Dataset Information

Sequential disruption of ALV host receptor genes reveals no sharing of receptors between ALV subgroups A, B, and J.

ABSTRACT:

Background

Previously, we showed that targeted disruption of viral receptor genes in avian leukosis virus (ALV) subgroups using clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated protein 9 (Cas9))-based genome editing confers resistance to ALV subgroups B and J. Here, we used the same strategy to target the receptor expressed by ALV subgroup A (TVA) and generate chicken cells resistant to infection by this virus.

Results

CRISPR/Cas9-based disruption of exon 2 within the tva gene of DF-1 fibroblasts conferred resistance to infection by ALV subgroup A regardless of whether frameshift mutations were introduced during editing. Conversely, overexpression of the wild-type TVA receptor (wtTVA) by tva-modified DF-1 clones restored susceptibility to ALV subgroup A. The results confirm that exon 2, which contains the low-density lipoprotein receptor class A domain of TVA, is critical for virus entry. Furthermore, we sequentially modified DF-1 cells by editing the tva, tvb, and Na⁺/H⁺ exchange 1 (chNHE1) genes, which are the specific receptors for ALV subgroups A, B, and J, respectively.

Conclusions

Simultaneous editing of multiple receptors to block infection by different subgroups of ALV confirmed that ALV subgroups A, B, and J do not share host receptors. This strategy could be used to generate cells resistant to multiple viral pathogens that use distinct receptors for cell entry.

SUBMITTER: Lee HJ

PROVIDER: S-EPMC6444617 | biostudies-literature | 2019

REPOSITORIES: biostudies-literature

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Publications

Sequential disruption of ALV host receptor genes reveals no sharing of receptors between ALV subgroups A, B, and J.

Lee Hong Jo HJ Park Kyung Je KJ Lee Kyung Youn KY Yao Yongxiu Y Nair Venugopal V Han Jae Yong JY

Journal of animal science and biotechnology 20190402

<h4>Background</h4>Previously, we showed that targeted disruption of viral receptor genes in avian leukosis virus (ALV) subgroups using clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated protein 9 (Cas9))-based genome editing confers resistance to ALV subgroups B and J. Here, we used the same strategy to target the receptor expressed by ALV subgroup A (TVA) and generate chicken cells resistant to infection by this virus.<h4>Results</h4>CRISPR/Cas9-based disrupti ...[more]

PMID: 30976416

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Project description:Background and purposeThe calcitonin (CT) receptor family is complex, comprising two receptors (the CT receptor [CTR] and the CTR-like receptor [CLR]), three accessory proteins (RAMPs) and multiple endogenous peptides. This family contains several important drug targets, including CGRP, which is targeted by migraine therapeutics. The pharmacology of this receptor family is poorly characterised in species other than rats and humans. To facilitate understanding of translational and preclinical data, we need to know the receptor pharmacology of this family in mice.Experimental approachPlasmids encoding mouse CLR/CTR and RAMPs were transiently transfected into Cos-7 cells. cAMP production was measured in response to agonists in the absence or presence of antagonists.Key resultsWe report the first synthesis and characterisation of mouse adrenomedullin, adrenomedullin 2 and βCGRP and of mouse CTR without or with mouse RAMPs. Receptors containing m-CTR had subtly different pharmacology than human receptors; they were promiscuous in their pharmacology, both with and without RAMPs. Several peptides, including mouse αCGRP and mouse adrenomedullin 2, were potent agonists of the m-CTR:m-RAMP3 complex. Pharmacological profiles of receptors comprising m-CLR:m-RAMPs were generally similar to those of their human counterparts, albeit with reduced specificity.Conclusion and implicationsMouse receptor pharmacology differed from that in humans, with mouse receptors displaying reduced discrimination between ligands. This creates challenges for interpreting which receptor may underlie an effect in preclinical models and thus translation of findings from mice to humans. It also highlights the need for new ligands to differentiate between these complexes.Linked articlesThis article is part of a themed issue on Advances in Migraine and Headache Therapy (BJP 75th Anniversary).. To view the other articles in this section visit http://onlinelibrary.wiley.com/doi/10.1111/bph.v179.3/issuetoc.

Dataset Information

Sequential disruption of ALV host receptor genes reveals no sharing of receptors between ALV subgroups A, B, and J.

Background

Results

Conclusions

Publications

Sequential disruption of ALV host receptor genes reveals no sharing of receptors between ALV subgroups A, B, and J.

Similar Datasets

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