scholarly journals CRISPR/Cas9 and Transgene Verification of Gene Involvement in Unfolded Protein Response and Recombinant Protein Production in Barley Grain

2021 ◽  
Vol 12 ◽  
Author(s):  
Michael Panting ◽  
Inger Baeksted Holme ◽  
Jón Már Björnsson ◽  
Yingxin Zhong ◽  
Henrik Brinch-Pedersen
Keyword(s):  
Recombinant Protein ◽  
Er Stress ◽  
Unfolded Protein Response ◽  
Recombinant Proteins ◽  
Recombinant Protein Production ◽  
Protein Production ◽  
Protein Accumulation ◽  
Unfolded Protein ◽  
Protein Response ◽  
Positive Effect

The use of plants as heterologous hosts to produce recombinant proteins has some intriguing advantages. There is, however, the potential of overloading the endoplasmic reticulum (ER) capacity when producing recombinant proteins in the seeds. This leads to an ER-stress condition and accumulating of unfolded proteins. The unfolded protein response (UPR) is activated to alleviate the ER-stress. With the aim to increase the yield of human epidermal growth factor (EGF) and mouse leukemia inhibitory factor (mLIF) in barley, we selected genes reported to have increased expression during ER-induced stress. The selected genes were calreticulin (CRT), protein disulfide isomerase (PDI), isopentenyl diphosphate isomerase (IPI), glutathione-s-transferase (GST), HSP70, HSP26, and HSP16.9. These were knocked out using CRISPR/Cas9 or overexpressed by conventional transgenesis. The generated homozygous barley lines were crossed with barley plants expressing EGF or mLIF and the offspring plants analyzed for EGF and mLIF protein accumulation in the mature grain. All manipulated genes had an impact on the expression of UPR genes when plantlets were subjected to tunicamycin (TN). The PDI knockout plant showed decreased protein body formation, with protein evenly distributed in the cells of the endosperm. The two genes, GST and IPI, were found to have a positive effect on recombinant protein production. mLIF expression was increased in a F2 homozygous GST knockout mutant background as compared to a F2 GST wild-type offspring. The overexpression of IPI in a F1 cross showed a significant increase in EGF expression. We demonstrate that manipulation of UPR related genes can have a positive effect on recombinant protein accumulation.

scholarly journals How to design an optimal sensor network for the unfolded protein response

2018 ◽  
Vol 29 (25) ◽  
pp. 3052-3062 ◽  
Author(s):  
Wylie Stroberg ◽  
Hadar Aktin ◽  
Yonatan Savir ◽  
Santiago Schnell
Keyword(s):  
Er Stress ◽  
Unfolded Protein Response ◽  
Cellular Protein ◽  
Protein Accumulation ◽  
Protein Homeostasis ◽  
Unfolded Protein ◽  
Stress Sensing ◽  
Coarse Model ◽  
The Unfolded Protein Response ◽  
Protein Response

Cellular protein homeostasis requires continuous monitoring of stress in the endoplasmic reticulum (ER). Stress-detection networks control protein homeostasis by mitigating the deleterious effects of protein accumulation, such as aggregation and misfolding, with precise modulation of chaperone production. Here, we develop a coarse model of the unfolded protein response in yeast and use multi-objective optimization to determine which sensing and activation strategies optimally balance the trade-off between unfolded protein accumulation and chaperone production. By comparing a stress-sensing mechanism that responds directly to the level of unfolded protein in the ER to a mechanism that is negatively regulated by unbound chaperones, we show that chaperone-mediated sensors are more efficient than sensors that detect unfolded proteins directly. This results from the chaperone-mediated sensor having separate thresholds for activation and deactivation. Finally, we demonstrate that a sensor responsive to both unfolded protein and unbound chaperone does not further optimize homeostatic control. Our results suggest a strategy for designing stress sensors and may explain why BiP-mitigated ER stress-sensing networks have evolved.

scholarly journals Crosstalk Between Endoplasmic Reticulum Stress and The Unfolded Protein Response During ZIKA Virus Infection

2019 ◽  
Author(s):  
Jonathan Turpin ◽  
Etienne Frumence ◽  
Wissal Harrabi ◽  
Chaker El Kalamouni ◽  
Philippe Desprès ◽  
...  
Keyword(s):  
Endoplasmic Reticulum ◽  
Er Stress ◽  
Host Cell ◽  
Unfolded Protein Response ◽  
Zika Virus ◽  
Special Focus ◽  
Protein Accumulation ◽  
Unfolded Protein ◽  
Cell Defense ◽  
Protein Response

Flaviviruses replicate in membranous factories associated with the endoplasmic reticulum (ER). The replication rate generates a high polyprotein integration that can contribute to ER stress. Therefore, the host cell can develop an Unfolded Protein Response (UPR) to this protein accumulation which will stimulate appropriate cellular responses in the infectious context (Adaptation, autophagy or cell death). These different stress responses can help to overcome the virus and usually support antiviral strategies initiated by infected cells. In the present study, we investigated the capacity of ZIKA virus (ZIKV) to induce ER stress in epithelial A549 cells with a special focus on the causal factors behind it. We observed that the cells respond to ZIKV infection by implementing an UPR through activation of the IRE1 and PERK pathway but surprisingly without activation of the ATF6 branch. When we examined the effects of modulating the ER stress response we found that UPR inducers significantly inhibit ZIKV replication. Interestingly, our findings provide evidence that ZIKV can altered the UPR in order to escape to the host cell defense system.

scholarly journals How to design an optimal sensor network for the unfolded protein response

2018 ◽  
Author(s):  
Wylie Stroberg ◽  
Hadar Aktin ◽  
Yonatan Savir ◽  
Santiago Schnell
Keyword(s):  
Er Stress ◽  
Unfolded Protein Response ◽  
Cellular Protein ◽  
Protein Accumulation ◽  
Protein Homeostasis ◽  
Unfolded Protein ◽  
Stress Sensing ◽  
Coarse Model ◽  
The Unfolded Protein Response ◽  
Protein Response

AbstractCellular protein homeostasis requires continuous monitoring of stress in the endoplasmic reticulum (ER). Stress detection networks control protein homeostasis by mitigating the deleterious effects of protein accumulation, such as aggregation and misfolding, with precise modulation of chaperone production. Here, we develop a coarse model of the unfolded protein response in yeast, and use multi-objective optimization to determine which sensing and activation strategies optimally balance the trade-off between unfolded protein accumulation and chaperone production. By comparing a stress-sensing mechanism that responds directly to the level of unfolded protein in the ER to a mechanism that is negatively regulated by unbound chaperones, we show that chaperone-mediated sensors are more efficient than sensors that detect unfolded proteins directly. This results from the chaperone-mediated sensor having separate thresholds for activation and deactivation. Lastly, we demonstrate that a sensor responsive to both unfolded protein and unbound chaperone does not further optimize homeostatic control. Our results suggest a strategy for designing stress sensors and may explain why BiP-mitigated ER stress sensing networks have evolved.

scholarly journals Endoplasmic Reticulum Stress Signaling and the Pathogenesis of Hepatocarcinoma

2021 ◽  
Vol 22 (4) ◽  
pp. 1799
Author(s):  
Juncheng Wei ◽  
Deyu Fang
Keyword(s):  
Endoplasmic Reticulum ◽  
Protein Synthesis ◽  
Er Stress ◽  
Unfolded Protein Response ◽  
Response To Treatment ◽  
Protein Accumulation ◽  
Primary Malignancy ◽  
Unfolded Protein ◽  
Protein Response ◽  
Er Homeostasis

Hepatocellular carcinoma (HCC), also known as hepatoma, is a primary malignancy of the liver and the third leading cause of cancer mortality globally. Although much attention has focused on HCC, its pathogenesis remains largely obscure. The endoplasmic reticulum (ER) is a cellular organelle important for regulating protein synthesis, folding, modification and trafficking, and lipid metabolism. ER stress occurs when ER homeostasis is disturbed by numerous environmental, physiological, and pathological challenges. In response to ER stress due to misfolded/unfolded protein accumulation, unfolded protein response (UPR) is activated to maintain ER function for cell survival or, in cases of excessively severe ER stress, initiation of apoptosis. The liver is especially susceptible to ER stress given its protein synthesis and detoxification functions. Experimental data suggest that ER stress and unfolded protein response are involved in HCC development, aggressiveness and response to treatment. Herein, we highlight recent findings and provide an overview of the evidence linking ER stress to the pathogenesis of HCC.

scholarly journals Effectors Targeting the Unfolded Protein Response during Intracellular Bacterial Infection

2021 ◽  
Vol 9 (4) ◽  
pp. 705
Author(s):  
Manal H. Alshareef ◽  
Elizabeth L. Hartland ◽  
Kathleen McCaffrey
Keyword(s):  
Bacterial Infection ◽  
Er Stress ◽  
Unfolded Protein Response ◽  
Host Defense ◽  
Effector Proteins ◽  
Dual Nature ◽  
Unfolded Protein ◽  
Bacterial Replication ◽  
The Unfolded Protein Response ◽  
Protein Response

The unfolded protein response (UPR) is a homeostatic response to endoplasmic reticulum (ER) stress within eukaryotic cells. The UPR initiates transcriptional and post-transcriptional programs to resolve ER stress; or, if ER stress is severe or prolonged, initiates apoptosis. ER stress is a common feature of bacterial infection although the role of the UPR in host defense is only beginning to be understood. While the UPR is important for host defense against pore-forming toxins produced by some bacteria, other bacterial effector proteins hijack the UPR through the activity of translocated effector proteins that facilitate intracellular survival and proliferation. UPR-mediated apoptosis can limit bacterial replication but also often contributes to tissue damage and disease. Here, we discuss the dual nature of the UPR during infection and the implications of UPR activation or inhibition for inflammation and immunity as illustrated by different bacterial pathogens.

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