Wnt signaling has emerged as a major regulator of tissue development by governing the self-renewal and maintenance of stem cells in most tissue types. As a key upstream regulator of the Wnt pathway, the transmembrane E3 ligase ZNRF3 has recently been established to play a role in negative regulation of Wnt signaling by targeting Frizzled (FZD) receptor for ubiquitination and degradation. However, the upstream regulation of ZNRF3, in particular the turnover of ZNRF3, is still unclear. Here we report that ZNRF3 is accumulated in the presence of proteasome inhibitor treatment independent of its E3-ubiquitin ligase activity. Furthermore, the Cullin 1-specific SCF complex containing β-TRCP has been identified to directly interact with and ubiquitinate ZNRF3 thereby regulating its protein stability. Similar with the degradation of β-catenin by β-TRCP, ZNRF3 is ubiquitinated by β-TRCP in both CKI-phosphorylation- and degron-dependent manners. Thus, our findings not only identify a novel substrate for β-TRCP oncogenic regulation, but also highlight the dual regulation of Wnt signaling by β-TRCP in a context-dependent manner where β-TRCP negatively regulates Wnt signaling by targeting β-catenin, and positively regulates Wnt signaling by targeting ZNRF3.
Cells, Cultured ; Humans ; Proteolysis ; Ubiquitin-Protein Ligases ; metabolism ; Ubiquitination ; beta-Transducin Repeat-Containing Proteins ; metabolism

OBJECTIVETo employ single nucleotide polymorphisms (SNP) microarray to detect copy number variations (CNVs) for the diagnosis of disease and molecular classification.

METHODSFor a patient with split-hand/split-foot malformation, genome-wide copy number variants SNP microarray was applied. Tiny copy number variations were verified by real-time fluorescent quantitative PCR.

RESULTSThe results of SNP microarray has revealed that the patient has carried a 0.39 Mb duplication in 10q24.31-24.32 (102 955 122-103 348 688), which has encompassed genes including LBX1, BTRC and POLL. By real-time fluorescent quantitative PCR, duplicate area encompassing the pathogenic genes have been verified. The results for LBX1, BTRC, POLL genes were all consistent with the SNP microarray test. Moreover, a duplication was detected in exon 9 of FBXW4 gene which is in nearby.

CONCLUSIONSNP chips can efficiently identify tiny CNVs (< 1.0 Mb). In combination with real-time fluorescence quantitative PCR, this may provide valuable information for prenatal diagnosis.

Adult ; Asian Continental Ancestry Group ; genetics ; China ; Chromosome Duplication ; DNA Copy Number Variations ; DNA Polymerase beta ; genetics ; Homeodomain Proteins ; genetics ; Humans ; Limb Deformities, Congenital ; genetics ; Male ; Polymorphism, Single Nucleotide ; Transcription Factors ; genetics ; beta-Transducin Repeat-Containing Proteins ; genetics

An F-box protein, beta-TrCP recognizes substrate proteins and destabilizes them through ubiquitin-dependent proteolysis. It regulates the stability of diverse proteins and functions as either a tumor suppressor or an oncogene. Although the regulation by beta-TrCP has been widely studied, the regulation of beta-TrCP itself is not well understood yet. In this study, we found that the level of beta-TrCP1 is downregulated by various protein kinase inhibitors in triple-negative breast cancer (TNBC) cells. A PI3K/mTOR inhibitor PI-103 reduced the level of beta-TrCP1 in a wide range of TNBC cells in a proteasome-dependent manner. Concomitantly, the levels of c-Myc and cyclin E were also downregulated by PI-103. PI-103 reduced the phosphorylation of beta-TrCP1 prior to its degradation. In addition, knockdown of beta-TrCP1 inhibited the proliferation of TNBC cells. We further identified that pharmacological inhibition of mTORC2 was sufficient to reduce the beta-TrCP1 and c-Myc levels. These results suggest that mTORC2 regulates the stability of beta-TrCP1 in TNBC cells and targeting beta-TrCP1 is a potential approach to treat human TNBC.
Cell Line, Tumor ; Cell Proliferation ; Cell Survival/drug effects ; Cyclin E/genetics/metabolism ; Dose-Response Relationship, Drug ; Female ; Furans/pharmacology ; Gene Knockdown Techniques ; Humans ; Models, Biological ; Multiprotein Complexes/antagonists & inhibitors ; Phosphatidylinositol 3-Kinases/*antagonists & inhibitors ; Phosphorylation/drug effects ; Protein Kinase Inhibitors/*pharmacology ; Proteolysis/drug effects ; Proto-Oncogene Proteins c-myc/genetics/metabolism ; Pyridines/pharmacology ; Pyrimidines/pharmacology ; TOR Serine-Threonine Kinases/*antagonists & inhibitors ; Triple Negative Breast Neoplasms/genetics/*metabolism ; beta-Transducin Repeat-Containing Proteins/genetics/*metabolism