ANTISENSE OLIGONUCLEOTIDES TO CUX-1, A CUT-RELATED HOMEOBOX GENE, CAUSE INCREASED APOPTOSIS IN MOUSE EMBRYONIC KIDNEY CULTURES

Cux-1 is a murine homeobox gene that is highly and transiently express ed in the developing kidney. To further evaluate the role of Cux-1 in mammalian kidney development, organotypic cultures of embryonic mouse kidney were incubated with phosphorothioate-coupled antisense Cux-1 ol igonucleotides (ODNs) in the presence of cationic liposomes. Inhibitio n of Cux-1 expression by antisense ODNs was verified by reverse transc ription-PCR. Metanephroi that were incubated with antisense Cu-l ODNs were 23% smaller than metanephroi that were incubated with sense Cux-1 ODNs. Morphologic analysis of metanephroi that were treated with anti sense Cux-1 ODNs revealed that ureteric buds and induced epithelial st ructures were present. However, extensive areas of cell. death contain ing shrunken cells with pyknotic nuclei were also evident. The presenc e of increased apoptosis was verified by ultrastructural and terminal transferase-mediated dUTP nick end labeling analyses. Two different an tisense Cux-1 ODNs targeting either the translation start codon or the homeobox produced increased apoptosis. In contrast, metanephroi incub ated with sense ODNs exhibited only occasional apoptotic cells. We con clude that the presence of antisense Cux-1 ODNs does not block nephron induction, but results instead in increased apoptosis. Proper regulat ion of Cux-1 expression may be necessary for normal kidney development .