Oncogenic H-Ras Up-Regulates Acid β-Hexosaminidase by a Mechanism Dependent on the Autophagy Regulator TFEB

Abstract

The expression of constitutively active H-RasV12 oncogene has been described to induce proliferative arrest and premature senescence in many cell models. There are a number of studies indicating an association between senescence and lysosomal enzyme alterations, e.g. lysosomal β-galactosidase is the most widely used biomarker to detect senescence in cultured cells and we previously reported that H-RasV12 up-regulates lysosomal glycohydrolases enzymatic activity in human fibroblasts. Here we investigated the molecular mechanisms underlying lysosomal glycohydrolase β-hexosaminidase up-regulation in human fibroblasts expressing the constitutively active H-RasV12. We demonstrated that H-Ras activation increases β-hexosaminidase expression and secretion by a Raf/extracellular signal-regulated protein kinase dependent pathway, through a mechanism that relies on the activity of the transcription factor EB (TFEB). Because of the pivotal role of TFEB in the regulation of lysosomal system biogenesis and function, our results suggest that this could be a general mechanism to enhance lysosomal enzymes activity during oncogene-induced senescence.

Extracellular vesicles released by fibroblasts undergoing H-Ras induced senescence show changes in lipid profile

“…HuDe (human dermal fibroblasts) were purchased from the Istituto Zooprofilattico Sperimentale, Brescia, Italy as previously reported [ 17 ]. Cells were cultured in Dulbecco’s modified Eagle’s medium (DMEM) containing 10% (v/v) heat-inactivated fetal bovine serum (FBS), 2 mM glutamine, 100 U/mL penicillin, 100 mg/mL streptomycin and grown at 37°C in a 5% CO 2 .…”

“…Different stimuli can induce cellular senescence; in particular, DNA damage appears to be a key factor. Besides, oncogene activation has been also found to prompt cellular senescence, leading to the definition of peculiar senescence process termed Oncogene-Induced Senescence (OIS) [ 17 ].…”

Metabolic alterations accompanying oncogene-induced senescence

“…demonstrated a significant increase in the transcription factor TFEB that leads to upregulation of HEXA and HEXB, which encode lysosomal glycohydrolase β-hexosaminidase. 76 Taken together, these studies indicate an increase in both autophagy and lysosomal activity in cells undergoing OIS.…”

The update on transcriptional regulation of autophagy in normal and pathologic cells: A novel therapeutic target

“…The transcription factor EB (TFEB) is a master gene for lysosomal biogenesis and also can coordinate autophagosome formation by driving expression of autophagy and lysosomal genes [98][99][100][101][102][103][104][105][106][107][108]. Ebrahimi-Fakhari et al [98] emphasized that further understanding of the biology of TFEB-mediated autophagy will identify novel drug targets and hopefully will provide new therapeutics for treating Parkinson s disease.…”

“…Cortes et al [106] further reported that interference with TFEB signaling could lead to impaired autophagy function and diminished lysosomal degradative capacity, undermining neuronal survival. More persuasive evidence [107,108] was that degradation of long-lived proteins was enhanced by TFEB overexpression, but this response was reduced in TFEB knockout mice, which showed a defect in increase of autophagy.…”