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Wnt regulation of gene expression requires binding of LEF/T-cell factor (LEF/TCF) transcription factors toWnt response elements (WREs) and recruitment of the activator ␤-catenin. There are significant differences in the abilities of LEF/TCF family members to regulate Wnt target genes. For example, alternatively spliced isoforms of TCF-1 and TCF-4 with a C-terminal "E" tail are uniquely potent in their activation of LEF1 and CDX1. Here we report that the mechanism responsible for this unique activity is an auxiliary 30-amino-acid DNA interaction motif referred to here as the "cysteine clamp" (or C-clamp). The C-clamp contains invariant cysteine, aromatic, and basic residues, and surface plasmon resonance (SPR) studies with recombinant C-clamp protein showed that it binds double-stranded DNA but not single-stranded DNA or RNA (equilibrium dissociation constant ‫؍‬ 16 nM). CASTing (Cyclic Amplification and Selection of Targets) experiments were used to test whether this motif influences WRE recognition. Full-length LEF-1, TCF-1E, and TCF-1E with a mutated C-clamp all bind nearly identical WREs (TYYCTTTGATSTT), showing that the C-clamp does not alter WRE specificity. However, a GC element downstream of the WRE (RCCG) is enriched in wild-type TCF-1E binding sites but not in mutant TCF-1E binding sites. We conclude that the C-clamp is a sequencespecific DNA binding motif. C-clamp mutations destroy the ability of ␤-catenin to regulate the LEF1 promoter, and they severely impair the ability of TCF-1 to regulate growth in colon cancer cells. Thus, E-tail isoforms of TCFs utilize two DNA binding activities to access a subset of Wnt targets important for cell growth.

BackgroundReducing sedentary time and increasing lifestyle activities, including light-intensity activity, may be an option to help prevent metabolic syndrome (MetS). The purpose of the present study was to examine whether objectively measured light-intensity lifestyle activity and sedentary time is associated with MetS, independent of moderate–vigorous intensity physical activity (MVPA).MethodsThe participants in this cross-sectional study were 483 middle-aged Japanese adults, aged 30–64 years. The participants were divided into those with or without MetS according to the Japanese criteria for MetS. A triaxial accelerometer was used to measure light-intensity lifestyle activity [1.6–2.9 metabolic equivalents (METs)] and sedentary time (≤1.5 METs). Logistic regression was used to predict MetS from the levels of light-intensity lifestyle activity and sedentary time with age, sex, smoking, calorie intake, accelerometer wear time, and MVPA as covariates.ResultsThe odds ratios (OR) for MetS in the highest and middle tertiles of light-intensity lifestyle activity were 0.44 [95% confidence interval (CI): 0.24 to 0.81] and 0.51 (95% CI: 0.29 to 0.89) relative to the lowest tertile, after adjustment for age, sex, smoking, calorie intake, accelerometer wear time and MVPA (Ptrend = 0.012). Sedentary time was also associated with the risk of MetS (Ptrend = 0.018). Among participants in the highest tertile of sedentary time, the risk of MetS was 2.27-times greater than that in the lowest tertile (95% CI: 1.25 to 4.11). The risk of MetS was not significantly increased in subjects in the middle tertile of sedentary time.ConclusionsWe found that light-intensity lifestyle activity and sedentary time were significantly associated with the risk of MetS, independent of MVPA. The results of our study suggest that public health messages and guidelines should be refined to include increases in light-intensity lifestyle activity and/or decreases in sedentary time, alongside promoting MVPA, to prevent MetS.

Alternative promoters within the LEF1 locus produce polypeptides of opposing biological activities. Promoter 1 produces full-length LEF-1 protein, which recruits ␤-catenin to Wnt target genes. Promoter 2 produces a truncated form that cannot interact with ␤-catenin and instead suppresses Wnt regulation of target genes. Here we show that promoter 1 is aberrantly activated in colon cancers because it is a direct target of the Wnt pathway. T-cell factor (TCF)-␤-catenin complexes bind to Wnt response elements in exon 1 and dynamically regulate chromatin acetylation and promoter 1 activity. Promoter 2 is delimited to the intron 2/exon 3 boundary and, like promoter 1, is also directly regulated by TCF-␤-catenin complexes. Promoter 2 is nevertheless silent in colon cancer because an upstream repressor selectively targets the basal promoter leading to destabilized TCF-␤-catenin binding. We conclude that the biological outcome of aberrant LEF1 activation in colon cancer is directed by differential promoter activation and repression.

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