Work

The Nonmonotonic Dose Dependence of Protein Expression in Cells Transfected with Self-Amplifying RNA

Year: 2022

Type: article

Abstract: Positive-sense RNA viruses often have a cytotoxic effect on their host cell because of the strength of their RNA replicase proteins, even though only one copy of their genome begins the viral life cycle in each cell. Noninfectious forms of them-replicons-which include just their RNA replication-related genes, are also strongly self-amplifying and cytotoxic. Accordingly, when replicons fused with nonviral genes of interest are transfected into cells to amplify expression of proteins of interest, one needs to keep the replicon "dose" sufficiently low. We demonstrate how to control the number of RNA replicons getting into transfected cells and that there is a sweet spot for the replicon dose that optimizes protein expression and cell viability. Examples are given for the case of Nodamura viral replicons with fluorescent protein reporter genes in a variety of mammalian cell lines, quantified by flow cytometry and live/dead cell assays.

Source: Journal of Virology

DOI: 10.1128/jvi.01858-21

Authors: Cheylene R. Tanimoto, Abby R. Thurm, Devin S. Brandt, Charles M. Knobler, William M. Gelbart

Institutions: University of California, Los Angeles, California NanoSystems Institute

Language: English

Topic: RNA Interference and Gene Delivery

Subfield: Molecular Biology

Field: Biochemistry, Genetics and Molecular Biology

Domain: Life Sciences

SDG: Good health and well-being

Open Access status: green

Funder: National Science Foundation