Activation of PERK-Nrf2 oncogenic signaling promotes Mdm2-mediated Rb degradation in persistently infected HCV culture
Authors: Milad Chedid
Luis A Balart
Yucel Aydin
Krzysztof Moroz
Suchitra Sumitran-Holgersson
Shuanghu Liu
Hua Lu
Curt H Hagedorn
Krzysztof Reiss
Srikanta Dash
Donkita Danielle Williams
Srinivas Chava
Journal: Scientific Reports (2017)
Discipline: Multidisciplinary
Keywords: Chronic hepatitis C virus (HCV) infection; hepatocellular carcinoma (HCC); PERK axis; endoplasmic reticulum (ER) stress; Nrf2
Affiliations: Department of Medicine, Division of Gastroenterology and Hepatology, New Orleans, Louisiana, USA., Department of Pathology and Laboratory Medicine, New Orleans, Louisiana, USA., Department of Medicinal Chemistry, College of Pharmacy, University of Utah, Salt Lake City, UT, USA., Department of Medicine and Genetics, University of Arkansas for Medical Sciences, Little Rock, Arkansas, USA., Laboratory of Transplantation Surgery and Regenerative Medicine, University of Gothenburg, Gothenburg, Sweden., School of Medicine, LSU Health Sciences Center, New Orleans, Louisiana, USA., Department of Biochemistry, Tulane University Health Sciences Center, New Orleans, Louisiana, USA., Department of Pathology and Laboratory Medicine, New Orleans, Louisiana, USA. ., Department of Medicine, Division of Gastroenterology and Hepatology, New Orleans, Louisiana, USA. .

Abstract:  The mechanism of how chronic hepatitis C virus (HCV) infection leads to such a high rate of hepatocellular carcinoma (HCC) is unknown. We found that the PERK axis of endoplasmic reticulum (ER) stress elicited prominent nuclear translocation of Nrf2 in 100% of HCV infected hepatocytes. The sustained nuclear translocation of Nrf2 in chronically infected culture induces Mdm2-mediated retinoblastoma protein (Rb) degradation. Silencing PERK and Nrf2 restored Mdm2-mediated Rb degradation, suggesting that sustained activation of PERK/Nrf2 axis creates oncogenic stress in chronically infected HCV culture model. The activation of Nrf2 and its nuclear translocation were prevented by ER-stress and PERK inhibitors, suggesting that PERK axis is involved in the sustained activation of Nrf2 signaling during chronic HCV infection. Furthermore, we show that HCV clearance induced by interferon-α based antiviral normalized the ER-stress response and prevented nuclear translocation of Nrf2, whereas HCV clearance by DAAs combination does neither. In conclusion, we report here a novel mechanism for how sustained activation of PERK axis of ER-stress during chronic HCV infection activates oncogenic Nrf2 signaling that promotes hepatocyte survival and oncogenesis by inducing Mdm2-mediated Rb degradation. 

Retracted: Yes

PubPeer comments:
  It is improbable that SARS-CoV-2 emerged through laboratory manipulation of a related SARS-CoV-like coronavirus. As noted above, the RBD of SARS-CoV-2 is optimized for binding to human ACE2 with an efficient solution different from those previously predicted 7,11. Furthermore, if genetic manipulation had been performed, one of the several reverse-genetic systems available for betacoronaviruses would probably have been used....Instead, we propose two scenarios that can plausibly explain the origin of SARS-CoV-2: (i) natural selection in an animal host before zoonotic transfer; and (ii) natural selection in humans following zoonotic transfer. Andersen et al are foreclosing on the possibility of direct manipulation of genetic sequences. But the alternatives they propose don t include one that neurobiologist J. Couey, in an [online journal club discussion] , persuasively argues is the most likely one. That would be, not natural, but *artificial* selection in an animal host before zoonotic transfer. Couey:  They don t cite the two most relevant papers. They don t cite the chimera paper from 2015....Already in 2011 there are literally multiple papers showing that we don t need the sequence...just by passing the virus through successive generations in a host the virus will evolve to better infect that host. So if you choose a host with similar receptor homology to us, like a ferret, then the virus that results is likely to also affect humans.  We have tried and true methods of using large populations of other hosts to allow the normal mutation rate and recombination rate of viruses to do all the work for us. Could the authors address this possibility? Update 3 28 In a [second video] addressing this paper, Couey specifies that the authors have neglected to cite highly relevant papers showing a. that  we ve actually used ferrets as an enrichment host to great success with avian flu [(Herfst et al (2012) Science)] and b.  we ve even created a respiratory version of avian flu that infects ferrets... [(IMai et al (2012) Nature] ) He also notes that *we have the genetic techniques to construct chimeric viruses*, citing [Menachery et al (2015) Nature Medicine] ; [Bi et al (2015) PLoS Pathog] , and that, while the highest homology of COVID-19 is with the bat virus, in the vicinity of the spike protein this similarity  crashes off and the sequence bears a much greater resemblance to the pangolin version. Andersen et al s argument seems to boil down to the rather weak assertion that:  If genetic manipulation had been performed, one of several reverse-genetic systems available for betacoronaviruses would probably have been used. Meaning a different one could, and possibly may, have been used. , 
 To even think that somebody or an entity could weaponize a virus by altering its genetic material using conventional reverse-genetic tools and expect not to get caught is absolutely stupid. This is so, especially if such a person or entity really cares about maintaining stealth. Reverse genetic tools would leave fingerprints in the virus that would be picked up even by basic bioinformatics analysis. However, as Lydia has pointed out above, this does not rule out the possibility of laboratory manipulation of SARS-COV-2. Itu2019s possible that a more stealthy approach was employed. Apart from just passaging this virus through a host with similar receptor homology to humans, it s also possible to create animal models that are engineered to carry the human receptors to which this virus binds. The virus would then be allowed to  naturally evolve in these animal models. Viral strains that evolve to efficiently bind to these human receptors would then be isolated and further propagated in the same models and later on, used to inoculate human populations. This would probably save a lot of time. However, what I saw in this article were authors who already made up their minds and strongly believed that this was not a human-customized virus. All they did was to systematically defend this belief, without taking an objective look at other possibilities. It s understandable that they did not want to sound like conspiracy theorists. But it would have helped if they did give this possibility the same weight as their  this virus is not a result of laboratory manipulation position. I strongly believe that they could have done this without necessarily pointing fingers at any government. In fact, where this virus and hence the COVID-19 first appeared does not mean anything to me. Any foreign government with ill intentions could have deployed it. And a city with a biosafety level four laboratory could have been a reasonable target because it would strengthen the narrative that this virus probably escaped from that lab. , 
 The authors present evidence support towards a natural origin for the COVID-19 virus. I agree. ![file] However, it is possible for the virus to have a laboratory origin; it does not mean that the virus was purposefully genetically engineered or that it was purposefully released. There is a novel insertion (polybasic cleavage site) that results in the peptide sequence P-R-R-A, which brings about a host of benefits for the virus, and an increased pathogenicity in the human host. ![file] A GenBank Analysis of PRRA along with the conserved sequences that straddle it, P-R-R-A-R-S show a high degree of similarity with mycoplasma sequences, particularly with those of Toxoplasma gondii, which is a parasite common in cats (particularly those pet cats that are allowed to  free-range ), that in turn infect their human owners. https: www.sciencedaily.com releases 2012 12 121206203240.htm ![file] ![file] Thus, if the virus was serially propagated (against guidelines) in a laboratory, in a human cell line that had mycoplasma contamination (quite possible, as this is a routinely discussed problem in cell-culture labs) or propagated in a human cell line that had Toxoplasma contamination (quite possible if handled by a careless laboratory researcher who had Toxoplasma infection), then it is only matter of time before a virus emerges in culture that has the required pathogenicity to start an epidemic. Analysis of the peptide sequence aligns with numerous mycoplasma common to pigeons, chicken, swine, canines, cattle, and of course Toxoplasma of cats. Although the authors hypothesize a non-laboratory origin for the virus, ![file] ![file] , 
 There is a long write up (non-peer reviewed, non-academic, but very well referenced) on the possibility of a lab origin for the virus. It turns out that  appropriate chimaeric viruses were being created and gain-of-function research carried out, including in the lab in Wuhan. Some of the most similar coronavirus strains (including pangolin) would plausibly have been available to the Wuhan lab, and in the case of the bat virus, was located at the lab and had not been made public. Two arguments against the lab origin are not totally water-tight. Thus, the reported initiation of the epidemic at the Wuhan seafood market seems not to have been verified for all of the initial patients. The relatively large numbers of point mutations differentiating the SARS-COV-2 sequence from the pangolin bat ones suggest some evolutionary distance, but are maybe not beyond the realms of possibility in culture. They latter argument anyway depends upon the complete veracity of the reported bat sequence and requires that no further changes were introduced. As the article concludes, there is certainly no proof at this stage, but some of the arguments against a lab origin are not as convincing as initially announced, and there are some intriguing coincidences, to say the least. https: medium.com @yurideigin lab-made-cov2-genealogy-through-the-lens-of-gain-of-function-research-f96dd7413748 , 
 [ The principal reason for media dismissals of the lab origin possibility is a review paper in Nature Medicine (Andersen et al., 2020). Although by Jun 29 2020 this review had almost 700 citations it also has major scientific shortcomings. ] Link is to a new article in Counterpunch discussing the possible origins of COVID and critiquing the handling of the issue by Andersen et al. , by two experts in relevant fields. ,