LpoB activation of PBP1B is required for spheroplast recovery. Spheroplasts lacking PBP1B (ΔmrcB) (A to E) or LpoB (ΔlpoB) (F to H) were grown on osmotically protected sucrose recovery medium, and the recovery process was monitored by time-lapse phase-contrast microscopy. The time after plating (in hours:minutes) is displayed in the upper right corner of each panel. The images are representative. (A) E. coli DR7V1(pAM238), vector-only control (n = 7); (B) E. coli DR7A(pSK17), wild-type PBP1A overexpressed from a low-copy-number vector (↓), 100 μM IPTG (n = 7); (C) E. coli MAJ595(pMAJ60), wild-type PBP1A overexpressed from a medium-copy-number vector (↑), 25 μM IPTG (n = 45); (D) E. coli DR7C(pSK12), PBP1B complementation with basal expression, no IPTG (n = 9); (E) E. coli DR7U(pUB2H), PBP1B-ΔUB2H overexpressed (n = 8); (F) E. coli DR8V(pDEV), vector-only control (n = 14); (G) E. coli DR8C(pLpoB), wild-type LpoB overexpressed (n = 5); (H) E. coli DR8CIM(pLpoB-IM), inner membrane-bound LpoB variant overexpressed (n = 15).