Convergence of Wnt, growth factor, and heterotrimeric G protein signals on the guanine nucleotide exchange factor Daple

Abstract

Cellular proliferation, differentiation, and morphogenesis are shaped by multiple signaling cascades; their concurrent dysregulation plays an integral role in cancer progression and is a common feature of many malignancies. Three such cascades that contribute to the oncogenic potential are those mediated by Wnt and Frizzled (FZD), growth factor receptor tyrosine kinases (RTKs), and trimeric G proteins and GPCRs. Daple is a Dishevelled (Dvl)- and FZD-binding protein and a guanine-nucleotide exchange factor (GEF) for the trimeric G protein, Gαi. Daple enhances β-catenin-independent Wnt signaling through its ability to activate the pathway downstream of the Wnt5/FZD7 pathway. Here we identified that Daple is a substrate of multiple RTKs and non-RTKs, and hence, a point of convergence for all three cascades. We show that phosphorylation by both RTKs and non-RTKs near the Dvl-binding motif in Daple dissociated Daple:Dvl complexes and augmented the ability of Daple to bind and activate Gαi which potentiated β-catenin-independent Wnt signals and triggered epithelial-mesenchymal transition (EMT) in a manner similar to that triggered by Wnt5A/FZD. Although Daple acts as a tumor suppressor in the healthy colon, but concurrent upregulation of Daple and epidermal growth factor receptor (EGFR) in colorectal tumors from patients was associated with poor prognosis. We conclude that Daple-dependent activation of Gαi and enhancement of β-catenin-independent Wnt signals is not just triggered by Wnt5a/FZD to suppress tumorigenesis, but also is hijacked by growth factor-RTKs to stoke tumor progression. Thus, this work defines a crosstalk paradigm amongst growth factor RTKs, trimeric G-proteins, and Wnt/FZD in cancer biology.

Tyrosine-based Signals Regulate the assembly of Daple•PARD3 Complex at Cell-cell Junctions during Polarized Planar Cell Migration

“…First discovered in a yeast-two hybrid screen through its ability to bind to the PDZ domain on Disheveled (Dvl), Daple has emerged as a key modulator of Wnt signaling [9,11]. The extreme C-terminus on Daple contains an 'atypical' PBM which we, and others, demonstrated to be necessary for the Daple•Dvl interaction [10][11][12]. We subsequently showed that Daple binds Gαi proteins and triggers GTPase signaling downstream of the Wnt receptor Frizzled-7 upon activation by Wnt5a; such signaling is necessary for the activation of the b-catenin-independent Wnt signaling pathway (a.k.a.…”

“…We began by asking if the newly discovered Daple•PARD3-PDZ interaction affects the Daple•Gai interaction. We had previously described an unexpected allosteric phenomenon in which the Daple•Dvl-PDZ and the Daple•Gai interactions are mutually exclusive, i.e., ternary complexes between all 3 proteins, Gai•Daple•Dvl is not feasible [10]. To our surprise, such allostery was not observed in the case of PARD3; instead, we found that Gai3 bound PARD3 exclusively in the presence of Daple, and not directly, indicative of the formation of tertiary Gai•Daple•PARD3 complexes (Figure 5G).…”

“…The two tyrosines immediately adjacent to Daple's PBM have previously been shown to serve as a platform for convergence of signaling downstream of multiple receptor and non-receptor tyrosine kinases (TKs) like Src [10] (Figure 6A). We asked if these phosphoevents can dictate which PDZ proteins and/or Gai may bind Daple.…”

“…Daple (CCDC88C) is a large, multi-modular PBM-containing scaffold protein [9,10]. First discovered in a yeast-two hybrid screen through its ability to bind to the PDZ domain on Disheveled (Dvl), Daple has emerged as a key modulator of Wnt signaling [9,11].…”

“…non-canonical Wnt signaling). More recently, we showed Dapledependent Wnt signaling is also shaped by Akt and multiple tyrosine kinases (TKs); in each case, phosphorylation of Daple by these kinases impacted its localization and signaling [9,10,13]. We also showed that Daple has two opposing roles during colorectal cancer (CRC) progression; much like the non-canonical Wnt pathway, Daple acts as a tumor suppressor in the normal epithelium and early during cancer initiation, but it serves as a potent trigger for EMT and tumor cell invasion later during cancer dissemination.…”