Gene editing as applied to prevention of reproductive porcine reproductive and respiratory syndrome

Whitworth, Kristin M.; Prather, Randall S.

doi:10.1002/mrd.22811

Cited by 31 publications

(19 citation statements)

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“…Through a clathrin-coated vesicle, PRRSV is delivered into early endosomes (EEs) where it interacts with CD163 to release its genome into the cytoplasm (Whitworth and Prather, 2017). The EE is a major sorting station where the contents are either sent back to the plasma membrane or proceed further down to late endosomes and lysosomes (Gruenberg and Van Der Goot, 2006).…”

Section: Introductionmentioning

confidence: 99%

CD163ΔSRCR5 MARC-145 Cells Resist PRRSV-2 Infection via Inhibiting Virus Uncoating, Which Requires the Interaction of CD163 With Calpain 1

Wei

Dong

et al. 2020

Front. Microbiol.

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Porcine alveolar macrophages without the CD163 SRCR5 domain are resistant to porcine reproductive and respiratory syndrome virus (PRRSV) infection. However, whether the deletion of CD163 SRCR5 in MARC-145 cells confers resistance to PRRSV and interaction of which of the host proteins with CD163 is involved in virus uncoating remain unclear. Here we deleted the SRCR5 domain of CD163 in MARC-145 cells using CRISPR/Cas9 to generate a CD163 SRCR5 MARC-145 cell line. The modification of CD163 had no impact on CD163 expression. CD163 SRCR5 cells were completely resistant to infection by PRRSV-2 strains Li11, CHR6, TJM, and VR2332. The modified cells showed no cytokine response to PRRSV-2 infection and maintained normal cell vitality comparable with the WT cells. The resistant phenotype of the cells was stably maintained through cell passages. There were no replication transcription complexes in the CD163 SRCR5 cells. SRCR5 deletion did not disturb the colocalization of CD163 and PRRSV-N in early endosomes (EE). However, the interaction of the viral proteins GP2a, GP3, or GP5 with CD163, which is involved in virus uncoating was affected. Furthermore, 77 CD163-binding cellular proteins affected by the SRCR5 deletion were identified by LC-MS/MS. Inhibition of calpain 1 trapped the virions in EE and forced then into late endosomes but did not block viral attachment and internalization, suggesting that calpain 1 is involved in the uncoating. Overall, CD163 SRCR5 MARC-145 cells are fully resistant to PRRSV-2 infection and calpain 1 is identified as a novel host protein that interacts with CD163 to facilitate PRRSV uncoating.

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Section: Introductionmentioning

confidence: 99%

CD163ΔSRCR5 MARC-145 Cells Resist PRRSV-2 Infection via Inhibiting Virus Uncoating, Which Requires the Interaction of CD163 With Calpain 1

Wei

Dong

et al. 2020

Front. Microbiol.

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“…Moreover, the key immune mechanisms that we identified may help to focus ongoing research about the role of the GBP5 candidate gene for this resistance SNP, which is currently poorly understood [45]. Furthermore, recent scientific breakthroughs in blocking the permissiveness of porcine alveolar macrophages to some PRRSv strains by gene editing highlight potential new avenues for genetic disease control [46, 47]. Our model suggests that considerable beneficial effects could already be achieved, even if gene editing only led to a partial reduction of target cell permissiveness.…”

Section: Discussionmentioning

confidence: 99%

How to prevent viremia rebound? Evidence from a PRRSv data-supported model of immune response

Touzeau

Islam

et al. 2019

BMC Syst Biol

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BackgroundUnderstanding what determines the between-host variability in infection dynamics is a key issue to better control the infection spread. In particular, pathogen clearance is desirable over rebounds for the health of the infected individual and its contact group. In this context, the Porcine Respiratory and Reproductive Syndrome virus (PRRSv) is of particular interest. Numerous studies have shown that pigs similarly infected with this highly ubiquitous virus elicit diverse response profiles. Whilst some manage to clear the virus within a few weeks, others experience prolonged infection with a rebound. Despite much speculation, the underlying mechanisms responsible for this undesirable rebound phenomenon remain unclear.ResultsWe aimed at identifying immune mechanisms that can reproduce and explain the rebound patterns observed in PRRSv infection using a mathematical modelling approach of the within-host dynamics. As diverse mechanisms were found to influence PRRSv infection, we established a model that details the major mechanisms and their regulations at the between-cell scale. We developed an ABC-like optimisation method to fit our model to an extensive set of experimental data, consisting of non-rebounder and rebounder viremia profiles. We compared, between both profiles, the estimated parameter values, the resulting immune dynamics and the efficacies of the underlying immune mechanisms. Exploring the influence of these mechanisms, we showed that rebound was promoted by high apoptosis, high cell infection and low cytolysis by Cytotoxic T Lymphocytes, while increasing neutralisation was very efficient to prevent rebounds.ConclusionsOur paper provides an original model of the immune response and an appropriate systematic fitting method, whose interest extends beyond PRRS infection. It gives the first mechanistic explanation for emergence of rebounds during PRRSv infection. Moreover, results suggest that vaccines or genetic selection promoting strong neutralising and cytolytic responses, ideally associated with low apoptotic activity and cell permissiveness, would prevent rebound.Electronic supplementary materialThe online version of this article (10.1186/s12918-018-0666-7) contains supplementary material, which is available to authorized users.

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“…Strategies that use the CRISPR/Cas9 technique to improve the reproduction in swine are becoming more prevalent. PRRSV, a virus associated with reproductive and respiratory disease, can cause severe unsuccessful reproductive outcomes in sows, decrease sperm quality in infected boars, and lower the birth rates of healthy piglets [32]. In 2016, Tao et al generated efficient biallelic mutation in porcine parthenotes by cytoplasmic injection of Cas9/sgRNA mixtures.…”

Section: Applications To Breeding and Reproductionmentioning

confidence: 99%

Application and Development of CRISPR/Cas9 Technology in Pig Research

Lin¹,

Deng²,

Li³

et al. 2019

Gene Editing - Technologies and Applications

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Pigs provide valuable meat sources, disease models, and research materials for humans. However, traditional methods no longer meet the developing needs of pig production. More recently, advanced biotechnologies such as SCNT and genome editing are enabling researchers to manipulate genomic DNA molecules. Such methods have greatly promoted the advancement of pig research. Three gene editing platforms including ZFNs, TALENs, and CRISPR/Cas are becoming increasingly prevalent in life science research, with CRISPR/Cas9 now being the most widely used. CRISPR/Cas9, a part of the defense mechanism against viral infection, was discovered in prokaryotes and has now developed as a powerful and effective genome editing tool that can introduce and enhance modifications to the eukaryotic genomes in a range of animals including insects, amphibians, fish, and mammals in a predictable manner. Given its excellent characteristics that are superior to other tailored endonucleases systems, CRISPR/Cas9 is suitable for conducting pig-related studies. In this review, we briefly discuss the historical perspectives of CRISPR/Cas9 technology and highlight the applications and developments for using CRISPR/Cas9-based methods in pig research. We will also review the choices for delivering genome editing elements and the merits and drawbacks of utilizing the CRISPR/Cas9 technology for pig research, as well as the future prospects.

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Gene editing as applied to prevention of reproductive porcine reproductive and respiratory syndrome

Cited by 31 publications

References 67 publications

CD163ΔSRCR5 MARC-145 Cells Resist PRRSV-2 Infection via Inhibiting Virus Uncoating, Which Requires the Interaction of CD163 With Calpain 1

CD163ΔSRCR5 MARC-145 Cells Resist PRRSV-2 Infection via Inhibiting Virus Uncoating, Which Requires the Interaction of CD163 With Calpain 1

How to prevent viremia rebound? Evidence from a PRRSv data-supported model of immune response

Application and Development of CRISPR/Cas9 Technology in Pig Research

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