Artificial riboswitches for gene expression and replication control of DNA and RNA viruses.
Aptazymes are small, ligand-dependent self-cleaving ribozymes that function independently of transcription factors and can be customized for induction by various small molecules. Here, we introduce these artificial riboswitches for regulation of DNA and RNA viruses. We hypothesize that they represent universally applicable tools for studying viral gene functions and for applications as a safety switch for oncolytic and live vaccine viruses. Our study shows that the insertion of artificial aptazymes into the adenoviral immediate early gene E1A enables small-molecule-triggered, dose-dependent inhibition of gene expression. Aptazyme-mediated shutdown of E1A expression translates into inhibition of adenoviral genome replication, infectious particle production, and cytotoxicity/oncolysis. These results provide proof of concept for the aptazyme approach for effective control of biological outcomes in eukaryotic systems, specifically in virus infections. Importantly, we also demonstrate aptazyme-dependent regulation of measles virus fusion protein expression, translating into potent reduction of progeny infectivity and virus spread. This not only establishes functionality of aptazymes in fully cytoplasmic genetic systems, but also implicates general feasibility of this strategy for application in viruses with either DNA or RNA genomes. Our study implies that gene regulation by artificial riboswitches may be an appealing alternative to Tet- and other protein-dependent gene regulation systems, based on their small size, RNA-intrinsic mode of action, and flexibility of the inducing molecule. Future applications range from gene analysis in basic research to medicine, for example as a safety switch for new generations of efficiency-enhanced oncolytic viruses.
Oncolytic Adenovirus Group, German Cancer Research Center (Deutsches Krebsforschungszentrum, DKFZ), 69120 Heidelberg, Germany.
Patrick
P
Ketzer
Johanna K
JK
Kaufmann
Sarah
S
Engelhardt
Sascha
S
Bossow
Christof
C
von Kalle
Jörg S
JS
Hartig
Guy
G
Ungerechts
Dirk M
DM
Nettelbeck
D000256:Adenoviridae / Q000235:genetics / Q000472:pathogenicity / Q000502:physiology; D017500:Adenovirus E1A Proteins / Q000235:genetics / Q000378:metabolism; D002460:Cell Line; D004267:DNA Viruses / Q000235:genetics* / Q000472:pathogenicity / Q000502:physiology*; D015967:Gene Expression Regulation, Viral; D005814:Genes, Viral / Q000235:genetics; D008024:Ligands; D008459:Measles virus / Q000235:genetics / Q000472:pathogenicity / Q000502:physiology; D050504:Oncolytic Viruses / Q000235:genetics / Q000502:physiology; D012328:RNA Viruses / Q000235:genetics* / Q000472:pathogenicity / Q000502:physiology*; D016337:RNA, Catalytic / Q000378:metabolism; D058928:Riboswitch / Q000235:genetics*; D014771:Virion / Q000502:physiology; D053586:Virus Internalization; D014779:Virus Replication / Q000235:genetics*
D017500:Adenovirus E1A Proteins; D008024:Ligands; D016337:RNA, Catalytic; D058928:Riboswitch