beta-catenin accumulates in the nucleus and binds
to the transcription factor, TCF/Lef, to activate Wnt target gene
transcription. In order for beta-catenin to bind TCF/Lef, however, the
transcriptional corepressor Groucho (Gro)/TLE, which is bound to TCF/Lef,
must be displaced. Work by Hanson et al. shows that X-linked Inhibitor of
Apoptosis (XIAP), an E3 ubiquitin ligase, is also recruited to TCF/Lef upon
Wnt signaling. XIAP ubiquitylates Groucho/TLE, thereby decreasing its
affinity for TCF/Lef and allowing for unfettered access of beta-catenin to
TCF/Lef. The figure illustrates a safety lock cover (Gro/TLE) that is used
to prevent accidental/unauthorized activation of a critical process (Wnt
signaling). Unlocking and removal of the cover (via ubiquitylation by XIAP)
allows the switch (TCF/Lef) to be activated (by beta-catenin).
The artist is
Wolfgang Wozniak, Creative Director of firstname.lastname@example.org
The enteric nervous system (ENS) arises from the coordinated migration, expansion and differentiation of vagal and sacral neural crest progenitor cells. During development, vagal neural crest cells enter the foregut and migrate in a rostro-to-caudal direction, colonizing the entire gastrointestinal tract and generating the majority of the ENS. Sacral neural crest contributes to a subset of enteric ganglia in the hindgut, colonizing the colon in a caudal-to-rostral wave. During this process, enteric neural crest-derived progenitors (ENPs) self-renew and begin expressing markers of neural and glial lineages as they populate the intestine. Our earlier work demonstrated that the transcription factor Foxd3 is required early in neural crest-derived progenitors for self-renewal, multipotency and establishment of multiple neural crest-derived cells and structures including the ENS. Here, we describe Foxd3 expression within the fetal and postnatal intestine: Foxd3 was strongly expressed in ENPs as they colonize the gastrointestinal tract and was progressively restricted to enteric glial cells. Using a novel Ednrb-iCre transgene to delete Foxd3 after vagal neural crest cells migrate into the midgut, we demonstrated a late temporal requirement for Foxd3 during ENS development. Lineage labeling of Ednrb-iCre expressing cells in Foxd3 mutant embryos revealed a reduction of ENPs throughout the gut and loss of Ednrb-iCre lineage cells in the distal colon. Although mutant mice were viable, defects in patterning and distribution of ENPs were associated with reduced proliferation and severe reduction of glial cells derived from the Ednrb-iCre lineage. Analyses of ENS-lineage and differentiation in mutant embryos suggested activation of a compensatory population of Foxd3-positive ENPs that did not express the Ednrb-iCre transgene. Our findings highlight the crucial roles played by Foxd3 during ENS development including progenitor proliferation, neural patterning, and glial differentiation and may help delineate distinct molecular programs controlling vagal versus sacral neural crest development.
Enterocyte Microvillus-Derived Vesicles Detoxify Bacterial Products and Regulate Epithelial-Microbial Interactions
David A. Shifrin, Jr., Russell E. McConnell, Rajalakshmi Nambiar, James N. Higginbotham, Robert J. Coffey, and Matthew J. Tyska
Current Biology; April 10, 2012
The continuous monolayer of intestinal epithelial cells (IECs) lining the gut lumen functions as the site of nutrient absorption and as a physical barrier to prevent the transloca- tion of microbes and associated toxic compounds into the peripheral vasculature . IECs also express host defense proteins such as intestinal alkaline phosphatase (IAP), which detoxify bacterial products and prevent intestinal inflammation [2–5]. Our laboratory recently showed that IAP is enriched on vesicles that are released from the tips of IEC microvilli and accumulate in the intestinal lumen [6, 7]. Here, we show that these native ‘‘lumenal vesicles’’ (LVs) (1) contain catalytically active IAP that can dephos- phorylate lipopolysaccharide (LPS), (2) cluster on the surface of native lumenal bacteria, (3) prevent the adherence of enteropathogenic E. coli (EPEC) to epithelial monolayers, and (4) limit bacterial population growth. We also find that IECs upregulate LV production in response to EPEC and other Gram-negative pathogens. Together, these results suggest that microvillar vesicle shedding represents a novel mechanism for distributing host defense machinery into the intestinal lumen and that microvillus-derived LVs modulate epithelial-microbial interactions.
Enteropathogenic E. coli intimately attached to the surface of Caco2BBE intestinal epithelial cells (Image selected for cover of Current Biology)
Immunofluorescence of Enteropathogenic E. coli intimately attached to the surface of HT-29 intestinal epithelial cells reveal actin pedestal formation and enrichment of intestinal alkaline phosphatase at sites of bacterial attachment. DNA (bacteria and HT-29) marked with DAPI in blue, IAP in green, actin marked with phalloidin in red.
Assembly of an integral Golgi complex is driven by microtubule (MT)-dependent transport. Conversely, the Golgi itself functions as an unconventional MT-organizing center (MTOC). This raises the question of whether Golgi assembly requires centrosomal MTs or can be self-organized, relying on its own MTOC activity. The computational model presented here predicts that each MT population is capable of gathering Golgi stacks but not of establishing Golgi complex integrity or polarity. In contrast, the concerted effort of two MT populations would assemble an integral, polarized Golgi complex. Indeed, while laser ablation of the centrosome did not alter already-formed Golgi complexes, acentrosomal cells fail to reassemble an integral complex upon nocodazole washout. Moreover, polarity of post-Golgi trafficking was compromised under these conditions, leading to strong deficiency in polarized cell migration. Our data indicate that centrosomal MTs complement Golgi self-organization for proper Golgi assembly and motile-cell polarization.
In humans, GLE1 is mutated in lethal congenital contracture syndrome 1 (LCCS1) leading to prenatal death of all affected fetuses. Although the molecular roles of Gle1 in nuclear mRNA export and translation have been documented, no animal models for this disease have been reported. To elucidate the function of Gle1 in vertebrate development, we used the zebrafish (Danio rerio) model system. gle1 mRNA is maternally deposited and widely expressed. Altering Gle1 using an insertional mutant or antisense morpholinos results in multiple defects, including immobility, small eyes, diminished pharyngeal arches, curved body axis, edema, underdeveloped intestine and cell death in the central nervous system. These phenotypes parallel those observed in LCCS1 human fetuses. Gle1 depletion also results in reduction of motoneurons and aberrant arborization of motor axons. Unexpectedly, the motoneuron deficiency results from apoptosis of neural precursors, not of differentiated motoneurons. Mosaic analyses further indicate that Gle1 activity is required extrinsically in the environment for normal motor axon arborization. Importantly, the zebrafish phenotypes caused by Gle1 deficiency are only rescued by expressing wild-type human GLE1 and not by the disease-linked FinMajor mutant form of GLE1. Together, our studies provide the first functional characterization of Gle1 in vertebrate development and reveal its essential role in actively dividing cells. We propose that defective GLE1 function in human LCCS1 results in both neurogenic and non-neurogenic defects linked to the apoptosis of proliferative organ precursors.
One of the most abundant components of the enterocyte brush border is the actin-based monomeric motor, myosin-1a (Myo1a). Within brush border microvilli, Myo1a carries out a number of critical functions at the interface between membrane and actin cytoskeleton. Proper physiological function of Myo1a depends on its ability to bind to microvillar membrane, an interaction mediated by a C-terminal tail homology 1 (TH1) domain. However, little is known about the mechanistic details of the Myo1a-TH1/membrane interaction. Structure-function analysis of Myo1a-TH1 targeting in epithelial cells revealed that an N-terminal motif conserved among class I myosins, and a C-terminal motif unique to Myo1a-TH1, are both required for steady state microvillar enrichment. Purified Myo1a bound to liposomes composed of phosphatidylserine (PS) and phosphoinositol (4,5) bisphosphate [PI(4,5)P2], with moderate affinity in a charge-dependent manner. Additionally, peptides of the N- and C-terminal regions required for targeting were able to compete with Myo1a for binding to highly charged liposomes in vitro. Single molecule total internal reflection fluorescence (TIRF) microscopy showed that these motifs are also necessary for slowing the membrane detachment rate in cells. Finally, Myo1a-TH1 co-localized with both lactadherin-C2 (a PS-binding protein) and PLCδ1-PH (a PI(4,5)P2-binding protein) in microvilli, but only lactaderin-C2 expression reduced brush border targeting of Myo1a-TH1. Together, our results suggest that Myo1a targeting to microvilli is driven by membrane binding potential that is distributed throughout TH1 rather than localized to a single motif. These data highlight the diversity of mechanisms that enable different class I myosins to target membranes in distinct biological contexts.
"In November 2011, the AAAS Council elected 539 members as Fellows of AAAS. These individuals will be recognized for their contributions to science and technology at the Fellows Forum to be held on 18 February 2012 during the AAAS Annual Meeting in Vancouver, Canada. The new Fellows will receive a certificate and a blue and gold rosette as a symbol of their distinguished accomplishments."
Kathleen Gould and her colleagues are studying genetic mutations in fission yeast to better understand the molecular mechanisms that regulate cell division, particularly cytokinesis, the final event of the cell cycle. Image shows fission yeast cytokinesis mutants with the multiple nuclei stained blue, spindle pole bodies pink, and F-actin patches green. (Courtesy Adam Bohnert.)
Jun-Song Chen1,2, Lucy X. Lu1,2, Melanie D. Ohi2, Kevin M. Creamer3,4, Chauca English2, Janet F. Partridge4, Ryoma Ohi2, and Kathleen L. Gould1,2
1 Howard Hughes Medical Institute &
2 Department of Cell and Developmental Biology, Vanderbilt University School of Medicine, Nashville, TN 37212
3 Integrated Program in Biomedical Sciences, University of Tennessee Health Science Center, Memphis, TN 38163
4 St. Jude Children’s Research Hospital, Memphis, TN 38105
Correspondence to Kathleen L. Gould: email@example.com
Cdk1 controls many aspects of mitotic chromosome behavior and spindle microtubule (MT) dynamics to ensure accurate chromosome segregation. In this paper, we characterize a new kinetochore substrate of fission yeast Cdk1, Nsk1, which promotes proper kinetochore–MT (k-MT) interactions and chromosome movements in a phosphoregulated manner. Cdk1 phosphorylation of Nsk1 antagonizes Nsk1 kinetochore and spindle localization during early mitosis. A nonphosphorylatable Nsk1 mutant binds prematurely to kinetochores and spindle, cementing improper k-MT attachments and leading to high rates of lagging chromosomes that missegregate. Accordingly, cells lacking nsk1 exhibit synthetic growth defects with mutations that disturb MT dynamics and/or kinetochore structure, and lack of proper phosphoregulation leads to even more severe defects. Intriguingly, Nsk1 is stabilized by binding directly to the dynein light chain Dlc1 independently of the dynein motor, and Nsk1–Dlc1 forms chainlike structures in vitro. Our findings establish new roles for Cdk1 and the Nsk1–Dlc1 complex in regulating the k-MT interface and chromosome segregation.
S.C.P., J.D.W., J.E.R., and D.M.M. designed research; S.C.P., J.D.W., and J.E.R. performed research; M.S. and W.W.W. contributed unpublished reagents/analytic tools; S.C.P., J.D.W., J.E.R., and D.M.M. analyzed data; S.C.P. and D.M.M. wrote the paper.
Although transcription factors are known to regulate synaptic plasticity, downstream genes that contribute to neural circuit remodeling are largely undefined. In Caenorhabditis elegans, GABAergic Dorsal D (DD) motor neuron synapses are relocated to new sites during larval development. This remodeling program is blocked in Ventral D (VD) GABAergic motor neurons by the COUP-TF (chicken ovalbumin upstream promoter transcription factor) homolog, UNC-55. We exploited this UNC-55 function to identify downstream synaptic remodeling genes that encode a diverse array of protein types including ion channels, cytoskeletal components, and transcription factors. We show that one of these targets, the Iroquois-like homeodomain protein, IRX-1, functions as a key regulator of remodeling in DD neurons. Our discovery of irx-1 as an unc-55-regulated target defines a transcriptional pathway that orchestrates an intricate synaptic remodeling program. Moreover, the well established roles of these conserved transcription factors in mammalian neural development suggest that a similar cascade may also control synaptic plasticity in more complex nervous systems.
- Received June 22, 2011.
- Revision received August 16, 2011.
- Accepted September 6, 2011.