Isolation of Suppressor Genes that Restore Retrovirus Susceptibility to a Virus-resistant Cell Line
Genetic selections in mammalian cell lines have recently been developed for the isolation of mutant cells that are refractory to infection by retroviruses. These selections have been used to recover lines that block early postentry stages of infection, either before reverse transcription or before nuclear entry. The mechanisms of action of these blocks remain unknown.
We have devised a method for the selection of genes from cDNA libraries that suppress the block to virus infection, and so restore virus susceptibility. The protocol involves the transformation of pools of resistant cells by cDNA expression libraries, followed by the selection for rare virus-sensitive cells, using multiple rounds of selection after infection by marked viral vector genomes. The suppressor genes were then recovered from these virus sensitive cells, and their ability to restore virus susceptibility was confirmed by reintroduction of these cDNAs into the resistant line.
The identities of these genes provide insights into the mechanism of virus resistance and will help to define new pathways used during retrovirus infection. The methods for gene isolation developed here will also permit the identification of similar suppressors that modify or override other recently identified virus resistance genes.
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- Academic Units
- Biochemistry and Molecular Biophysics
- Pathology and Cell Biology
- BioMed Central
- Published Here
- August 24, 2014