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. 2017 Jul 6;29(7):1748–1772. doi: 10.1105/tpc.17.00044

Figure 1.

Figure 1.

MfNACsa Is Associated with Membranes.

(A) Confocal imaging of transgenic M. truncatula hairy roots expressing MfNACsa-GFP fusion proteins driven by the constitutive CaMV 35S promoter (i) or the MfNACsa endogenous promoter (ii). GFP signal was detected with an Olympus FluoView FV1000 confocal laser scanning microscope. Bars = 50 μm.

(B) Cellular fractionation and immunoblot analysis of ProMfNACsa:MfNACsa-GFP protein in N. benthamiana leaves. Soluble and membrane proteins (i), and cytoplasmic and nuclear proteins were separated and subjected the immunoblot analysis (ii). H+-ATPase, Histone H3, and cFBPase were used as PM, nuclear, and cytosolic fraction markers, respectively.

(C) MfNACsa localizes to the PM and ER compartment in onion epidermal cells. Colocalization of MfNACsa-eGFP fusion proteins driven by CaMV35S promoter with the RFP-tagged PIP2A PM marker (i) or the HDEL ER marker (ii), and costaining with the endocytic dye FM4-64 (iii) or nuclear dye DAPI (iv). The signal patterns were observed at 488 nm (eGFP: green), 546 nm (RFP or FM4-64: magenta), or 358 nm (DAPI: blue) fluorescence excitation wavelength by confocal imaging (Olympus FluoView FV1000). Bars = 50 μm. The eGFP-labeled MfNACsa is green, RFP-labeled PIP2A or HDEL or FM4-64 dye is magenta, and merged image is yellow. The ICQs of MfNACsa-eGFP against PIP2A-RFP, RFP-HDEL, FM4-64 dye, and DAPI are 0.363, 0.341, 0.239 and –0.154, respectively.