WT1 – A Complex Life

1. WT1 – A Complex Life By: Chen Zhu

2. WT1 – Outline • Introduction • Function • Structure • Role in development • In mice • In humans • Role in cancer • Wilm’s Tumor, related illnesses

3. Introduction • Wilms’ Tumor 1 (WT1)is found on human chromosome 11p13 • WT1 is important for the development of the genitourinary system and mesothelial tissues. • The inactivation of WT1 is responsible for ~10-15% of Wilms’ tumor

4. Function • WT1 is a transcriptional and post-transcriptional regulator • It contains signals that allow it to localize inside the nucleus • It contains domains that can bind to several DNA sequences

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6. Structure • WT1 is a 50kb gene with ten exons, together encoding a mRNA of ~3kb. • The N-terminus of WT1 contains a proline and glutamine-rich region that facilitates homodimerization • There are two alternatively spliced exons in WT1

7. Exon 5 • Exon 5 encodes 17 amino acids between the N-terminus and the first Zn finger domain • Encodes a protein-protein interaction domain

8. Exon 9 • Encodes the sequence KTS between the third and fourth zinc fingers of the protein • WT1 (-KTS) variants have been shown to act as transcriptional regulators1 • WT1 (+KTS) may play a role in RNA processing.

9. Adapted from Figure 5 in Keith W. Brown and Karim T.A. Malik, “The molecular biology of Wilms’ Tumor,” Exp. Rev. Mol. Med. 14 May, http://www.expertreviews.org/01003027h.htm

10. Alternate Splicing • 24 alternate isoforms of WT1 have been described • (+KTS)/(-KTS) is by far the most important alternate splicing mechanism

11. Function • Many target genes of WT1 have been identified • The role of WT1 in transcriptional regulation, however, is not well understood

12. Function • We do understand, however, that WH1 is involved in many different pathways: • The differentiation of leukemic cell lines • The differentiation of pheochromocytoma cells • Both the induction and prevention of apoptosis

13. WT1 Target Genes2

14. Role In Development • WT1 is expressed during development in the following systems: • Urinogenital system • Epicardium and subepicardial mesenchyme (heart) • Kidneys • Spleen • Parts of the brain • Spinal chord • Mesothelial organs • Diaphram

15. Jordan A. Kreidberg, et al., “WT-1 is required for early liver development,” Cell 74: 679-691 (1993)

16. Knockout Mice • WT1 (-/-) die from embryonic day 12 to the end of gestation • Many defects are evident in many of the areas mentioned in the previous slide • Mutant embryos lack kidneys and gonads

17. Heterozygous Mice • WT1 (+/-) mice usually die after several months due to renal insufficiency • The severity of symptoms can be correlated with WT1 levels • Symptoms are similar to the human Frasier Syndrome

18. Jordan A. Kreidberg, et al., “WT-1 is required for early liver development,” Cell 74: 679-691 (1993)

19. Kidney Development • Loose mesenchyme forms epithelial condenses around ureteric bud tips • S-shaped bodies develop and eventually elongate to attach to the collecting duct

20. Kidney Development -The role of WT1 • Without WT1, the loose mesenchyme becomes apoptotic • WT1 may thus act as a survival factor for populations of embryonic kidney cells during development • An antiapoptotic protein encoded by Bcl-2 may be an upstream mediator of WT1

21. Heart Development • WT1(-/-) mutants show smaller hearts and thinning of muscular walls • The epicarium, especially, requires WT1 • WT1 is thought to play a role in the transformation of epicardial to mesenchymal cells

22. Jordan A. Kreidberg, et al., “WT-1 is required for early liver development,” Cell 74: 679-691 (1993)

23. Role In Cancer • WAGR: heterozygous deletions at chromosome 11p13 • Frasier Syndrome : heterozygous point mutations in intron 9 • Leukemia: heterozygous WT1 mutations

24. WAGR • WAGR is a syndrome for affected children predisposed to develop: Wilms’ tumor Aniridia Genitourinary anomalies mental Retardation http://bms.brown.edu/pedisurg/images/ImageBank/Abdomen/WilmsCT8yo.jpg

25. WAGR • Aniridia • Genitourinary Anomalies • Mental Retardation http://www.abdn.ac.uk/clsm/UserFiles/Image/collinson-eye-large.jpg

26. Wilm’s Tumor • Wilms’ Tumor is a cancer of the kidney • About 500 cases are diagnosed yearly • Onset is early, with nearly everybody diagnosed under the age of 25 • Survival with therapy is very good: ~85% • WAGR patients should undergo frequent ultrasound testing

27. Wilm’s Tumor • Wilm’s tumor arises from mesenchyme that fails to differentiate into the epithelial component of the nephron • Inactivation of WT1 produces foci of primitive renal cells called nephrogenic rests • Sometimes called tri-phasic type, consisting of undifferentiated mesenchyme, stromal and epiphelial cells

28. Leukemia • WT1 is inappropriately expressed in many Leukemia cell lines (~15%) • WT1 is usually expressed in stem cells of bone marrow, but not in normal mature blood cells, linking it to an immature state for leukemia cells • WT1 expression levels increase during the progression of acute myeloid leukemia • Patients with low WT1 mRNA had a better prognosis than those with high WT1 mRNA

29. Questions?

30. Bibliography • Holger Scholz and Karin M. Kirschner, “A Role for the Wilms’ Tumor Protein WT1 in Organ Development,” Physiology 20: 54-59 (2005) • Volkher Scharnhorst, et al., “WT1 proteins: functions in growth and differentiation,” Gene 273: 141-161 (2001) • Kay-Dietrich Wagner, et al., “The complex life of WT1,” Journal of Cell Science 116: 1653-1658 (2003) • Jordan A. Kreidberg, et al., “WT-1 is required for early liver development,” Cell 74: 679-691 (1993) • National Cancer institute, “Wilms’ Tumor and Other Childhood Kidney Tumors,” November 27, 2006. <http://www.cancer.gov/cancertopics/pdq/treatment/wilms/Patient> • International WAGR Syndrome Association, “What is WAGR Syndrome?” Accessed April 27, 2007. <http://www.wagr.org/> • Monika L. Metzger, Jeffrey S. Dome., “Current Therapy for Wilms’ Tumor,” Oncologist 10: 815-826 (2005)