Figure 6.
Increased holdfast deacetylation increases holdfast binding in high-ionic-strength environments
(A) Schematic of the experimental setup.
(B–D) Cells were grown exponentially for 2 h in PYE with 0.03% xylose (A-B), or 0 μM, 50 μM, or 250 μM CuSO4 (C) and shed holdfast were collected from the culture supernatant. The purified holdfasts were mixed with different concentration of NaCl and incubated on glass slides for 4 h. The percentage of holdfasts bound per field of view was quantified at different concentrations of NaCl. The number of holdfasts bound per field of view at 0 mM NaCl was standardized to 100%. Data are expressed as an average of five independent biological replicates with five technical replicates each. Error bars represent the standard error of the mean. (B) Purified holdfasts from C. crescentus ΔhfaB with pMR10 (empty vector, blue dashed line) as a control, C. crescentus ΔhfaB ΔhfsH complemented with HfsH from C. crescentus under the control of the xylose-inducible promoter (PxylCC-hfsHCC, green), C. crescentus ΔhfaB ΔhfsH cross-complemented with HfsH from H. baltica under the control of the xylose-inducible promoter (PxylCC-hfsHHB, red), and H. baltica ΔhfaB with pMR10 (empty vector, yellow). (C) Purified holdfasts from H. baltica ΔhfaB with pMR10 (empty vector, black dashed line) as a control, H. baltica ΔhfaB ΔhfsH complemented with HfsHHB under the control of the xylose-inducible promoter (PxylHB-hfsHHB, maroon), and H. baltica ΔhfaB ΔhfsH cross-complemented with HfsHCC under the control of the xylose-inducible promoter (PxylHB-hfsHCC, blue). (D) Purified holdfasts from H. baltica ΔhfaB ΔhfsH complemented with HfsHHB under the control of the copper-inducible promoter (PCu-hfsHHB) and H. baltica ΔhfaB ΔhfsH cross-complemented with HfsHCC under the control of the copper-inducible promoter (PCu-hfsHCC). PCu-hfsHCC was induced with 0 μM (black), 50 μM (blue), and 250 μM CuSO4 (green), and PCu-hfsHHB was induced with 0 μM (maroon), 50 μM (purple), and 250 μM CuSO4 (red).