MetSVORF12's potential to promote viral replication of MetSV in Methanosarcina mazei

Archaeal viruses are still the most enigmatic and unstudied part of microbiology. In recent years, novel archaeal viruses were isolated and characterized demonstrating a high diversity not only in morphology but also in genome structure. One of these was the Methanosarcina spherical virus (MetSV), a lytic virus of the Tectiviridae specifically infecting the methanoarchaeal model organism Methanosarcina mazei. MetSV's linear dsDNA genome has a size of ~ 10.5 kbp encoding 23 ORFs, of which 12 are classified as small ORFs^1,2 (sORFs). For a better understanding of the virus-host interaction, we searched for anti-defense proteins thus focusing on the sORFs and looked for possible interactions of the encoded proteins with the hosts proteome or DNA molecules revealing MetSVORF12s ability to bind DNA. Overexpression of MetSVORF12 in the host M. mazei showed a strong phenotype pointing towards an interference with its replication. Recently, we have established a genetic system for MetSV now allowing genetic approaches for functional characterizations². Consequently, we created a deletion of ORF12 by side-directed mutagenesis in the plasmid-derived virus. Reinfection with ΔMetSVORF12 through liposome-mediated transformation into M. mazei resulted in culture lysis indicating a nonessential function but still reducing the virus efficiency compared to the wildtype. Currently, we are studying more potential interaction partners. Based on our results we propose a role of MetSVORF12 in the MetSV replication.

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