Fig. 7. (i) Confocal sections of HeLa cells injected with expression vectors encoding YFPSmB or GFPASF/SF2 in the presence or absence of LMB and incubated for 8 hours. In cells injected with YFPSmB in the presence of LMB (A-D), expressed YFPSmB does not accumulate in nuclear bodies or speckles (C). Nuclear speckles are still seen using antibodies to endogenous snRNPs (A). p80-coilin is found predominantly in the nucleolus (arrows in B). Control cells, injected with pYFPSmB in the absence of LMB (E-H) show a speckled pattern of YFPSmB (G), with endogenous Sm proteins in speckles and Cajal bodies (E) and p80-coilin in Cajal bodies (F). Cells injected with pGFPASF/SF2 in the presence of LMB (I-L) show the uptake of GFPASF/SF2 into speckles (K). p80-coilin is seen in the nucleoli (arrows in J,L). Bar, 10 µm. (ii) Immonoprecipitation of snRNPs from cells transfected in the presence (A) or absence (B) of LMB using anti-TMG antibodies, detected using anti-GFP antibodies (top panels). In the presence of LMB, GFP-SmD1 is not incorporated into snRNPs (compare lower band in bound versus unbound fraction). In the absence of LMB, GFP-SmD1 is seen in the bound, snRNP, fraction. The lower panels show 3D projections of serial confocal sections through cells treated with LMB (A) and control cells (B). In LMB-treated cells, GFP-SmD1 signal is diffuse, whereas in control cells (B), GFPSmD1 is seen in Cajal bodies and nucleoli.
