Regulation of Wilms Tumor Gene 1 (WT1) expression

Abstract

The Wilms Tumor gene 1 codes a powerful transcription factor that appears to play a pivotal role in the regulation of cell proliferation and differentiation. The physiological expression of WT1 is restricted to a few types of developing tissues: blood precursor cells, kidney precursors and cells in the mammary ducts. The abnormal expression of WT1 had been found in various human neoplasias. WT1 expressed in neoplasias lacks mutations and reduction of its expression blocks proliferation and leads to apoptosis or cell differentiation. There are few possible causes that could explain the abnormal expression of WT1: the gene could be mutated, which could lead to constitutive expression, an abnormal expression of physiological activators could activate WT1 expression, and an epigenetic modification of DNA could activate WT1. All three possible causes for pathological upregulation of WT1 gene were investigated in this work. The genes PAX2, PAX8 and GATA1 are physiological activators and regulators of WT1 expression. The expression patterns of these genes were analyzed in carcinoma cell lines and human carcinoma tissue samples. A close correlation was observed between expression of WT1 and PAX2 and PAX8. However, the correlation was not linear but rather of a threshold type. WT1 expression appeared when activators where expressed at certain levels. A model of an on off regulation is proposed in which the expression of an activator had to reach certain level (termed activation threshold) to activate expression of WT1. The method of RNA interference was used to further analyze the interactions between WT1 and activator genes. The genes WT1, GATA1, PAX2 and PAX8 were silenced in a series of experiments. It was shown that RNAi reduce the growth in WT1 expressing cells by 50%. However, the reduction of WT1 expression achieved with RNAi did not lead to an increase in apoptosis. Targeting of each of the regulator genes caused similar effects downregulation of the activator was followed first by downregulation of WT1 and later by downregulation of housekeeping genes. When the WT1 regulators were targeted the relative downregulation of WT1 was lower than in the case of direct targeting. When regulators were downregulated also the growth reduction was less pronounced than in cases with direct targeting of WT1. Taken together, it can be implied that WT1 is regulated by PAX2, PAX8 and GATA1.

Die Arbeit beschreibt Aspekte der Expressionsregulation vom Wilms Tumor Gen 1(WT1) bei humanen Neoplasien. Die Hauptregulatoren sind die Gene PAX2, PAX8 und GATA1. Die Experimente bezueglich RNA Interferenz zeigen den Zusammenhang zwischen der Expression von PAX2, PAX8, GATA1 und WT1. Mutationen im WT1-Gen und in den Promotoren von PAX2, PAX8 und GATA1 spielen keine Rolle bei WT1 Tumorexpression. Die Methylierung von WT1 Enhancer ist dagegen ein wichtiger Faktor.