Retinoblastoma protein regulates the crosstalk between autophagy and apoptosis, and favors glioblastoma resistance to etoposide

Abstract

Glioblastomas (GBMs) are devastating tumors of the central nervous system, with a poor prognosis of 1-year survival. This results from a high resistance of GBM tumor cells to current therapeutic options, including etoposide (VP-16). Understanding resistance mechanisms may thus open new therapeutic avenues. VP-16 is a topoisomerase inhibitor that causes replication fork stalling and, ultimately, the formation of DNA double-strand breaks and apoptotic cell death. Autophagy has been identified as a VP-16 treatment resistance mechanism in tumor cells. Retinoblastoma protein (RB) is a classical tumor suppressor owing to its role in G1/S cell cycle checkpoint, but recent data have shown RB participation in many other cellular functions, including, counterintuitively, negative regulation of apoptosis. As GBMs usually display an amplification of the EGFR signaling involving the RB protein pathway, we questioned whether RB might be involved in mechanisms of resistance of GBM cells to VP-16. We observed that RB silencing increased VP-16-induced DNA double-strand breaks and p53 activation. Moreover, RB knockdown increased VP-16-induced apoptosis in GBM cell lines and cancer stem cells, the latter being now recognized essential to resistance to treatments and recurrence. We also showed that VP-16 treatment induced autophagy, and that RB silencing impaired this process by inhibiting the fusion of autophagosomes with lysosomes. Taken together, our data suggest that RB silencing causes a blockage on the VP-16-induced autophagic flux, which is followed by apoptosis in GBM cell lines and in cancer stem cells. Therefore, we show here, for the first time, that RB represents a molecular link between autophagy and apoptosis, and a resistance marker in GBM, a discovery with potential importance for anticancer treatment.

The Drosophila retinoblastoma protein, Rbf1, induces a debcl and drp1-dependent mitochondrial apoptosis.

“…However, the role of pRb in cancer is not limited to cell cycle regulation and appears to be more complex. Indeed, pRb can be pro-apoptotic (Bowen et al, 1998;Hilgendorf et al, 2013;Ianari et al, 2009) or anti-apoptotic (Biasoli et al, 2013;Morgenbesser et al, 1994;Tsai et al, 1998), and the regulation of these opposite roles of pRb towards apoptosis remains poorly understood. The ability of pRb to induce apoptosis fits with its tumor suppressor function.…”

TAp63gamma is required for the late stages of myogenesis

“…[9][10][11][12][13][14] While early myogenesis markers can be also expressed in proliferating cells, late differentiation markers, like myosin heavy chain (MHC), are induced only after cell cycle arrest. 15 The retinoblastoma protein (Rb) and cyclin D, plays a critical role in myoblasts cycle arrest; indeed myocytes and muscle cells lacking Rb fail to exit the cell cycle, [16][17][18][19][20][21][22][23][24][25][26] altering the late phases of skeletal myogenesis. In addition, the cyclin-dependent-kinase inhibitors p21…”

Inhibition of pRB Pathway Differentially Modulates Apoptosis in Esophageal Cancer Cells

“…However, the autophagy induced by 5-FU can function either as a survival or as a death mechanism, apparently depending on the combination of genetic alterations in each cell [40], [41], [42]. We and others have previously shown that pRB regulates the completion of the autophagic process induced by anticancer drugs [13], [18], [21], [43].…”

“…The extracted protein (containing 15–40 μg of protein per sample) was diluted in sample buffer (10% SDS; 10 mM β-mercaptoethanol; 20% glycerol; 0.2 M Tris–HCl, pH 6.8; and 0.05% bromophenol blue) and electrophoresed in polyacrylamide gel (SDS-PAGE) according to our previous paper [13]. The antibodies used were polyclonal anti-RB 851 (1:5000, kindly given by Dr. Jean Y. J. Wang), phospho-RB at Ser807/811 (1:1000, Cell Signaling Technology), and anti-α-tubulin (1:10,000, Sigma-Aldrich).…”