Presence of WT1, the Wilm's Tumor Suppressor Gene Product, in Nuclear Poly(A)+ Ribonucleoprotein
Ladomery, M., Slight, J., McGhee, S. and Hastie, N. (1999) Presence of WT1, the Wilm's Tumor Suppressor Gene Product, in Nuclear Poly(A)+ Ribonucleoprotein. Journal of Biological Chemistry, 274 (51). pp. 36520-36526. ISSN 0021-9258 Available from: http://eprints.uwe.ac.uk/10909
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Publisher's URL: http://dx.doi.org/10.1074/jbc.274.51.36520
The tumor suppressor gene WT1 encodes a zinc finger protein, which consists of four C-terminal C2-H2 zinc fingers of the Krüppel type, and at the N terminus a Q/P-rich trans-regulatory domain, both characteristic of transcription factors. However, recent findings suggest that WT1 may also be involved in a post-transcriptional process. Specifically, WT1 isoforms containing the alternatively spliced exon 9 (+lysine-threonine-serine (KTS)) preferentially associate with nuclear speckles and co-immunoprecipitate splicing antigens (Larsson, S. H., Charlieu, J.-P., Miyagawa, K., Engelkamp, D., Rassoulzadegan, M., 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 fractionated nuclear extracts to see if particles containing WT1 have the properties of ribonucleoprotein (RNP). In summary, WT1 is enriched by oligo(dT) chromatography, as are U2AF65, the U5 small nuclear RNP-associated protein p116 and hnRNP A1. Gel filtration and sedimentation profiles suggest that WT1 is present in RNase-sensitive particles, >2 MDa in size, peaking at ∼60 S, and ∼1.27 g/cm3 on Nycodenz. Similar results were obtained from two cell lines expressing WT1, fetal kidneys (day E17), and transiently transfected cells, suggesting that the presence of WT1 protein in nuclear poly(A)+ RNP is a general aspect of WT1 function.