Fig 4.

Zn²⁺ induces transcription of eptA_Pa. A) Cells were grown in LB broth. Major ³²P-labeled lipid A species are indicated with a cartoon corresponding to the lipid A structure; colors of modification groups are the same as those used in Fig. 1. Both heterologous expression of eptA_Pa as well as addition of 2mM ZnSO₄ to the media results in pEtN addition to lipid A. This modification is not detectable in the eptA_Pa mutant, but restored upon complementation with peptA_nprom. B) Relative gene expression of eptA_Pa and arnT in response to Zn²⁺. Transcription of eptA_Pa is induced by 2mM ZnSO₄ approximately 21-fold. Zn²⁺ downregulates arnT transcription by >4-fold. Ratios were standardized relative to expression of the housekeeping control gene, clpX. C), D) and E). MALDI-TOF MS analysis of lipid A prepared from cells grown in LB broth. C) Analysis of PA14 + 2mM ZnSO₄ reveals pEtN addition to lipid A, while D) ΔeptA_Pa + 2mM ZnSO₄ shows no pEtN modification, but instead L-Ara4N addition. E) Complementation of ΔeptA_Pa with peptA_nprom restores pEtN addition to the lipid A in response to Zn²⁺. The fractions most representative of pEtN modification are shown.