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 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 approximately 60 S, and approximately 1.27 g/cm(3) 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.
Ladomery
M R
MR
MRC Human Genetics Unit, Western General Hospital, Crewe Road, Edinburgh EH4 2XU, Scotland, United Kingdom.
Slight
J
J
Mc Ghee
S
S
Hastie
N D
ND
D000818:Animals; D002460:Cell Line; D002845:Chromatography; D004268:DNA-Binding Proteins / Q000737:chemistry* / Q000235:genetics; D016162:Genes, Wilms Tumor / Q000235:genetics; D051379:Mice; D012261:Ribonucleoproteins / Q000737:chemistry* / Q000235:genetics; D014157:Transcription Factors / Q000737:chemistry* / Q000235:genetics; D014162:Transfection; D025721:WT1 Proteins; D016335:Zinc Fingers / Q000235:genetics
D004268:DNA-Binding Proteins; D012261:Ribonucleoproteins; D014157:Transcription Factors; D025721:WT1 Proteins