Knock-Down of Mucolipin 1 Channel Promotes Tumor Progression and Invasion in Human Glioblastoma Cell Lines

Abstract

Among cancers that affect the central nervous system, glioblastoma is the most common. Given the negative prognostic significance of transient receptor potential mucolipin 1 (TRPML1) channel reduction in patients with glioblastoma, as discussed in previous publications, the aim of the current study was to investigate the biological advantage of TRPML1 loss for glioma cells. Human glioblastoma primary cancer cells (FSL and FCL) and glioblastoma cell lines (T98 and U251) were used for that purpose. TRPML1 silencing in T98 cells induces defective autophagy, nitric oxide (NO) production, and cathepsin B-dependent apoptosis in the first 48 h and then apoptotic-resistant cells proliferate with a high growth rate with respect to control cells. In U251 cells, knock-down of TRPML1 stimulates NO generation and protein oxidation, arrests cell cycle at G2/M phase, and induces autophagy leading to cathepsin B-dependent senescence. Finally, in both cell lines, the long-term effects of TRPML1 silencing promote survival and invasion capacity with respect to control cells. Silencing of TRPML1 also affects the phenotype of glioblastoma primary cells. FSL cells show increased proliferation ability, while FCL cells enter into senescence associated with an increased invasion ability. In conclusion, although the molecular heterogeneity among different glioblastoma cell lines mirrors the intercellular heterogeneity in cancer cells, our data support TRPML1 downregulation as a negative prognostic factor in glioblastoma.

Functional In Vitro Assessment of VEGFA/NOTCH2 Signaling Pathway and pRB Proteasomal Degradation and the Clinical Relevance of Mucolipin TRPML2 Overexpression in Glioblastoma Patients

“…The pRB protein shows high susceptibility to cytosolic proteases, with CatB-like proteolytic activity [ 44 ]; moreover, we and other researchers [ 43 , 66 ], have previously reported that TRPML1 knock-down results in the leak of mature lysosomal CatB. In this regard, increased levels of mature CatB were evidenced in the cytosol fraction of siTRPML2 T98 cells, but not U251 cells compared with siGLO control cells.…”

Highly Potent and Selective Dopamine D4 Receptor Antagonists Potentially Useful for the Treatment of Glioblastoma

“…GBM cell lines T98 and U251 (grade IV) were obtained as previously described. 49 Cells were grown until 80% of confluence in Eagle’s minimum essential medium (EMEM, Sigma, Merck Life Science S.r.l. Milano, Italy) plus 10% heat-inactivated fetal calf serum (HIFCS, Life Technologies, Monza, Italy), penicillin (100 U/mL), and streptomycin (50 μg/mL) in a humidified atmosphere of 5% CO 2 at 37 °C.…”

The relevance of organelle interactions in cellular senescence

“…The impairment of the function of TRPML1 leads to enlarged lysosomes, lysosomal calcium accumulation, and increased pH, which further contributes to lysosomal malfunction and cellular senescence 119 , 126 . It has been reported that mitochondrial ROS can activate TRPML1, inducing lysosomal calcium release 85 , further inducing autophagy and lysosomal biogenesis 127 and delaying cellular senescence. However, excess ROS levels in the mitochondria have the opposite effect, leading to lysosomal dysfunction and cellular senescence 126 , 127 .…”

“…It has been reported that mitochondrial ROS can activate TRPML1, inducing lysosomal calcium release 85 , further inducing autophagy and lysosomal biogenesis 127 and delaying cellular senescence. However, excess ROS levels in the mitochondria have the opposite effect, leading to lysosomal dysfunction and cellular senescence 126 , 127 .…”