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Figure 3


Fig. 3. Intracellular localization of DRM-tagged CLIMP-63 related constructs in M3/18 cells. M3/18 cells grown at 39.5°C were transfected with recombinant plasmids encoding the corresponding fusion proteins. Images were collected with an LSM510 microscope equipped with a Plan-Apochromate 100x/1.4 oil DIC lens at scan-zoom set at `1' and line average set at `4'. Due to low fluorescent intensity of stably expressed GFP-Dad1 the pinholes were set wide-open. Expression of GFP-Dad1 was visualized using the argon laser (a,d,g,j,m). The HeNe1 laser was used to visualize DRM-tagged constructs (b,e,h,k,n). To assess colocalization of two proteins, the images of both channels were merged (c,f,i,l,o). Expression of p63-DRM and {Delta}Lp63-DRM fusion constructs revealed that these fusion proteins are localizing to the ER (a-c and d-f, respectively). No changes in the ER morphology were observed. Expression of the soluble {Delta}TLp63-DRM and DRM revealed that these proteins are localizing to the cytosol (j-l and m-o, respectively). The morphology of the ER is not altered. Expression of the DRM-{Delta}Cp63 construct lead to some accumulation in a perinuclear area, while the morphology of the ER appeared was not affected (g-i). {Delta}TLp63-DRM apparently had cytosolic localization (j-l) similar to that of red fluorescent tag, DRM (m-o). Bar, 20 µm.





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