Inactivation of porcine endogenous retrovirus in pigs using CRISPR-Cas9.

There is no author summary for this article yet. Authors can add summaries to their articles on ScienceOpen to make them more accessible to a non-specialist audience.

Abstract

Xenotransplantation is a promising strategy to alleviate the shortage of organs for human transplantation. In addition to the concerns about pig-to-human immunological compatibility, the risk of cross-species transmission of porcine endogenous retroviruses (PERVs) has impeded the clinical application of this approach. We previously demonstrated the feasibility of inactivating PERV activity in an immortalized pig cell line. We now confirm that PERVs infect human cells, and we observe the horizontal transfer of PERVs among human cells. Using CRISPR-Cas9, we inactivated all of the PERVs in a porcine primary cell line and generated PERV-inactivated pigs via somatic cell nuclear transfer. Our study highlights the value of PERV inactivation to prevent cross-species viral transmission and demonstrates the successful production of PERV-inactivated animals to address the safety concern in clinical xenotransplantation.

Related collections

Most cited references 11

Record: found
Abstract: found
Article: not found

Genome editing assessment using CRISPR Genome Analyzer (CRISPR-GA).

Marc Guell, Luhan Yang, George Church (2014)

Clustered regularly interspaced short palindromic repeats (CRISPR)-based technologies have revolutionized human genome engineering and opened countless possibilities to basic science, synthetic biology and gene therapy. Albeit the enormous potential of these tools, their performance is far from perfect. It is essential to perform a posterior careful analysis of the gene editing experiment. However, there are no computational tools for genome editing assessment yet, and current experimental tools lack sensitivity and flexibility. We present a platform to assess the quality of a genome editing experiment only with three mouse clicks. The method evaluates next-generation data to quantify and characterize insertions, deletions and homologous recombination. CRISPR Genome Analyzer provides a report for the locus selected, which includes a quantification of the edited site and the analysis of the different alterations detected. The platform maps the reads, estimates and locates insertions and deletions, computes the allele replacement efficiency and provides a report integrating all the information. CRISPR-GA Web is available at http://crispr-ga.net. Documentation on CRISPR-GA instructions can be found at http://crispr-ga.net/documentation.html mguell@genetics.med.harvard.edu. © The Author 2014. Published by Oxford University Press. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.

0 comments Cited 60 times – based on 0 reviews      Review now

Bookmark

Record: found
Abstract: found
Article: not found

Multiple groups of novel retroviral genomes in pigs and related species.

Y Takeuchi, W. Heneine, T Switzer … (2001)

In view of the concern over potential infection hazards in the use of porcine tissues and organs for xenotransplantation to humans, we investigated the diversity of porcine endogenous retrovirus (PERV) genomes in the DNA of domestic pigs and related species. In addition to the three known envelope subgroups of infectious gamma retroviruses (PERV-A, -B, and -C), classed together here as PERV group gamma 1, four novel groups of gamma retrovirus (gamma 2 to gamma 5) and four novel groups of beta retrovirus (beta 1 to beta 4) genomes were detected in pig DNA using generic and specific PCR primers. PCR quantification indicated that the retroviral genome copy number in the Landrace x Duroc F(1) hybrid pig ranged from 2 (beta 2 and gamma 5) to approximately 50 (gamma 1). The gamma 1, gamma 2, and beta 4 genomes were transcribed into RNA in adult kidney tissue. Apart from gamma 1, the retroviral genomes are not known to be infectious, and sequencing of a small number of amplified genome fragments revealed stop codons in putative open reading frames in several cases. Analysis of DNA from wild boar and other species of Old World pigs (Suidae) and New World peccaries (Tayassuidae) showed that one retrovirus group, beta 2, was common to all species tested, while the others were present among all Old World species but absent from New World species. The PERV-C subgroup of gamma1 genomes segregated among domestic pigs and were absent from two African species (red river hog and warthog). Thus domestic swine and their phylogenetic relatives harbor multiple groups of hitherto undescribed PERV genomes.

0 comments Cited 36 times – based on 0 reviews      Review now

Bookmark

Record: found
Abstract: found
Article: found

Is Open Access

How Active Are Porcine Endogenous Retroviruses (PERVs)?

Joachim Denner (2016)

Porcine endogenous retroviruses (PERVs) represent a risk factor if porcine cells, tissues, or organs were to be transplanted into human recipients to alleviate the shortage of human transplants; a procedure called xenotransplantation. In contrast to human endogenous retroviruses (HERVs), which are mostly defective and not replication-competent, PERVs are released from normal pig cells and are infectious. PERV-A and PERV-B are polytropic viruses infecting cells of several species, among them humans; whereas PERV-C is an ecotropic virus infecting only pig cells. Virus infection was shown in co-culture experiments, but also in vivo, in the pig, leading to de novo integration of proviruses in certain organs. This was shown by measurement of the copy number per cell, finding different numbers in different organs. In addition, recombinations between PERV-A and PERV-C were observed and the recombinant PERV-A/C were found to be integrated in cells of different organs, but not in the germ line of the animals. Here, the evidence for such in vivo activities of PERVs, including expression as mRNA, protein and virus particles, de novo infection and recombination, will be summarised. These activities make screening of pigs for provirus number and PERV expression level difficult, especially when only blood or ear biopsies are available for analysis. Highly sensitive methods to measure the copy number and the expression level will be required when selecting pigs with low copy number and low expression of PERV as well as when inactivating PERVs using the clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated nuclease (CRISPR/Cas) technology.

0 comments Cited 34 times – based on 0 reviews      Review now

Bookmark

All references

Author and article information

Journal

PubMed ID:: 28798043

PMC ID:: 5813284

DOI:: 10.1126/science.aan4187

Data availability:

Comments

Comment on this article

scite_

Cited by 251

See all cited by

Inactivation of porcine endogenous retrovirus in pigs using CRISPR-Cas9.

Read this article at

Abstract

Related collections

Genome Engineering using CRISPR

Most cited references 11

Genome editing assessment using CRISPR Genome Analyzer (CRISPR-GA).

Multiple groups of novel retroviral genomes in pigs and related species.

How Active Are Porcine Endogenous Retroviruses (PERVs)?

Author and article information

Journal

Comments

Comment on this article

Similar content 320

Cited by 251