Figure 2.
UCC1 Defines a New Central Sub-domain in the Casparian Strip
(A) Diagram representing the construct pUCC1::mCherry-UCC1 (UTR, untranslated region; SP, signal peptide; H Cter, hydrophobic C terminus for GPI anchoring).
(B) Maximum intensity projection, orthogonal, median, and surface views of confocal sections of plants expressing pUCC1::mCherry-UCC1 (red) in cleared roots. In the case of maximum intensity projection (maximum projection), the figure represents different regions of the root measured as number of cells after the onset of elongation. For the orthogonal, median, and surface views, cell walls were stained with Calcofluor white (gray in the figures). Scale bar, 20 μm for the maximum projection and orthogonal views. Scale bar, 5 μm for the median and surface views. Ep, epidermis; Co, cortex; End, endodermis (see also Figure S2A).
(C) Maximum intensity projection, median, and surface view of confocal sections of plants expressing CASP1-GFP (cyan) and mCherry-UCC1 (red). Signal was captured at the 10th endodermal cell after the onset of elongation observed in vivo. Scale bar, 20 μm for maximum projection and 3 μm for median and surface view (see also Figure S2B).
(D) In vivo observation of the surface view of an endodermal cell expressing pESB1::ESB1-mCherry or pCASP1::CASP1-GFP. Scale bar, 2 μm.
(E) Immunolocalization assay of UCC1 protein (red) in plant expressing pCASP1::CASP1-GFP (cyan). A primary polyclonal antibody targeting UCC1 was used in combination with a secondary antibody conjugated with Dylight 633. Scale bar, 2 μm see also Figures S2C–S2H.
(F) Graph presenting the distribution of normalized pixels intensity (relative pixel intensity, 0–1) across the Casparian strip (distance in μm) for CASP1-GFP fluorescence (cyan) and UCC1 immunofluorescence (red, Dylight 633). Light curves represent individual replicates coming from individual plants (n = 4). Each replicate is the average pixel intensity across a segment of 25 μm along the Casparian strip axis. Dark curves represent the mean values for CASP1-GFP and UCC1 immunofluorescence.