Results: 2

1.

Figure. From: Physiological role of stalk lengthening in Caulobacter crescentus.

Figure 1. StpABCD complexes are not required for cell survival upon stalk detachment. (A) Stalk elongation upon phosphate starvation. Caulobacter cells were grown overnight in defined HIGG media 15 containing either 1 mM or 1 µM phosphate. Scale bar: 2 µm. (B) Schematic of a Caulobacter cell showing the physiological separation of the cell body from the stalk by protein diffusion barriers. (C) Mechanical removal of stalks does not affect viability. Stalks of wild-type (WT, CB15N) 16 and diffusion barrier-deficient (ΔstpAB) cells, 5 grown in low-phosphate medium (M2G−P) 5 for 24 h, were sheared off in a pre-cooled Waring blender for 3 min at maximum speed at 4°C. Successful removal of stalks was confirmed by DIC microscopy (data not shown). Suspensions of the resulting stalk-less cells were adjusted to equal optical densities, serially diluted, spotted onto PYE agar and incubated at 28°C.

Eric A Klein, et al. Commun Integr Biol. 2013 July 1;6(4):e24561.
2.

Figure. From: Physiological role of stalk lengthening in Caulobacter crescentus.

Figure 2. Models for the physiological role of stalk elongation. (A) The red pathway describes the previously held model of phosphate uptake, in which PstS shuttles phosphate from the stalk to the cell body, where it is imported by the PstCAB transporter. 6 Our discovery of a diffusion barrier which blocks PstS shuttling contradicts this model. 5 An alternative model, shown as the green pathway, assumes that previously undetected PstCAB complexes do exist in the stalk, allowing the uptake of phosphate into the cytoplasmic core of the stalk and its subsequent diffusion to the cell body. (B) Using kinetic and diffusion parameters, the distance that PstS-phosphate complexes can diffuse before phosphate release is approximately 0.7 µm. This distance is far shorter than the stalk length under phosphate-limiting conditions. (C) Localization of PstA-GFP in the stalk. Strain YB4062 (CB15N pMR10-Ppst-pstCA-gfp) was grown for 36 h in HIGG medium containing 30 µM phosphate. PstA-GFP was visualized by fluorescence microscopy. Arrowheads indicate stalks containing the fusion protein. Scale bar: 2 µm. (D) Subcellular localization of Heat Shock Protein 20 (HSP20, IbpA homolog) in wild-type and ΔstpAB cells grown in high-phosphate (PYE) and in low-phosphate (M2G−P) medium. To test for the segregation of damaged proteins into the stalk, cells of strains SS419 (CB15N Pxyl::Pxyl-ibpA-venus) and SS420 (CB15N ΔstpAB Pxyl::Pxyl-ibpA-venus) were first grown in M2G−P for 12 h. Production of IbpA-Venus was induced by adding 0.3% xylose for 1 h prior to a heat shock. Cells were shifted to 40°C for 1 h, followed by a growth period of 8 h at 28°C. Untreated cells were cultured at 28°C for 9 h. Images show overlays of DIC and false-colored fluorescence images. Scale bar: 3 µm. (E) Stalk elongation may function to elevate single cells away from surfaces. As the cell distances itself from the surface, fluid velocity (blue gradient) and nutrient flux (blue arrows) increase. Thus, stalk elongation may ensure greater nutrient availability. (F) Caulobacter cells may co-colonize surfaces with other organisms. By distancing themselves from the surface, they may have greater access to nutrients relative to nearby surface-associated species, thereby increasing their competitiveness.

Eric A Klein, et al. Commun Integr Biol. 2013 July 1;6(4):e24561.

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