Membrane-anchored plakoglobins have multiple mechanisms of action in Wnt signaling

In Wnt signaling, beta-catenin and plakoglobin transduce signals to the nuc leus through interactions with TCF-type transcription factors. However, whe n plakoglobin is artificially engineered to restrict it to the cytoplasm by fusion with the transmembrane domain of connexin (cnxPg), it efficiently i nduces a Wnt-like axis duplication phenotype in Xenopus. Ln Xenopus embryos , maternal XTCF3 normally represses ventral expression of the dorsalizing g ene Siamois. Two models have been proposed to explain the Wnt-like activity of cnxPg: 1) that cnxPg inhibits the machinery involved in the turnover of cytosolic beta-catenin, which then accumulates and inhibits maternal XTCF3 , and 2) that cnxPg directly acts to inhibit XTCF3 activity. To distinguish between these models, we created a series of N-terminal deletion mutations of cnxPg and examined their ability to induce an ectopic axis in Xenopus, activate a TCF-responsive reporter (OT), stabilize beta-catenin, and coloca lize with components of the Wnt signaling pathway. cnxPg does not colocaliz e with the Wnt pathway component Dishevelled, but it does lead to the redis tribution of APC and Axin, two proteins involved in the regulation of beta- catenin turnover. Expression of cnxPg increases levels of cytosolic beta-ca tenin; however, this effect does not completely explain its signaling activ ity. Although cnxPg and Wnt-1 stabilize beta-catenin to similar extents, cn xPg activates OT to 10- to 20-fold higher levels than Wnt-1. Moreover, alth ough LEF1 and TCF4 synergize with beta-catenin and plakoglobin to activate OT, both suppress the signaling activity of cnxPg. In contrast, XTCF3 suppr esses the signaling activity of both beta-catenin and cnxPg. Both exogenous XLEF1 and XTCF3 are sequestered in the cytoplasm of Xenopus cells by cnxPg . Based an these data, we conclude that, in addition to its effects on beta -catenin, cnxPg interacts with other components of the Wnt pathway, perhaps TCFs, and that these interactions contribute to its signaling activity.