as a potential therapeutic target for glioma.

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Glucose metabolism is an essential process for energy production and cell survival for both normal and abnormal cellular metabolism. Several glucose transporter/ solute carrier 2A (GLUT/SLC2A) superfamily members, including glucose transporter 1 (GLUT1), have been shown to mediate the cellular uptake of glucose in diverse cell types. GLUT1-mediated glucose uptake is a transient and rapid process; thus, the real-time monitoring of GLUT1 trafficking is pivotal for a better understanding of GLUT1 expression and GLUT1-dependent glucose uptake. In the present study, we established a rapid and effective method to visualize the trafficking of GLUT1 between the plasma membrane (PM) and endolysosomal system in live cells using an mCherry-EGFP-GLUT1 tandem fluorescence tracing system. We found that GLUT1 localized at the PM exhibited both red (mCherry) and green (EGFP) fluorescence (yellow when overlapping). However, a significant increase in red punctate fluorescence (mCherry is resistant to acidic pH), but not green fluorescence (EGFP is quenched by acidic pH), was observed upon glucose deprivation, indicating that the mCherry-EGFP-GLUT1 functional protein was trafficked to the acidic endolysosomal system. Besides, we were able to calculate the relative ratio of mCherry to EGFP by quantification of the translocation coefficient, which can be used as a readout for GLUT1 internalization and subsequent lysosomal degradation. Two mutants, mCherry-EGFP-GLUT1-S226D and mCherry-EGFP-GLUT1-ΔC4, were also constructed, which indirectly confirmed the specificity of mCherry-EGFP-GLUT1 for monitoring GLUT1 trafficking. By using a series of endosomal (Rab5, Rab7, and Rab11) and lysosomal markers, we were able to define a model of GLUT1 trafficking in live cells in which upon glucose deprivation, GLUT1 dissociates from the PM and experiences a pH gradient from 6.8−6.1 in the early endosomes to 6.0−4.8 in the late endosomes and finally pH 4.5 in lysosomes, which is appropriate for degradation. In addition, our proof-of-concept study indicated that the pmCherry-EGFP-GLUT1 tracing system can accurately reflect endogenous changes in GLUT1 in response to treatment with the small molecule, andrographolide. Since targeting GLUT1 expression and GLUT1-dependent glucose metabolism is a promising therapeutic strategy for diverse types of cancers and certain other glucose addiction diseases, our study herein indicates that pmCherry-EGFP-GLUT1 can be utilized as a biosensor for GLUT1-dependent functional studies and potential small molecule screening.

Osteosarcoma (OS) is the most common bone malignancy in adolescents and has poor clinical outcomes. Protein arginine methyltransferase 5 (PRMT5) has recently been shown to be aberrantly expressed in various cancers, yet its role in OS remains elusive. Here, we found that PRMT5 was overexpressed in OS and its overexpression predicted poor clinical outcomes. PRMT5 knockdown significantly triggered pronounced senescence in OS cells, as evidenced by the increase in senescence-associated β-galactosidase (SA-β-gal)-stained cells, induction of p21 expression, and upregulation of senescence-associated secretory phenotype (SASP) gene expression. In addition, we found that PRMT5 plays a key role in regulating DNA damaging agents-induced OS cell senescence, possibly, via affecting the repair of DNA damage. Furthermore, we found that TXNIP acts as a key factor mediating PRMT5 depletion-induced DNA damage and cellular senescence. Mechanistically, TRIM21, which interacts with PRMT5, was essential for the regulation of TXNIP/p21 expression. In summary, we propose a model in which PRMT5, by interaction with TRIM21, plays a key role in regulating the TXNIP/p21 axis during senescence in OS cells. The present findings suggest that PRMT5 overexpression in OS cells might confer resistance to chemotherapy and that targeting the PRMT5/TRIM21/TXNIP signaling may enhance the therapeutic efficacy in OS.

p -Aminosalicylic acid (PAS) is an important second-line antibiotic for treating multidrug-resistant tuberculosis (MDR-TB). Due to gastrointestinal disturbance and intolerance, its potent and efficacy in the treatment of extensively drug-resistant (XDR)-TB commonly are poor. Thus, it is important to reveal the mechanism of susceptibility and resistance of Mycobacterium tuberculosis (Mtb) to this drug. Herein, we screened and established PAS-resistant (PAS r ) folC mutated and un-mutated Mtb strains, then utilized a multi-omics (genome, proteome, and metabolome) analysis to better characterize the mechanisms of PAS resistance in Mtb. Interestingly, we found that promotion of SAM-dependent methyltransferases and suppression of PAS uptake via inhibiting some drug transport associated membrane proteins were two key pathways for the folC mutated strain evolving into the PAS r Mtb strain. However, the folC un-mutated strain was resistant to PAS via uptake of exogenous methionine, mitigating the role of inhibitors, and promoting DfrA, ThyA and FolC expression. Beyond these findings, we also found PAS resistance in Mtb might be associated with the increasing phenylalanine metabolism pathway. Collectively, our findings uncovered the differences of resistant mechanism between folC mutated and un-mutated Mtb strains resistant to PAS using multi-omics analysis and targeting modulators to these pathways may be effective for treatment of PAS r Mtb strains.