A yeast model system for functional analysis of beta-catenin signaling.
We have developed a novel Saccharomyces cerevisiae model system to dissect the molecular events of beta-catenin (beta-cat) signaling. Coexpression of mammalian beta-cat with TCF4 or LEF1 results in nuclear accumulation of these proteins and a functional complex that activates reporter gene transcription from constructs containing leukocyte enhancer factor (LEF)/T cell factor (TCF) response elements. Reporter transcription is constitutive, requires expression of both beta-cat and TCF4 or LEF1, and is not supported by mutated LEF/TCF binding elements or by TCF4 or LEF1 mutants. A cytoplasmic domain of E-cadherin or a functional fragment of adenomatous polyposis coli (APC) protein (APC-25) complexes with beta-cat, reduces beta-cat binding to TCF4, and leads to increased cytoplasmic localization of beta-cat and a reduction in reporter activation. Systematic mutation of putative nuclear export signal sequences in APC-25 decreases APC-25 binding to beta-cat and restores reporter gene transcription. Additional beta-cat signaling components, Axin and glycogen synthase kinase 3beta, form a multisubunit complex similar to that found in mammalian cells. Coexpression of the F-box protein beta-transducin repeat-containing protein reduces the stability of beta-cat and decreases reporter activation. Thus, we have reconstituted a functional beta-cat signal transduction pathway in yeast and show that beta-cat signaling can be regulated at multiple levels, including protein subcellular localization, protein complex formation, and protein stability.
Lee
Margaret S
MS
Aventis Pharmaceuticals, Cambridge Genomics Center, Cambridge, MA 02139, USA.
D'Amour
Karen A
KA
Papkoff
Jackie
J
D021581:Active Transport, Cell Nucleus; D025601:Adenomatous Polyposis Coli Protein / Q000378:metabolism; D000409:Alanine / Q000378:metabolism; D019943:Amino Acid Substitution; D060466:Axin Protein; D001693:Biological Transport, Active; D015820:Cadherins / Q000378:metabolism; D017871:Calcium-Calmodulin-Dependent Protein Kinases / Q000737:chemistry / Q000378:metabolism; D003593:Cytoplasm / Q000378:metabolism; D003598:Cytoskeletal Proteins / Q000378:metabolism / Q000502:physiology*; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism; D017353:Gene Deletion; D005786:Gene Expression Regulation; D017930:Genes, Reporter; D038341:Glycogen Synthase Kinases; D006801:Humans; D051821:Lymphoid Enhancer-Binding Factor 1; D008954:Models, Biological; D009842:Oligopeptides; D010455:Peptides / Q000378:metabolism; D011506:Proteins / Q000737:chemistry / Q000378:metabolism; D012097:Repressor Proteins; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000378:metabolism*; D015398:Signal Transduction; D051820:TCF Transcription Factors; D015534:Trans-Activators / Q000378:metabolism / Q000502:physiology*; D058118:Transcription Factor 7-Like 2 Protein; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism; D015293:Transducin / Q000378:metabolism; D051176:beta Catenin
D025601:Adenomatous Polyposis Coli Protein; D060466:Axin Protein; C495270:CTNNB1 protein, human; D015820:Cadherins; D003598:Cytoskeletal Proteins; D004268:DNA-Binding Proteins; C497573:LEF1 protein, human; D051821:Lymphoid Enhancer-Binding Factor 1; D009842:Oligopeptides; D010455:Peptides; D011506:Proteins; D012097:Repressor Proteins; D051820:TCF Transcription Factors; C544894:TCF7L2 protein, human; D015534:Trans-Activators; D058118:Transcription Factor 7-Like 2 Protein; D014157:Transcription Factors; D051176:beta Catenin; C085706:FLAG peptide; D038341:Glycogen Synthase Kinases; D017871:Calcium-Calmodulin-Dependent Protein Kinases; D015293:Transducin; D000409:Alanine