Various structural and non-structural HCV proteins were shown to localize inside the ER lumen or integrated FTY720 manufacturer to ER membrane and to undergo ER-specific modifications [5], [6]. The ER is the cellular site for folding and modification of membrane-bound and secreted proteins. The flux of newly synthesized proteins into the ER is dynamic. Conditions of ER stress occur when the amount of proteins entering the ER exceeds its folding capacity. This imbalance induces a cyto-protective signaling cascade collectively termed the unfolded protein response (UPR) [7], [8]. The mammalian UPR operates in several parallel pathways, whose sensors are IRE1, PERK and ATF6. When misfolded proteins accumulate in the ER, IRE1 is activated by trans-autophosphorylation and splices the mRNA of XBP-1, which converts the unspliced XBP-1 (XBP-1u), a highly unstable protein into its spliced form (XBP-1s).

XBP-1s is a potent transcription factor responsible for the transcriptional activation of the majority of UPR target genes [9]. PERK, a second ER stress sensor, is activated in a similar fashion to IRE1 and is responsible for the modulation of protein synthesis in response to ER stress. Activated PERK phosphorylates eIF2�� and leads to general suppression of protein synthesis and activation of the integrated stress response [10], [11]. ATF6, the third sensor, operates both as a stress sensor and as a transcription factor. In response to ER stress, ATF6 translocates from the ER to the Golgi apparatus, where its N-terminus is released from the membrane by regulated intramembrane proteolysis [12].

Activation of the UPR leads to a coordinated and highly regulated response which improves the efficiency of protein folding, processing and export. In order to reduce protein load in the ER, the UPR attenuates the flow of proteins into the ER and decreases protein synthesis, while up-regulating the expression of chaperones. Finally, it removes misfolded proteins via activation of ER associated degradation (ERAD). The UPR is intimately linked to the apoptotic machinery. Both IRE1 and PERK, when strongly activated, lead to apoptosis [13], [14]. While acute ER stress conditions signal for cell death, chronic sub-lethal ER stress causes cellular adaptation and eventually resistance to apoptosis in a mechanism, which was suggested to involve modulation of mRNA localization and stability of UPR components [15].

It was previously shown that in cells stably expressing the HCV replicon, XBP-1 mRNA undergoes splicing, indicative of ER stress. However the expression of specific target genes downstream to XBP-1s responsible for enhanced degradation of misfolded Carfilzomib ER proteins was not elevated. These results suggested that HCV manipulates the IRE1-XBP-1 pathway of the UPR. It was further shown that ER stress develops in response to expression of specific HCV proteins such as the core protein E1, E2, NS2 and NS4B [16]�C[19].