Transcriptional Organization of the Salmonella Typhimurium Phage P22 pid ORFan Locus

Abstract

Many phage genes lack sequence similarity to any other open reading frame (ORF) in current databases. These enigmatic ORFan genes can have a tremendous impact on phage propagation and host interactions but often remain experimentally unexplored. We previously revealed a novel interaction between phage P22 and its Salmonella Typhimurium host, instigated by the ORFan gene pid (for phage P22 encoded instigator of dgo expression) and resulting in derepression of the host dgoRKAT operon. The pid gene is highly expressed in phage carrier cells that harbor a polarly located P22 episome that segregates asymmetrically among daughter cells. Here, we discovered that the pid locus is fitted with a weak promoter, has an exceptionally long 5′ untranslated region that is instructive for a secondary pid mRNA species, and has a 3′ Rho-independent termination loop that is responsible for stability of the pid transcript.

Superinfection exclusion factors drive a history-dependent switch from vertical to horizontal phage transmission

“…A similar experiment was set up as in Figure 2A, but this time LT2 and P22 were replaced with LT2K7 and P22-pid CÀ183A so that established P22 lysogens become easily detectable as Lac + (i.e., constitutively blue) colonies after plating on LB X-Gal. More specifically, P22-pid CÀ183A constitutively overexpresses its Pid protein, thereby constitutively activating the dgo::lacZ reporter in LT2K7 (Wolput et al, 2022). As such, the trend of P22 lysogens versus non-lysogens under sustained high phage-to-host exposure (i.e., P22-pid CÀ183A phages were added at t = 0 h and once more at t = 2 h; Figure 2B, triangles) could readily be monitored.…”

“…In this assay, non-lysogenic colonies produce blue colored streaks after overnight incubation at 37 C because of ongoing lysis (Maloy et al, 1996;Davis et al, 1980). In experiments where LT2K7 was used as host, colonies of P22-pid CÀ183A lysogens could easily be discriminated from those of P22 lysogens or non-lysogens because of their blue color on LB X-Gal (Wolput et al, 2022). Similarly plaques of P22-pid CÀ183A on LB X-Gal soft-agar lawns of LT2K7 can be readily identified by their blue color (Wolput et al, 2022).…”