Radioactive labeling did much to inform our deep understanding of the transition from host to phage metabolism after T4 infection. With RNAseq, metabolomics and proteomics now facilitating exploring that transition in exciting ways in various phage-host systems, we review radiolabeling approaches that are still uniquely useful and provide a range of complementary data while remaining relatively affordable if equipment is available.
1. Protein synthesis: 2-D protein gels of pulse-labeled T4 infections show that all synthesis of host proteins is shut off within the first minute or so of T4 infection and clearly detail the patterns and quantities of early, middle and late proteins. Gels of amber mutants and large deletion mutants of T4 facilitated identification of specific proteins.
2. T4 Infection of stationary phase E. coli in Hibernation Mode: When T4 infects stationary phase cells, it can pause part way through the infection cycle, making no new phage until nutrients are provided then making hundreds of phage per cell. By pre-labeling the host DNA with 3HdT, we found that it is gradually broken down in the first hour, as happens, though more rapidly, in exponential phase. This can only be observed using a mutant blocked in DNA synthesis; otherwise, the dT is quickly re-incorporated into phage DNA, implying that the phage program is paused only after middle-mode enzymes are made and the Nucleotide Synthetic Complex is functional.
3. T4 effects on host membrane lipid synthesis: In addition to drastic changes in nucleic acid and protein changes, T4 infection also substantially stimulates the synthesis of membrane lipid phosphatidyl glycerol for the first 15 minutes after infection. There is ongoing synthesis but no stimulation of phosphatidyl ethanolamine. Both responses are controlled genetically by T4; they depend on different parts of the deletable region between genes 59 and rIIB.