FIG. 6.

D. melanogaster NXF1-2xUBA partially rescues export in cells depleted of NXF1. (A to F) SL2 cell lines constitutively expressing GFP fusions with wild-type NXF1 (wt), NXF1-1xUBA, or NXF1-2xUBA were fixed in formaldehyde. Bulk poly(A)⁺ RNA was detected by FISH (B, D, and F). (G to P) SL2 cells stably transfected with plasmids expressing GFP or GFP-NXF1-2xUBA were transfected with a dsRNA corresponding to the NTF2-like domain of D. melanogaster NXF1 (G to L) or to the coding region of D. melanogaster p15(M to P). Five days after transfection, poly(A)⁺ RNA was detected by FISH with an indocarbocyanine-labeled oligo(dT) probe. In the GFP-expressing cell line, depletion of NXF1 causes nuclear accumulation of poly(A)⁺ RNA regardless of whether GFP is expressed (G and H). In the GFP-NXF1-2xUBA cell line, a strong nuclear accumulation of poly(A)⁺ RNA was observed in cells in which the protein was not detected, but when D. melanogaster NXF1-2xUBA was expressed at detectable levels, mRNA export was restored (I to L, arrowheads). This restoration of export was also observed when p15 was depleted (M to P).