Fig 2: Western blot analysis of PDDI2-GFP gene products. (A) Cells harboring YCp- and YEp-based plasmids in response to 20 mM CY treatment. (B) Dose response of YCpU-PDDI2-GFP-transformed cells in response to CY. (C) Response of YEpU-PDDI2-GFP-transformed cells in response to high CY doses. (D) Response of YEpU-PDDI2-GFP-transformed cells in response to low CY doses. (E) Cells harboring YCp- and YEp-based plasmids in response to 0.015% MMS treatment. (F) Dose response of YCpU-PDDI2-GFP-transformed cells in response to MMS. (G) Response of YEpU-PDDI2-GFP-transformed cells in response to high MMS doses. (H) Response of YEpU-PDDI2-GFP-transformed cells in response to low MMS doses. The experimental protocol is described in Materials and Methods and all treatments were for 2 h. The anti-GFP monoclonal antibody B-2 was purchased from Santa Cruz (sc-9996) and the yeast anti-Pgk1 polyclonal antibody was a generous gift from Dr. W. Li (Institute of Zoology, Chinese Academy of Sciences, Beijing).

Fig 3: Mutations in MinE2–9 affected lipid binding in vitro and protein stability in vivo.(a) Full-length MinE carrying a single residue substitution (F6D) to weaken the amphipathicity of MinE2–9 showed reduced co-sedimentation with liposomes. S, supernatant; P, pellet. (b) Statistical analysis of the ability of MinEF6D to co-sediment with liposomes. Three experiments were used to quantify the supernatant and pellet fractions for analysis. (c) Cellular localization of Yfp-MinD, MinEA2E/L3S/F6D-Cfp, and wild-type MinE-Cfp. Western blot analyses showed that the mutant protein MinEA2E/L3S/F6D was unstable in cells (right panel) and influenced MinDE localization (left panel). Membrane association through the correct folding of MinE2–9 may be critical for MinE protein stability in cells. Monoclonal anti-GFP antibody (Santa Cruz Biotechnology, sc-9996) was used for detecting the fusion proteins.

Fig 4: Subcellular distribution of selected OsUev1s. a Subcellular localization of GFP-OsUev1s and their derivatives as illustrated in the right panel. The GFP-tagged OsUev1s were expressed in N. benthamiana leaves by an Agrobacterium-mediated infiltration method. Photos were taken after 2–3 days of infiltration. A representative image is shown for each transformant. b The above transformed N. benthamiana leaves were analyzed by a protein-partitioning assay as described in Methods. GFP-OsUev1A and GFP-OsUev1A-?CT were detected by an anti-GFP antibody (B-2, sc-9996, Santa Cruz). A: aqueous phase; D: detergent phase