Fig. 4. SYMREM1 stabilizes membrane tubulation and curvature in a cell wall-independent manner.

a–c N. benthamiana leaf protoplasts ectopically expressing YFP-SYMREM1 develop multiple membrane tubes as shown by confocal laser-scanning microscopy (a) and scanning electron microcopy (b) and close-up in (c). d–i Protoplasts expressing different SYMREM1 variants: only IDR domain (SYMREM1^IDR, d), only C-terminal including RemCA (SYMREM1^Cterm, e), full-length sequence lacking the RemCA domain (SYMREM1^FLΔRemCA, f), RemCA domain only (SYMREM1^RemCA, g, h), and the RemCA domain bearing a mutation of the palmitoylated Cys197 residue (SYMREM1^{RemCA C197A}, i). Scale bars indicate 10 μm. All the confocal images are shown as maximal projections. Magenta signals derive from chlorophyll autofluorescence within chloroplasts. Numbers indicate frequencies of all observations. Protoplast isolation experiments were at least performed in four biological replicates, with similar tube formation frequencies found in each replicate.