Cellular unfolded protein response against viruses used in gene therapy

Abstract

Viruses are excellent vehicles for gene therapy due to their natural ability to infect and deliver the cargo to specific tissues with high efficiency. Although such vectors are usually “gutted” and are replication defective, they are subjected to clearance by the host cells by immune recognition and destruction. Unfolded protein response (UPR) is a naturally evolved cyto-protective signaling pathway which is triggered due to endoplasmic reticulum (ER) stress caused by accumulation of unfolded/misfolded proteins in its lumen. The UPR signaling consists of three signaling pathways, namely PKR-like ER kinase, activating transcription factor 6, and inositol-requiring protein-1. Once activated, UPR triggers the production of ER molecular chaperones and stress response proteins to help reduce the protein load within the ER. This occurs by degradation of the misfolded proteins and ensues in the arrest of protein translation machinery. If the burden of protein load in ER is beyond its processing capacity, UPR can activate pro-apoptotic pathways or autophagy leading to cell death. Viruses are naturally evolved in hijacking the host cellular translation machinery to generate a large amount of proteins. This phenomenon disrupts ER homeostasis and leads to ER stress. Alternatively, in the case of gutted vectors used in gene therapy, the excess load of recombinant vectors administered and encountered by the cell can trigger UPR. Thus, in the context of gene therapy, UPR becomes a major roadblock that can potentially trigger inflammatory responses against the vectors and reduce the efficiency of gene transfer.

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“…Our results are consistent with a previous study reporting that BiP overexpression attenuates PERK and IRE1 activation, whereas a reduced level of BiP activates the UPR [49,50]. When the ER stress cannot restore the cellular homeostasis, C/EBP homologous protein (CHOP) is activated, leading to cell apoptosis [34,51]. Indeed, we found that CHOP expression was activated in response to VV-Onco induced cell stress.…”

“…The major ER chaperone binding immunoglobulin protein (BiP), also known as glucose-regulated protein78 (GRP78), is the master regulator of the UPR [14,34]. The anti-apoptotic property and cytoprotective function of BiP is well established [36][37][38][39].…”

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“…UPR有3条信号通路分支, 分别由3种内质网 驻膜蛋白所介导, 即IRE1(inositol-requiring protein 1), ATF6(activating transcription factor 6) 和 PERK(protein kinase RNA-like ER kinase). 在正常细胞中, 这3种重要 的信号分子都能够与内质网主要分子伴侣BiP(binding 起始的重要调节因子, 激活的eIF2α会抑制蛋白质的 合成 [51,52] .…”

“…痘苗病毒的K3L蛋 白由于与eIF2α序列具有约28%的一致性, 因此会被 eIF2α的激酶PKR(pancreatic eIF2α kinase)当做底物从 而抑制了PKR的活性, 阻止了eIF2α的磷酸化. 进一步 研究显示, K3L也能抑制PERK(PKR-like ER kinase)的 活性 [51,55] . 此外, 在RK13和HeLa细胞中, 痘苗病毒K1 蛋白能够抑制病毒诱导的eIF2α的磷酸化和PKR的磷 酸化 [56] , 这些有利于蛋白质的重新合成.…”

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“…This contrasts with most gene therapy approaches which replace functional loss with a WT gene product but do not remove the mutant, allowing the continued impact on the UPR due to the mutant and the viral vector used to insert the replacement gene, even though function may be restored [35]. …”