Presence of WT1, the Wilm's tumor suppressor gene product, in nuclear Poly(A)(+) ribonucleoprotein

The tumor suppressor gene WT1 encodes a zinc finger protein, which consists of four C-terminal C-2-H-2 zinc fingers of the Kruppel type, and at the N terminus a Q/P-rich trans-regulatory domain, both characteristic of transcr iption factors. However, recent findings suggest that WT1 may also be invol ved in a post-transcriptional process. Specifically, WT1 isoforms containin g the alternatively spliced exon 9 (+lysine-threonine-serine (BTrS)) prefer entially associate with nuclear speckles and coimmunoprecipitate splicing a ntigens (Larsson, S. H., Charlieu, J.-P., Miyagawa, K., Engelkamp, D., Rass oulzadegan, IM., Ross, A., Cuzin, F., van Heyningen, V., and Hastie, N. D. (1995) Cell 81, 391-401); furthermore, WT1 has been shown to interact with the ubiquitous splicing factor U2AF65 (Davies, R. C., Calvo, C., Larsson, S . H, Lamond, A. I., and Hastie, N. D. (1998) Genes Dev. 12, 3217-3225) and binds to RNA in vitro (Caricasole, A., Duarte, A., Larsson, S. H., Hastie, N. D., Little, M., Holmes, G., Todorov, I., and Ward, A. (1996) Proc. Natl. Acad, Sci. U: S. A. 93, 7562-7566; Bardeesy, N., and Pelletier, J. (1998) Nucleic Acids Res. 26, 1784-1792), To extend these findings, we have fracti onated nuclear extracts to see if particles containing WT1 have the propert ies of ribonucleoprotein (RNP). In summary, WT1 is enriched by oligo(dT) ch romatography, as are U2AF65, the U5 small nuclear RNP-associated protein pi le and hnRNP Al. Gel filtration and sedimentation profiles suggest that WT1 is present in RNase-sensitive particles, >2 MDa in size, peaking at simila r to 60 S, and similar to 1.27 g/cm(3) on Nycodenz. Similar results were ob tained from two cell lines expressing WT1, fetal kidneys (day E17), and tra nsiently transfected cells, suggesting that the presence of WT1 protein in nuclear poly(A)(+) RNP is a general aspect of WT1 function.