The standard model of Wnt signaling specifies that after receipt of a Wnt ligand at the membranous receptor complex, downstream mediators inhibit a cytoplasmic destruction complex, allowing β-catenin to accumulate in the cytosol and nucleus and co-activate Wnt target genes. Unexpectedly, shortly after Wnt treatment, we detected the dephosphorylated form of β-catenin at the plasma membrane, where it displayed a discontinuous punctate labeling. This pool of β-catenin could only be detected in E-cadherin-/- cells, because in E-cadherin+/+ cells Wnt-induced, membranous β-catenin was concealed by a constitutive junctional pool. Wnt-signaling-dependent dephosphorylated β-catenin colocalized at the plasma membrane with two members of the destruction complex - APC and axin - and the activated Wnt co-receptor LRP6. β-catenin induced through the Wnt receptor complex was significantly more competent transcriptionally than overexpressed β-catenin, both in cultured cells and in early Xenopus embryos. Our data reveal a new step in the processing of the Wnt...]]>
Cortical compartments partition proteins and membrane at the cell surface to define regions of specialized function. Here we ask how cortical compartments arise along the plasma membrane furrows that cellularize the early Drosophila embryo, and investigate the influence that this compartmentalization has on furrow ingression. We find that the zygotic gene product Nullo aids the establishment of discrete cortical compartments, called furrow canals, which form at the tip of incipient furrows. Upon nullo loss-of-function, proteins that are normally restricted to adjacent lateral regions of the furrow, such as Neurotactin and Discs large, spread into the furrow canals. At the same time, cortical components that should concentrate in furrow canals, such as Myosin 2 (Zipper) and Anillin (Scraps), are missing from some furrows. Depletion of these cortical components from the furrow canal compartments precipitates furrow regression. Contrary to previous models, we find that furrow compartmentalization does not require cell-cell junctions that...]]>
Securin is a chaperone protein with bifunctional properties. It binds to separase to inhibit premature sister chromatid separation until the onset of anaphase, and it also takes part in cell-cycle arrest after UV irradiation. At metaphase-to-anaphase transition, securin is targeted for proteasomal destruction by the anaphase-promoting complex or cyclosome (APC/C), allowing activation of separase. However, although securin is reported to undergo proteasome-dependent degradation after UV irradiation, the ubiquitin ligase responsible for securin ubiquitylation has not been well characterized. In this study, we show that UV radiation induced a marked reduction of securin in both the nucleus and cytoplasm. Moreover, we show that GSK-3β inhibitors prevent securin degradation, and that CUL1 and βTrCP are involved in this depletion. We also confirmed that SKP1-CUL1-βTrCP (SCFβTrCP) ubiquitylates securin in vivo, and identified a conserved and unconventional βTrCP recognition motif (DDAYPE) in the securin primary amino acid sequence of humans, nonhuman primates and rodents....]]>
The development of luminal organs begins with the formation of spherical cysts composed of a single layer of epithelial cells. Using a model three-dimensional cell culture, this study examines the role of a cytoskeletal motor, myosin II, in cyst formation. Caco-2 and SK-CO15 intestinal epithelial cells were embedded into Matrigel, and myosin II was inhibited by blebbistatin or siRNA-mediated knockdown. Whereas control cells formed spherical cysts with a smooth surface, inhibition of myosin II induced the outgrowth of F-actin-rich surface protrusions. The development of these protrusions was abrogated after inhibition of F-actin polymerization or of phospholipase C (PLC) activity, as well as after overexpression of a dominant-negative ADF/cofilin. Surface protrusions were enriched in microtubules and their formation was prevented by microtubule depolymerization. Myosin II inhibition caused a loss of peripheral F-actin bundles and a submembranous extension of cortical microtubules. Our findings suggest that inhibition of myosin II eliminates the cortical...]]>