Conditional mutations of beta-catenin and APC reveal roles for canonical Wnt signaling in lens differentiation.
OBJECTIVE
Previous studies indicate that the Wnt/beta-catenin-signaling pathway is active and functional during murine lens development. In this study, the consequences of constitutively activating the pathway in lens during development were investigated.


METHODS
To activate Wnt/beta-catenin signaling, beta-catenin (Catnb) and adenomatous polyposis coli (Apc) genes were conditionally mutated in two Cre lines that are active in whole lens (MLR10) or only in differentiated fibers (MLR39), from E13.5. Lens phenotype in mutant lenses was investigated by histology, immunohistochemistry, BrdU labeling, quantitative RT-PCR arrays, and TUNEL.


RESULTS
Only intercrosses with MLR10 resulted in ocular phenotypes, indicating Wnt/beta-catenin signaling functions in lens epithelium and during early fiber differentiation. Mutant lenses were characterized by increased progression of epithelial cells through the cell cycle, as shown by BrdU labeling, and phosphohistone 3 and cyclin D1 labeling, and maintenance of epithelial phenotype (E-cadherin and Pax6 expression) in the fiber compartment. Fiber cell differentiation was delayed as shown by reduced expression of c-maf and beta-crystallin and delay in expression of the CDKI, p57(kip2). From E13.5, there were numerous cells undergoing apoptosis, and by E15.5, there was evidence of epithelial-mesenchymal transition with numerous cells expressing alpha-smooth muscle actin. Quantitative PCR analyses revealed large changes in expression of Wnt target genes (Lef1, Tcf7, T (Brachyury), and Ccnd1), Wnt inhibitors (Wif1, Dkk1, Nkd1, and Frzb) and also several Wnts (Wnt6, Wnt10a, Wnt8b, and Wnt11).


CONCLUSIONS
These data indicate that the Wnt/beta-catenin pathway plays key roles in regulating proliferation of lens stem/progenitor cells during early stages of fiber cell differentiation.
Martinez
Gemma
G
Ocular Development Laboratory, Department of Anatomy and Cell Biology, University of Melbourne, Victoria, Australia.
Wijesinghe
Mary
M
Turner
Kirsty
K
Abud
Helen E
HE
Taketo
Makoto M
MM
Noda
Tetsuo
T
Robinson
Michael L
ML
de Iongh
Robb U
RU
D025601:Adenomatous Polyposis Coli Protein / Q000235:genetics*; D000818:Animals; D017209:Apoptosis; D001973:Bromodeoxyuridine / Q000378:metabolism; D018797:Cell Cycle Proteins / Q000378:metabolism; D002454:Cell Differentiation / Q000502:physiology*; D049109:Cell Proliferation; D004847:Epithelial Cells / Q000166:cytology; D005260:Female; D019084:Fluorescent Antibody Technique, Indirect; D016368:Frameshift Mutation; D020287:In Situ Nick-End Labeling; D007908:Lens, Crystalline / Q000196:embryology*; D008297:Male; D051379:Mice; D008822:Mice, Transgenic; D010641:Phenotype; D020133:Reverse Transcriptase Polymerase Chain Reaction; D015398:Signal Transduction / Q000502:physiology*; D051153:Wnt Proteins / Q000502:physiology*; D051176:beta Catenin / Q000235:genetics*
D025601:Adenomatous Polyposis Coli Protein; D018797:Cell Cycle Proteins; D051153:Wnt Proteins; D051176:beta Catenin; D001973:Bromodeoxyuridine