The Sp family of transcription factors in the regulation of the human and mouse MUC2 gene promoters

Modulation of mucin gene expression is an important component both in the e arly steps of colon cancer development and in later tumor progression. Prev ious work from our laboratory and others has suggested that the Sp family o f transcription factors mag play an important role in the regulation of the human MUC2 gene. To determine whether this was an essential element, we ex tended our work to the cloning and analysis of 3.5 lib of the 5'-flanking r egion of the mouse Muc2 (mMuc2) gene. Comparative analysis between the mous e and human MUC2 promoter regions has identified a strong sequence homology between the mouse and human genes, including the presence of GC-rich boxes , the location and composition of which are maintained in the mouse and hum an genes. We show that these GC boxes are binding sites for Sp-family trans cription factors and are functionally important since mithramycin, an inhib itor of Sp1/Sp3 binding, blocks MUC2 gene expression in HT29 cells. Further more, by a combination bf gel shift analysis and site-directed mutagenesis, we have identified the relative contribution of individual GC boxes, and o f the factors they bind, to the regulation of the mouse Muc2 promoter, whic h appears to be different in the mouse and human genes. Finally, we demonst rate by overexpressing Spl and Sp3 that the functional difference between t he proximal promoter region of the MUC2 gene in the two species is not attr ibutable to differential ability of this region to bind members of the Sp f amily of transcription factors, but rather to the different anatomy of the individual GC boxes in the mouse and human proximal promoters.