Congress Abstracts - Society for Developmental Biology

acts as a positive regulator of this complex, participating in its assembly/stability and modulating its activity. We show that Ccdc28b
regulates cilia length in vivo, at least in part, through its interaction with Sin1 and that depletion of Rictor, another core component of
mTORC2, does not result in shortened cilia. Thus, we describe a previously unknown role of SIN1 in cilia biology, which is
independent of its mTORC2-related function. Taken together, our findings implicate CCDC28B in the regulation of mTORC2 and
provide mechanistic insight to understand the role of CCDC28B in the regulation of cilia length uncovering a novel function of the
core mTORC2 component SIN1 in this process.
Program/Abstract # 82
Molecular Mechanism of Gprk2-dependent Smoothened Regulation in Drosophila
Dominic Maier, Shuofei Cheng, David Hipfner (IRCM, Canada)
Hedgehog (Hh) signaling plays a conserved and essential role in regulating development and homeostasis of numerous tissues.
Cytoplasmic signaling is initiated by Smoothened (Smo), a G-protein-coupled receptor (GPCR) family member, whose levels and
activity are regulated by the Hh receptor Patched (Ptc). In response to Hh binding to Ptc, Smo accumulates at the membrane in a
hyper-phosphorylated, active state. cAMP-dependent Protein kinase A (Pka) and Casein kinase I (CkI) are crucial kinases in this step.
However, phosphorylation of Smo by Pka and CkI accounts only for a fraction of total Smo phosphorylation suggesting that other
kinases might also be involved in Smo regulation. G-protein-coupled receptor kinases (GRKs) are known to phosphorylate activated
GPCRs, leading to the termination of G-protein-dependent signaling and receptor internalization. We have shown that Gprk2, a
member of the GRK family in Drosophila melanogaster, promotes Smo phosphorylation, most likely directly. By using two
independent approaches we have mapped multiple Gprk2-dependent phosphorylation sites in the Smo C-terminus. Because Hh target
gene expression is reduced in gprk2 mutant flies, we hypothesized that phosphorylation of Smo by Gprk2 is required for full
activation of Smo and consequently of Hh target genes. Consistent with this, we find that depletion of Gprk2 in Drosophila S2-R+
cells also decreases Hh target gene expression as measured with a transcriptional reporter assay. Interestingly, expression of Smo
constructs in which Gprk2 phosphorylation sites have been mutated to alanine also decreases Hh target gene expression, mimicking
Gprk2 depletion. Based on these findings we propose that complete Smo activation depends on at least three kinases: Pka and CkI as
well as Gprk2.
Program/Abstract # 83
Identification and regulation of adrenocortical stem cells
Ed Laufer, Salma Begum, Alex Goldberg, Alex Paul (Columbia, USA)
The adrenal cortex is an endocrine organ that produces steroid hormones critical for regulating ionic balances and blood volume, and
modulating stress responses. We previously defined both Shh-expressing cells within the zona glomerulosa at the cortical periphery
and Shh-responsive, Gli1+ cells in the overlying capsule mesenchyme as progenitors of all steroidogenic cell types. We have used
long-term genetic lineage tracing techniques to ask whether either population contains bona-fide adrenocortical stem cells. We find
that Shh lineage cells persist throughout life, and continuously contribute to all steroidogenic cell types, while the Gli1 lineage
generates steroidogenic cells only during embryogenesis. Thus only the Shh population contains cells with adrenocortical stem cell
properties. The canonical Wnt pathway has been implicated as a potential regulator of adrenocortical stem cells at the gland
periphery. We find that Shh-expressing cells have elevated levels of beta catenin, and are also marked by a transcriptional reporter of
canonical Wnt signaling. To address the relationship of Wnt signaling and maintenance of the Shh population, we deleted a
conditional beta catenin allele from the cortical population using cre recombinase expressed under the control of promoters expressed
throughout the steroidogenic lineage. If we delete beta catenin prior to the condensation of the gland, then the cortical cells disperse,
and adrenal formation fails. However if we delete beta catenin after gland formation, then the cortex shrinks over a few weeks, with a
progressive outer to inner loss of cortical populations. These data are consistent with canonical Wnt signals helping define the
adrenocortical stem cell niche.
Program/Abstract # 84
Early insights into the morphogenesis of the activated zone for regeneration or repair in the axolotl and in the mouse
Saori L. Haigo (UCSF, USA); Aida Rodrigo-Albors, Akira Tazaki, Elly M. Tanaka (DFG Ctr. for Regenerative Therapies Dresden,
Germany); Jeremy F. Reiter (UCSF, USA)
Despite a recent renaissance of interest in regenerative biology, we still have a limited understanding of basic cellular mechanisms that
underlie tissue repair in animals. Among vertebrates, urodele amphibians have the remarkable ability to regenerate damaged organs,
while mammals show restricted tissue repair with scarring of damaged organs. To begin to elucidate shared and divergent
mechanisms utilized by regenerating or repairing vertebrates, we sought to identify the activated zone of cells surrounding a fullthickness
wound in the mouse skin. Similar to the axolotl spinal cord, which activates a 500 micrometer zone adjacent to the
amputation plane sufficient to reconstruct the lost spinal cord following tail amputation, our preliminary observations suggest that the
mouse skin may have a similar 500 micrometer zone adjacent to the wound that is activated to repair the epidermis following injury.
Moreover, this activated zone maintains this 500 micrometer distance from the wound edge despite changing the size of initial injury.
We also present data on real time imaging of the initial morphogenesis of spinal cord regeneration in the axolotl. Finally, we provide
a comparative perspective on the initial cellular events that underlie regeneration or repair in vertebrates.
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