Figure 7.
Subcellular Localization of AtRAC10.
(A) N. benthamiana leaves that expressed GFPAtRAC10 (AtRAC10) or GFPAtrac10mSS202+208 (Atrac10mSS) were plasmolyzed. The majority of the wild-type protein was detected in the plasma membrane (PM), with smaller fractions in nuclei (n) (AtRAC10, I and II). In a few cells, cytoplasmic thickening (C) was detected (AtRAC10, II). The mutant protein (Atrac10mSS) was detected only in nuclei and cytoplasmic thickening.
(B) Immunoblot analysis of a protein with anti-GFP monoclonal antibodies after transient expression of GFPAtRAC10 (AtRAC10) or free GFP (GFP) in leaves of N. benthamiana, centrifugal separation to soluble (S) and insoluble pellet (P) fractions, and separation by SDS-PAGE.
(C) A mutant of AtRAC8 containing the four C-terminal residues of AtRAC10 (Atrac8m10C′) was localized only in the plasma membrane, similar to wild-type AtRAC8.
Images in (A) are differential interference contrast and fluorescence overlays. All images are confocal scans taken with a confocal laser scanning microscope. Numbers at left in (B) denote molecular mass in kD. Bars = 20 μm.