Congress Abstracts - Society for Developmental Biology

through its channel activities
Ishikawa, Masaki; Yamada, Yoshihiko (NIH/NIDCR, USA)
Pannexin 3 (Panx3), a new member of the pannexin gap junction family, is expressed in the perichondrium/periosteum and in
osteoblast. Previously, we reported that Panx3 is induced in the transition stage from proliferation and differentiation of
osteoprogenitor cells and promotes osteoblast differentiation through its hemichannel, endoplasmic reticulum (ER) Ca 2+ channel, and
gap junction. Canonical Wnt/ β-catenin signaling is essential for osteoprogenitor cell proliferation. Here we show that Panx3 inhibits
proliferation and promotes cell cycle exit of osteogenic C2C12 cells, primary calvarial cells, and newborn calvaria explants.
Overexpression of Panx3 reduced osteoprogenitor cell proliferation, whereas the inhibition of endogenous Panx3 increased the
proliferation. We found that this inhibition was mediated through reduced Wnt signaling by β-catenin degradation and GSK3β
activation, which was caused by reduced cAMP/PKA signaling through the Panx3 hemichannel. The Panx3 hemichannel also
contributes to the inhibition of proliferation, by reducing cAMP/PKA/CREB signaling which induces the expression of genes such as
cyclinD1 for cell progression. Furthermore, the released ATP promotes PI3K/Akt signaling, which activates the Panx3 ER Ca 2+
channel to increase intracellular Ca 2+ that increases p21 transcription and phosphorylation by promoting Smad signaling through the
calmodulin signaling pathway activated by Panx3 ER Ca 2+ channel. Our results revealed multi-functional roles of Panx3 for
osteoprogenitor proliferation and cell cycle exit. Thus, Panx3 plays a critical role in switching from proliferation to differentiation of
osteoblasts.
Program/Abstract # 439
Identification and expression of novel Wnt signaling-associated protein kinases
Park, Edmond; Shin, Eun-Young (Korea Basic Science Inst., Rep. of Korea); Shin, Ju-Hyun (Chungnam National Univ. Hosp., Rep.
of Korea); Kim, Gun-Hwa (Korea Basic Science Inst., Rep. of Korea)
The Wnt pathway is an evolutionarily conserved signaling network that is critical for mammalian development and adult tissue
maintenance. In addition, aberrant activation of the Wnt signaling is implicated in driving the formation of various human cancers,
particularly those of the digestive tract. Inhibition of aberrant Wnt pathway activity in cancer cell lines efficiently blocks their growth,
highlighting the great potential of therapeutics designed to achieve this in cancer patients. In this study, we try to identify novel
protein kinases that are associated with canonical Wnt signaling pathway by using TOPflash reporter assay system and human protein
kinase (~500 genes) library. As the result, we identified numbers of protein kinases that positively regulate Wnt signaling pathway.
The novel protein kinases would be possible and valuable target for regulating Wnt pathway in cancer and mammalian development.
Program/Abstract # 440
Axin-stimulated Wnt signaling in mouse embryogenesis and intestinal progenitor cells
Parrish, Angela; Mahaffey, James; Anderson, Kathryn (Sloan-Kettering Institute, USA)
The Wnt signaling pathway is essential for embryonic development and adult stem cell maintenance and is misregulated in many
cancer types. Although the pathway has been extensively studied, the mechanism of pathway activation remains controversial. In the
early mouse embryo, Wnt signals specify and maintain the primitive streak (the site of gastrulation) and progenitor cells in the streak
and tail bud. Axin and Axin2 are considered to be negative regulators of the canonical Wnt pathway. However, we previously showed
that a protein-stabilizing Axin2 point mutation (canopus) leads to a decrease of Wnt signaling in the head but an increase in the late
primitive streak, assessed by the expression of the transgenic Wnt reporter Topgal. To confirm elevated canonical Wnt signaling in the
primitive streak, we are examining the intracellular localization of β-catenin in the heads and tails of Axin-stabilized embryos. We
find that limiting Wnt production with small molecules blocks the increase in Wnt signaling caused by Axin stabilization; genetic
experiments to confirm that Axin-dependent pathway activation depends on ligand are in progress. Elevated Wnt signaling has a
critical role in colon cancer through activation of intestinal stem/progenitor fates. We find that adult canopus heterozygotes show an
increase in the number of β-catenin+ cells in small intestinal crypts, suggesting that stabilized Axin2 can also increase Wnt signaling
in intestinal progenitor cells. We conclude that Axin proteins can tissue specifically activate or repress Wnt signaling, and this will be
important for potential use of Axin-stabilizing drugs to treat cancer.
Program/Abstract # 441
Early Endocytic Trafficking in Control of Developmental Signaling
Gerstner, Norman; Zimyanin, Vitaly; Wieffer, Marnix; Zerial, Marino (MPI-CBG, Germany)
During gastrulation, cells simultaneously process different signaling inputs to pattern the early embryonic body axis. Several
conserved signaling pathways, fundamental for morphogenesis and other developmental functions, are regulated by endocytosis.
Ligand-activated receptors are internalized via different entry routes and compartmentalized into early endosomes. We have
previously performed a genome-wide screen on endocytosis (Collinet et al., Nature 2010) and identified several genes that selectively
regulate transport of signaling cargo to different types of endosomes, including early endosomes involved in signal transduction. We
hypothesize that cargo sorting between distinct early endosomes is essential for the precise regulation of developmental signaling. By
interfering with a novel Rab5-effector that regulates the transport of signaling cargo between endosomal compartments, we
investigated the role of endosomal trafficking in modulating signaling strength and specificity during zebrafish gastrulation. Since
Wnt/beta-catenin signaling and D/V patterning were strongly affected upon such perturbation, we focused on the Wnt/beta-catenin
pathway and trafficking of Wnt-receptor complexes. We quantified trafficking and signaling defects at the level of the whole embryo,
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