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. 2001 Mar 5;152(5):877–894. doi: 10.1083/jcb.152.5.877

Figure 3.

Figure 3

FTCD targets to the Golgi complex and organelles into fibers. COS-7 cells were transfected with GFP-tagged (A) or -untagged (B and C) rFTCD and grown for 18–48 h. (A) Cells were processed for immunofluorescence with polyclonal anti-GM130 antibodies. (B and C) Cells were processed for immunofluorescence with monoclonal anti-FTCD and polyclonal anti-GM130 antibodies. In cells expressing low levels of FTCD (A and B), the recombinant FTCD is concentrated in the Golgi region. In cells expressing high levels of FTCD (C), the recombinant FTCD also localizes to thick fibers originating from the Golgi region. (D) COS-7 cells were transfected with GFP–FTCD for 48 h and fractionated by equilibrium density centrifugation. An equivalent amount of each fraction was processed by SDS-PAGE, transferred to NC, and immunoblotted with anti-FTCD antibodies to detect the 58-kD endogenous FTCD and the 86-kD recombinant GFP–FTCD. The NC was also probed with anti-GM130 antibodies. The relative recovery of endogenous FTCD, GFP–FTCD, and GM130 in each lane were quantitatively evaluated by densitometry. Endogenous FTCD and GFP–FTCD were predominantly recovered in the high density sucrose load (fractions 9–15) containing soluble cytosolic proteins. A proportion of FTCD and GFP–FTCD were recovered in low density sucrose (fractions 5–7) enriched in the Golgi marker GM130.