P53-Dependent PUMA to DRAM antagonistic interplay as a key molecular switch in cell-fate decision in normal/high glucose conditions

Alessia Garufi,Giuseppa Pistritto,Gabriella D’Orazi,Mara Cirone,Gabriele Toietta,Silvia Baldari

doi:10.1186/s13046-017-0596-z

Abstract

BackgroundAs an important cellular stress sensor phosphoprotein p53 can trigger cell cycle arrest and apoptosis and regulate autophagy. The p53 activity mainly depends on its transactivating function, however, how p53 can select one or another biological outcome is still a matter of profound studies. Our previous findings indicate that switching cancer cells in high glucose (HG) impairs p53 apoptotic function and the transcription of target gene PUMA.Methods and resultsHere we report that, in response to drug adriamycin (ADR) in HG, p53 efficiently induced the expression of DRAM (damage-regulated autophagy modulator), a p53 target gene and a stress-induced regulator of autophagy. We found that ADR treatment of cancer cells in HG increased autophagy, as displayed by greater LC3II accumulation and p62 degradation compared to ADR-treated cells in low glucose. The increased autophagy in HG was in part dependent on p53-induced DRAM; indeed DRAM knockdown with specific siRNA reversed the expression of the autophagic markers in HG. A similar outcome was achieved by inhibiting p53 transcriptional activity with pifithrin-α. DRAM knockdown restored the ADR-induced cell death in HG to the levels obtained in low glucose. A similar outcome was achieved by inhibition of autophagy with cloroquine (CQ) or with silencing of autophagy gene ATG5. DRAM knockdown or inhibition of autophagy were both able to re-induce PUMA transcription in response to ADR, underlining a reciprocal interplay between PUMA to DRAM to unbalance p53 apoptotic activity in HG. Xenograft tumors transplanted in normoglycemic mice displayed growth delay after ADR treatment compared to those transplanted in diabetics mice and such different in vivo response correlated with PUMA to DRAM gene expression.ConclusionsAltogether, these findings suggest that in normal/high glucose condition a mutual unbalance between p53-dependent apoptosis (PUMA) and autophagy (DRAM) gene occurred, modifying the ADR-induced cancer cell death in HG both in vitro and in vivo.

Highlights

As an important cellular stress sensor phosphoprotein p53 can trigger cell cycle arrest and apoptosis and regulate autophagy
High glucose (HG) switched the adriamycin (ADR)-induced p53 transcriptional activity from p53 upregulated modulator of apoptosis (PUMA) to Damage-regulated autophagy modulator (DRAM) We first evaluated whether the glucose amount influenced the p53 transcriptional activity in response to drug
The results of reverse transcription polymerase chain reaction (RT-PCR) analyses of mRNA levels show that PUMA was greatly induced by ADR in low glucose (LG), as expected [25], while was not induced in high glucose (HG) (Fig. 1a); on the contrary and opposite to PUMA, DRAM was induced by ADR in HG while was not induced in LG (Fig. 1a)

Summary

Introduction

As an important cellular stress sensor phosphoprotein p53 can trigger cell cycle arrest and apoptosis and regulate autophagy. In response to several types of genotoxic stress the p53 oncosuppressor is activated to control, as transcription factor, genes regulating different cellular outcomes such as cell-cycle arrest and apoptosis [1]. In this manner, p53 protects cells from genomic instability leading to Garufi et al Journal of Experimental & Clinical Cancer Research (2017) 36:126 response, both cooperating in the achievement of clinically relevant effects [5, 6]. The absence of PUMA has been shown to cause high resistance of cancer cells to apoptosis induced by DNA-damaging agents, such as adriamycin, 5-fluorouracil, cisplatin, etc. [11, 12]

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