Eaf1 and Eaf2 negatively regulate canonical Wnt/β-catenin signaling.
Eaf factors play a crucial role in tumor suppression and embryogenesis. To investigate the potential mechanism of Eaf activity, we performed loss- and gain-of-function assays in zebrafish using morpholino and mRNA injections, respectively. We found that eaf1 and eaf2 inhibit Wnt/β-catenin signaling, thereby modulating mesodermal and neural patterning in the embryo. Moreover, ectopic expression of eaf1 and eaf2 in embryos and cultured cells blocked β-catenin reporter activity. By immunoprecipitation, we also observed that Eaf1 and Eaf2 bound to the Armadillo repeat region and C-terminus of β-catenin, as well as to other β-catenin transcription complex proteins, such as c-Jun, Tcf and Axin, suggesting the formation of a novel complex. In addition, the N-terminus of Eaf1 and Eaf2 bound to β-catenin and exhibited dominant-negative activity, whereas the C-terminus appeared to either harbor a suppression domain or to recruit a repressor. Both the N- and C-terminus must be intact for Eaf1 and Eaf2 suppressive activity. Lastly, we demonstrate a conservation of biological activities for Eaf family proteins across species. In summary, our evidence points to a novel role for Eaf1 and Eaf2 in inhibiting canonical Wnt/β-catenin signaling, which might form the mechanistic basis for Eaf1 and Eaf2 tumor suppressor activity.
Key Laboratory of Biodiversity and Conservation of Aquatic Organisms, Institute of Hydrobiology, Chinese Academy of Sciences, Wuhan, 430072, PR China.
Jing-Xia
JX
Liu
Dawei
D
Zhang
Xunwei
X
Xie
Gang
G
Ouyang
Xing
X
Liu
Yonghua
Y
Sun
Wuhan
W
Xiao
D000818:Animals; D030801:Animals, Genetically Modified; D019521:Body Patterning / Q000235:genetics; D019556:COS Cells; D002478:Cells, Cultured; D002522:Chlorocebus aethiops; D003598:Cytoskeletal Proteins / Q000235:genetics / Q000378:metabolism / Q000502:physiology; D015536:Down-Regulation / Q000235:genetics; D004625:Embryo, Nonmammalian; D018507:Gene Expression Regulation, Developmental; D057809:HEK293 Cells; D006801:Humans; D051379:Mice; D054258:Neural Plate / Q000196:embryology / Q000378:metabolism; D051153:Wnt Proteins / Q000235:genetics / Q000378:metabolism / Q000502:physiology; D060449:Wnt Signaling Pathway / Q000235:genetics*; D015027:Zebrafish / Q000196:embryology / Q000235:genetics / Q000378:metabolism; D029961:Zebrafish Proteins / Q000235:genetics / Q000378:metabolism / Q000502:physiology*
D003598:Cytoskeletal Proteins; C542877:EAF1 protein, zebrafish; C542878:EAF2 protein, zebrafish; D051153:Wnt Proteins; D029961:Zebrafish Proteins; C094521:wnt8a protein, zebrafish