Transcriptional program induced by Wnt protein in human fibroblasts suggests mechanisms for cell cooperativity in defining tissue microenvironments - PubMed (original) (raw)

Transcriptional program induced by Wnt protein in human fibroblasts suggests mechanisms for cell cooperativity in defining tissue microenvironments

Zach Klapholz-Brown et al. PLoS One. 2007.

Abstract

Background: The Wnt signaling system plays key roles in development, regulation of stem cell self-renewal and differentiation, cell polarity, morphogenesis and cancer. Given the multifaceted roles of Wnt signaling in these processes, its transcriptional effects on the stromal cells that make up the scaffold and infrastructure of epithelial tissues are of great interest.

Methods and results: To begin to investigate these effects, we used DNA microarrays to identify transcriptional targets of the Wnt pathway in human lung fibroblasts. Cells were treated with active Wnt3a protein in culture, and RNA was harvested at 4 hours and 24 hours. Nuclear accumulation of ss-Catenin, as shown by immunofluorescence, and induction of AXIN2 demonstrate that fibroblasts are programmed to respond to extracellular Wnt signals. In addition to several known Wnt targets, we found many new Wnt induced genes, including many transcripts encoding regulatory proteins. Transcription factors with important developmental roles, including HOX genes, dominated the early transcriptional response. Furthermore, we found differential expression of several genes that play direct roles in the Wnt signaling pathway, as well as genes involved in other cell signaling pathways including fibroblast growth factor (FGF) and bone morphogenetic protein (BMP) signaling. The gene most highly induced by Wnt3a was GREMLIN2, which encodes a secreted BMP antagonist.

Conclusions: Elevated expression of GREMLIN2 suggests a new role for Wnt signals in the maintenance of stem cell niches, whereby Wnt signals induce nearby fibroblasts to produce a BMP antagonist, inhibiting differentiation and promoting expansion of stem cells in their microenvironment. We suggest that Wnt-induced changes in the gene expression program of local stromal cells may play an important role in the establishment of specialized niches hospitable to the self-renewal of normal or malignant epithelial stem cells in vivo.

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Conflict of interest statement

Competing Interests: The authors have declared that no competing interests exist.

Figures

Figure 1. Immunofluorescence analysis comparing ß-Catenin localization in Wnt-treated and mock-treated human dermal fibroblasts.

Immunofluorescence staining shows accumulation of nuclear ß-Catenin in Wnt-treated cells, but no expression in “mock” treated cells (after 4 hrs of Wnt treatment). DAPI was used to stain cell nuclei (red).

Figure 2. Wnt-regulated genes in human lung fibroblasts.

A) SAM analysis was performed to select for genes that were differentially expressed in at 4 hour, 24 hour or both (averages), in Wnt-treated cells compared to the corresponding time zero and mock-treated fibroblasts, using an FDR of less than 1% as the cut-off. In this display, each row represents a specific gene, and each column a sample from the Wnt-treatment time course experiment. A red color indicates an increase in transcript levels of a given gene relative to the pre-treatment (time zero) level and green indicates decreased transcript levels relative to time zero. Grey represents missing or excluded data. The legend bar maps color versus magnitude of changes in transcript levels relative to the pre-treatment level. A version of this figure with every row labeled by gene name is available as Supplemental Figure S1. B) Shows expression patterns of Wnt-responsive genes encoding DNA binding proteins/transcription factors and transcriptional cofactors. C) Shows expression patterns of Wnt-responsive genes encoding proteins with direct roles in the Wnt signaling system. D) Shows expression patterns of Wnt-responsive genes encoding extracellular signaling molecules. E) Shows expression of Wnt-responsive genes encoding cell surface molecules and receptors.

Figure 3. A possible dual role for Wnt in the maintenance of stem cell niches.

We hypothesize that Wnt may promote self-renewal directly by raising nuclear ß-Catenin levels in stem cells and indirectly by activating the BMP inhibitor Gremlin 2 in nearby fibroblasts, thereby blocking target BMPs from promoting differentiation and loss of self-renewal capacity.

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Transcriptional program induced by Wnt protein in human fibroblasts suggests mechanisms for cell cooperativity in defining tissue microenvironments - PubMed (original) (raw)