The endoplasmic reticulum (ER) has recently emerged as an alternative target to induce cell death in tumours, because prolonged ER stress results in the induction of apoptosis even in chemoresistant transformed cells. Here, we show that the DNAdamage-responsive pro-apoptotic factor E2F1 is unexpectedly downregulated during the ER stress-mediated apoptotic programme. E2F1 decline is a late event during the ER response and is mediated by the two unfolded protein response (UPR) sensors ATF6 and IRE1 (also known as ERN1). Whereas ATF6 directly interacts with the E2F1 promoter, IRE1 requires the involvement of the known E2F1 modulator E2F7, through the activation of its main target Xbp-1. Importantly, inhibition of the E2F1 decrease prevents ER-stress-induced apoptosis, whereas E2F1 knockdown efficiently sensitises cells to ER stress-dependent apoptosis, leading to the upregulation of two main factors in the UPR pro-apoptotic execution phase, Puma and Noxa (also known as BBC3 and PMAIP1, respectively). Our results point to a novel key role of E2F1 in the cell survival/death decision under ER stress, and unveil E2F1 inactivation as a valuable novel potential therapeutic strategy to increase the response of tumour cells to ER stress-based anticancer treatments.

The endoplasmic reticulum (ER) consists of the nuclear envelope and a peripheral network of tubules and membrane sheets. The tubules are shaped by a specific class of curvature stabilizing proteins, the reticulons and DP1; however it is still unclear how the sheets are assembled. The ER is the cellular compartment responsible for secretory and membrane protein synthesis. The reducing conditions of ER lead to the intra/inter-chain formation of new disulphide bonds into polypeptides during protein folding assessed by enzymatic or spontaneous reactions. Moreover, ER represents the main intracellular calcium storage site and it plays an important role in calcium signaling that impacts many cellular processes. Accordingly, the maintenance of ER function represents an essential condition for the cell, and ER morphology constitutes an important prerogative of it. Furthermore, it is well known that ER undergoes prominent shape transitions during events such as cell division and differentiation. Thus, maintaining the correct ER structure is an essential feature for cellular physiology. Now, it is known that proper ER-associated proteins play a fundamental role in ER tubules formation. Among these ER-shaping proteins are the reticulons (RTN), which are acquiring a relevant position. In fact, beyond the structural role of reticulons, in very recent years new and deeper functional implications of these proteins are emerging in relation to their involvement in several cellular processes.

Endoplasmic reticulum (ER) is the primary site for the synthesis and folding of secreted and membrane-bound proteins. Accumulation of unfolded and misfolded proteins in ER underlies a wide range of human neurodegenerative disorders. Hence, molecules regulating the ER stress response represent potential candidates as drug targets for tackling these diseases. Protein disulphide isomerase (PDI) is a chaperone involved in ER stress pathway, its activity being an important cellular defense against protein misfolding. Here, we demonstrate that human neuroblastoma SH-SY5Y cells overexpressing the reticulon protein 1-C (RTN1-C) reticulon family member show a PDI punctuate subcellular distribution identified as ER vesicles. This represents an event associated with a significant increase of PDI enzymatic activity. We provide evidence that the modulation of PDI localization and activity does not only rely upon ER stress induction or upregulation of its synthesis, but tightly correlates to an alteration in its nitrosylation status. By using different RTN1-C mutants, we demonstrate that the observed effects depend on RTN1-C N-terminal region and on the integrity of the microtubule network. Overall, our results indicate that RTN1-C induces PDI redistribution in ER vesicles, and concomitantly modulates its activity by decreasing the levels of its S-nitrosylated form. Thus RTN1-C represents a promising candidate to modulate PDI function.