Madaline McPherson, Indiana University

Madaline McPherson, Qin Liao, Xindan Wang, Pranav Danthi

Cell Biology of Viral Infection

Bacteriophage phi6 contains an envelope, a segmented dsRNA genome, and unique replication cycle that requires this phage to display biology that is distinct from most other phages. Although aspects of phi6 replication are understood, the host factors that regulate infection by this phage are unknown. We used transposon mutagenesis coupled with deep sequencing (Tn-seq) of P. syringae pv. phaseolicola HB10Y to perform a genome-wide screen of host genes that regulate phi6 infection. This screen identified 74 candidates that render HB10Y resistant to infection. We investigated the role of mdoG and mdoH in phi6 replication. The mdoGH genes are involved in the biosynthesis of osmoregulated periplasmic glucans (OPGs). OPGs are synthesized in the periplasm and serve as osmoprotectants in some bacteria. Analysis of mdoG- and mdoH-deficient mutants revealed that phi6 produces small plaques in these strains. Infectious center assays revealed that mdoG and mdoH mutants and wild-type HB10Y have the same proportion of infected cells. The small plaque size was not due to a smaller proportion of infected cells. Our experiments measuring progeny phage release revealed that mdoG and mdoH mutants supported replication and release of phi6 to the same efficiency as in wild-type cells. Surprisingly, we found that unlike wild-type cells, mdoG and mdoH mutants don't succumb to infection. These experiments reveal a role for mdoG and mdoH in controlled phi6-induced cell death. Our results uncouple phage replication from cell death and reveal that phi6 can be released without lysis from a bacterial host with two membranes.