Figure 5: Tcf3 promotes cell migration non-cell autonomously and independently of its role as a cofactor to β-catenin.
From: Tcf3 promotes cell migration and wound repair through regulation of lipocalin 2
(a) Real-time PCR analysis of level of expression of transduced tet-inducible Tcf3 and its mutant versions, Tcf3ΔN and Tcf3ΔG, in response to Dox. Keratinocytes were transduced with GFP-tagged, tet-inducible lentivectors carrying myc-tagged Tcf3 or its mutant versions. Transduced cells were isolated by fluorescent cell sorting and were treated with Dox or vehicle control for 24 h prior to being harvested for messenger RNA analysis. Mean expression values from biological duplicates were calculated. Data are mean±s.d. (Student’s t-test). (b) Images of transduced keratinocytes expressing tet-inducible Tcf3, Tcf3ΔN, Tcf3ΔG 16 h after the initiation of migration assay with or without Dox. Black bar denotes 200 μm. (c) Graph quantifying the area migrated by transduced cells treated with Dox relative to the area migrated by cells treated with vehicle control. For each sample, over 30 non-overlapping fields were measured at each time point; and each experiment was repeated twice. Data are mean±s.d. **P<0.001 (Student’s t-test). (d) Tcf3-induced cells secrete factors that promote cell migration. Keratinocytes from tet-inducible Tcf3 (K14-rtTA;TRE-mycTcf3) and control (K14-rtTA) mice were cultured in the presence or absence of Dox for 48 h and their conditioned media were collected. Wild-type keratinocytes were subjected to a migration assay in the presence of specified conditioned media. Images were taken 38 h after the initiation of migration assay. Scale bar, 200 μm. (e) Conditioned media from Tcf3-induced cells (K14-rtTA;TRE-mycTcf3 +Dox) promotes migration of wild-type cells. Graph quantifying the migrated area of wild-type cells that were incubated with Dox-treated conditioned media during the migration assay relative to the area migrated by cells treated with vehicle-treated conditioned media. For each sample, over 30 non-overlapping fields were measured at each time point; and each experiment was repeated three times. Data are mean±s.d. **P<0.001 (Student’s t-test).
