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Figure 5


Fig. 5. MLN51 downregulation alters SG assembly. (A) HeLa Tet-On MLN51Fl (a-l) and HeLa Tet-On MLN51Nt (m-x) were induced for 24 hours with doxycyclin and incubated in normal medium (untreated) or with arsenite for 30 minutes before fixation. Cells were double-stained with anti-MLN51 (green) and anti-FMRP (red) antibodies or with anti-MLN51 (green) antibody and a fluorescent oligo(dT) probe (red) to detect poly(A)+ RNA as indicated on the top. Note that cells expressing MLN51Nt treated with arsenite do not have FMRP or poly(A)+ RNA cytoplasmic foci compared with the non-induced cells present in the same fields (asterisks). (B) HeLa cells, transfected with Flag-tagged MLN51Nt (a-c), EYFP-tagged MLN51Nt (d-f, green) and as control EYFP-tagged MLN51Fl (g-i, green) plasmids, were treated with arsenite before fixation and probed with anti-Flag (a,c, green) and/or anti-FMRP (red, b,c,e,f,h,i) antibodies. Note that in the merged images (c,f), cells overexpressing Flag-MLN51Nt or EYFP-MLN51Nt have no FMRP-positive cytoplasmic foci (arrows). (C) HeLa control cells (a-c) were treated with hippuristanol before fixation and probed with anti-MLN51 (green, a,c) and anti-FMRP (red, b,c) antibodies. HeLa Tet-On MLN51Fl (d-f) and HeLa Tet-On MLN51Nt (g-i) were induced for 24 hours with doxycyclin and incubated with hippuristanol before fixation. Cells were double-stained with anti-MLN51 (green) and anti-FMRP (red) antibodies. Note that cells overexpressing MLN51Nt have no FMRP-positive cytoplasmic foci (arrows).





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