(A) Translation was arrested (by addition of 100 µg/ml chloramphenicol) in sporulating wild-type PY79 cells (strain RL3), and samples were withdrawn at the indicated times. SpoIIE was detected by western blot using affinity purified rat α-SpoIIE polyclonal antibody. (B) Degradation of SpoIIE was monitored in vegetative cells induced to produce untagged SpoIIE (strain RL6040, wt) (strain RL6041, ∆ftsH) as in A. (C) Domains of SpoIIE-FLAG were serially deleted or replaced as diagramed at left and expressed in wt or ∆ftsH vegetatively growing cells. Translation was arrested by chloramphenicol addition, and samples were collected at times indicated. SpoIIE was detected by western blotting with α-FLAG monoclonal antibody. ∆Tag removes residues 11–37 (strain RL5880, wt) (strain RL5884, ∆ftsH), MalF-tm replaces the transmembrane domain (residues 38–319) with the first two transmembrane domains of E. coli MalF (strain RL5881, wt) (strain RL5885, ∆ftsH), ∆Reg removes the regulatory domain (residues 320–568) (strain RL5882, wt) (strain RL5886, ∆ftsH), and ∆PP2C removes the phosphatase domain (residues 590–827) (strain RL5883, wt) (strain RL5887, ∆ftsH). Western blots for strains shown in Figure 2E are duplicated here for comparison.