Project:  Imaging of viral mRNA using sequence specific fluorescent Peptide Nucleic Acids 

Susann Kummer 

Viral infections are spread all over the world and cause a variety of diseases. Understanding the molecular mechanisms of viral replication inside the host cell provides the basis for the development of new therapeutic treatments.
This Project is focused on the genesis of enveloped viruses. An early consequence of virus infection is the formation of viral mRNA. To follow both its formation and intracellular location we utilize PNA (Peptide Nucleic Acid) molecules, tagged with a fluorescent moiety. Upon complementation of the PNA with the respective mRNA sequence, intercalation of the fluorophore is accompanied by strong enhancement of quantum efficiency, which can be detected by (Confocal Laser Scanning) Fluorescence Microscopy in living infected cells. The advantage of these RNA probes is the stability of the peptide backbone. We studied the infection of BHK-21 cells by Vesicular Stomatitis Virus (VSV) as a model system for this technique (Fig. 2). VSV is a negative-sense single-stranded RNA virus that causes a zootic vesicular disease. This virus offers several advantages: (i) the existence of highly conserved sequences in the N- and L- gene as targets for PNAs, (ii) a cytosolic mechanism of transcription and replication and (iii) the convenience of simple cultivation and production of infective particles with high titres in different cell lines. Employing time-lapse imaging we could follow the production of VSV mRNA in infected cells.

	Fig.2: Detection of VSV mRNA in infected BHK-21 cells using PNA

References: