A prophage-encoded actin-like protein required for efficient viral DNA replication in bacteria

Catriona Donovan,Antonia Heyer,Anja Wittmann,Julia Frunzke,Tino Polen,Reinhard Krämer,Eugen Pfeifer,Marc Bramkamp

doi:10.1093/nar/gkv374

Abstract

In host cells, viral replication is localized at specific subcellular sites. Viruses that infect eukaryotic and prokaryotic cells often use host-derived cytoskeletal structures, such as the actin skeleton, for intracellular positioning. Here, we describe that a prophage, CGP3, integrated into the genome of Corynebacterium glutamicum encodes an actin-like protein, AlpC. Biochemical characterization confirms that AlpC is a bona fide actin-like protein and cell biological analysis shows that AlpC forms filamentous structures upon prophage induction. The co-transcribed adaptor protein, AlpA, binds to a consensus sequence in the upstream promoter region of the alpAC operon and also interacts with AlpC, thus connecting circular phage DNA to the actin-like filaments. Transcriptome analysis revealed that alpA and alpC are among the early induced genes upon excision of the CGP3 prophage. Furthermore, qPCR analysis of mutant strains revealed that both AlpA and AlpC are required for efficient phage replication. Altogether, these data emphasize that AlpAC are crucial for the spatio-temporal organization of efficient viral replication. This is remarkably similar to actin-assisted membrane localization of eukaryotic viruses that use the actin cytoskeleton to concentrate virus particles at the egress sites and provides a link of evolutionary conserved interactions between intracellular virus transport and actin.

Highlights

DNA of viral origin, including fully functional prophages, cryptic prophage elements or phage morons, represents a common element of bacterial genomes [1,2].Upon prophage induction, e.g. triggered by the host SOS response, temperate phages enter the lytic pathway leading to the excision of the viral DNA from the genome, replication, virion assembly and lysis of the host cell [3,4]
We identified and characterized a novel actin-like protein encoded by the cryptic prophage CGP3 located in the genome of Corynebacterium glutamicum strain ATCC 13032
Our labs have shown that the C. glutamicum ATCC 13032 prophage CGP3 is able to excise from the chromosome and replicate autonomously [44]

Summary

Introduction

DNA of viral origin, including fully functional prophages, cryptic (degenerated) prophage elements or phage morons, represents a common element of bacterial genomes [1,2].Upon prophage induction, e.g. triggered by the host SOS response, temperate phages enter the lytic pathway leading to the excision of the viral DNA from the genome, replication, virion assembly and lysis of the host cell [3,4]. The genetic organization of par loci is similar for both chromosome- and plasmid-encoded systems. The segrosomes are recognized and segregated by the action of a partition protein, which, depending on the plasmid partitioning system, is either a Walker-A P-loop ATPase (ParA, Type I), an actin-like ATPase (ParM, Type II) or a tubulin-like GTPase (TubZ, Type III) [14,15,20,21,22]. Walker-A P-loop ATPase, on the other hand, have been proposed to employ a diffusionratchet model In this model, the nucleotide bound from of the ATPase determines its affinity for the nucleoid. Combined with associated regulatory elements and intrinsic ATPase activity, gradients of the Walker-A ATPases form on the nucleoid.

Methods

Results

Conclusion