MG132 plus apoptosis antigen-1 (APO-1) antibody cooperate to restore p53 activity inducing autophagy and p53-dependent apoptosis in HPV16 E6-expressing keratinocytes

Abstract

The E6 oncoprotein can interfere with the ability of infected cells to undergo programmed cell death through the proteolytic degradation of proapoptotic proteins such as p53, employing the proteasome pathway. Therefore, inactivation of the proteasome through MG132 should restore the activity of several proapoptotic proteins. We investigated whether in HPV16 E6-expressing keratinocytes (KE6 cells), the restoration of p53 levels mediated by MG132 and/or activation of the CD95 pathway through apoptosis antigen-1 (APO-1) antibody are responsible for the induction of apoptosis. We found that KE6 cells underwent apoptosis mainly after incubation for 24 h with MG132 alone or APO-1 plus MG132. Both treatments activated the extrinsic and intrinsic apoptosis pathways. Autophagy was also activated, principally by APO-1 plus MG132. Inhibition of E6-mediated p53 proteasomal degradation by MG132 resulted in the elevation of p53 protein levels and its phosphorylation in Ser46 and Ser20; the p53 protein was localized mainly at nucleus after treatment with MG132 or APO-1 plus MG132. In addition, induction of its transcriptional target genes such as p21, Bax and TP53INP was observed 3 and 6 h after treatment. Also, LC3 mRNA was induced after 3 and 6 h, which correlates with lipidation of LC3B protein and induction of autophagy. Finally, using pifithrin alpha we observed a decrease in apoptosis induced by MG132, and by APO-1 plus MG132, suggesting that restoration of APO-1 sensitivity occurs in part through an increase in both the levels and the activity of p53. The use of small molecules to inhibit the proteasome pathway might permit the activation of cell death, providing new opportunities for CC treatment.

High cell density increases glioblastoma cell viability under glucose deprivation via degradation of the cystine/glutamate transporter xCT (SLC7A11)

“…It has been suggested that there is crosstalk between the lysosomal and the proteasomal degradation systems (42,43). For example, proteasomal inhibitors induce compensatory actions to cause autophagy and lysosomal degradation (44)(45)(46). On the other hand, lysosomal inhibition was found to increase or reduce proteasomal activity (47,48).…”

Crosstalk and Interplay between the Ubiquitin-Proteasome System and Autophagy

“…Proteasome inhibition can induce autophagy via the metabolic stabilization of the transcriptional factor p53 ( Gu et al, 2014 ; Lagunas-Martinez et al, 2017 ). Upon accumulation, the nuclear subpopulation of p53 acts as a transcription factor for autophagy housekeeping genes such as damage-regulated autophagy modifier (DRAM) ( Mrschtik et al, 2015 ; Zhang et al, 2013 ).…”

Associations between Huwe1 and autophagy in rat cerebral neuron oxygen‑glucose deprivation and reperfusion injury

“…Huwe1 is involved in the ubiquitination and degradation of multiple proteins, including P53 (19). Proteasome inhibition in neurons has been shown to induce autophagy via the metabolic stabilization of the transcription factor P53 (9,61,74). recently, it has been revealed that apoptosis signaling exhibits substantial molecular crosstalk with the uPS, as well as autophagy.…”

“…However, accumulating evidence has revealed crosstalk between the two degradation pathways (6,7). Previous studies investigating this crosstalk have focused on the compensatory and complementary relationship between autophagy and the uPS (7)(8)(9). it has been shown that uPS dysfunction can activate autophagy in vitro in cell culture and in vivo in animal models via the direct or indirect modulation of proteins associated with cell survival and death (10)(11)(12).…”