ABSTRACT
Constitutive activation of the Wnt signaling pathway is a root cause of many colon cancers. Activation of this pathway is caused by genetic mutations that stabilize the beta-catenin protein, allowing it to accumulate in the nucleus and form complexes with any member of the lymphoid enhancer factor (LEF1) and T-cell factor (TCF1, TCF3, TCF4) family of transcription factors (referred to collectively as LEF/TCFs) to activate transcription of target genes. Target genes such as MYC, CCND1, MMP7 and TCF7 (refs. 5-9) are normally expressed in colon tissue, so it has been proposed that abnormal expression levels or patterns imposed by beta-catenin/TCF complexes have a role in tumor progression. We report here that LEF1 is a new type of target gene ectopically activated in colon cancer. The pattern of this ectopic expression is unusual because it derives from selective activation of a promoter for a full-length LEF1 isoform that binds beta-catenin, but not a second, intronic promoter that drives expression of a dominant-negative isoform. beta-catenin/TCF complexes can activate the promoter for full-length LEF1, indicating that in cancer high levels of these complexes misregulate transcription to favor a positive feedback loop for Wnt signaling by inducing selective expression of full-length, beta-catenin-sensitive forms of LEF/TCFs.
Subject(s)
Colonic Neoplasms/genetics , Cytoskeletal Proteins/metabolism , DNA-Binding Proteins/genetics , Trans-Activators , Transcription Factors/genetics , Zebrafish Proteins , DNA-Binding Proteins/biosynthesis , Gene Expression Regulation, Neoplastic , Humans , Introns , Lymphoid Enhancer-Binding Factor 1 , Molecular Sequence Data , Neoplasm Proteins/biosynthesis , Neoplasm Proteins/genetics , Promoter Regions, Genetic , Protein Binding , Protein Isoforms , Proto-Oncogene Proteins/metabolism , Signal Transduction , T Cell Transcription Factor 1 , Transcription Factors/biosynthesis , Wnt Proteins , beta CateninABSTRACT
Proteins are directed to the nucleus by their nuclear localization sequences (NLSs) in a multistep process. The first step, which is to dock the NLS-containing protein to the nuclear pore, is carried out in part by a recently identified NLS receptor named Srp1/importin-alpha. Using the high mobility group (HMG) DNA binding domain of human lymphoid enhancer factor-1 (hLEF-1) as bait in a yeast two-hybrid screen, we have identified two different mouse Srp1 proteins (pendulin/importin-alpha and mSrp1) that each bind to a 9-amino acid sequence in hLEF-1 called the B box. We show that the B box of hLEF-1, a region essential for high affinity DNA binding, is also necessary and sufficient for nuclear localization, lending support to the model that NLSs can function both in nuclear transport and DNA binding. Pendulin and mSrp1 are the mouse homologues of hRch1/hSrp1alpha/importin-alpha and hSrp1/karyopherin alpha/NPI-1, respectively, and show considerable sequence divergence from each other. We find a surprising and significant difference in the expression pattern of pendulin and mSrp1 mRNA, suggesting that these two Srp1 proteins are distinguishable in function as well as sequence.
