Wnt1 expression slows growth and increases cell size in monolayer cultures of NIKs cells. The growth of NIKs cells transduced either with a control retrovirus (lacZ) or a Wnt1 expression vector (Wnt1-IRES-lacZ) was compared (A: total cell counts). When cultures were stained for lacZ expression (B), the number of large and very large cells expressing Wnt1 was greatly increased (C). Canonical Wnt signaling was measured as fold induction of a consensus TCF-βcatenin reporter (TOP-FLASH) with respect to FOP-FLASH (a scrambled consensus reporter, all normalized for transfection efficiency as previously described). The TOP/FOP ratio was increased in NIKs cells by 2× when cells were incubated with soluble (crude) Wnt3A for 22 hours, and reduced if the cells were transfected with the canonical Wnt inhibitor, dkk1, prior to Wnt3A incubation (D). Cell strains stably transduced with retroviral constructs (LZ, lacZ control; W, Wnt1-lacZ test vector) were transfected with either FOP-FLASH or TOP-FLASH reporters (E). The Wnt1-expressing culture showed a significant increase in Wnt-dependent transactivation (n = 3), though less than the naïve cultures treated with soluble Wnt3A (probably because of negative feedback).