Abstracts - Society for Developmental Biology

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patterning. Signaling pathways involved in these processes include Wnt, Nodal and Bmp. We found that loss of zygotic
Zimp7 with a morpholino results in dorsalization of embryos and upregulation of markers of the organizer and mesoderm
(including boz and squint). In contrast overexpression of zimp7 by mRNA injection, leads to repression of these same
genes and causes axial mesoderm defects resembling previously described mutants that affect the organizer such as
ichabod and boz. Loss of dorsal expression of the no tail gene and the presence of cyclopia in the gain-of-function situation
suggest that Zimp7 could interact with the Nodal-related pathway.
Program/Abstract # 453
Direct visualization of retinoic acid gradients in zebrafish embryos
Sosnik, Julian; Gratton, Enrico; Schilling, Thomas, University of California Irvine, United States;
Retinoic acid (RA) is a derivative of vitamin A that plays important signaling roles in development, regeneration and
disease. Because it is not synthesized de novo, tracking and determining its endogenous localization has been elusive. The
only data available for the distribution of RA are indirect, based on the localization of proteins involved in its synthesis,
transport, signaling and degradation, or based on reporters that indicate RA responses. To address this we have taken a
novel approach using Fluorescent Lifetime Image Microscopy (FLIM) in combination with a phasor approach to data
analysis. Due to RA’s unique lifetime of fluorescence, this methodology allows us to make direct observations in vivo of
its endogenous distribution in developing embryos. FLIM analysis of zebrafish embryos during gastrulation and
somitogenesis revealed higher levels of RA in somites where it is synthesized by Aldh1a2, and levels were reduced in
Aldh1a2 mutants. In addition, we observed graded reductions both anteriorly, across the presumptive hindbrain, as well as
posteriorly in the presomitic mesoderm. These observations are consistent with previous models in which RA acts as a
graded morphogen both in hindbrain and somite segmentation. These measurements strongly suggest that the graded
responses previously observed using transgenic reporters form at the level of the RA ligand itself. Taken together, these
data constitute some of the first direct observations of the endogenous distribution of RA in vivo in the developing embryo.
Present work includes the use of FLIM in the analysis of the crosstalk between RA and other signaling pathways and their
effects in modulating the spatial and temporal distribution of RA.
Program/Abstract # 454
The role for proteoglycans in regulating early embryonic patterning and morphogenesis
Superina, Simone; Ciruna, Brian, Hospital for Sick Children, Toronto, Canada
Growth factors and morphogens regulate embryonic patterning, cell fate specification, cell migration, and morphogenesis.
The activity and behaviour of these signalling molecules are regulated in the extracellular space through interactions with
proteoglycans. Proteoglycans are high molecular-weight proteins consisting of a core protein with covalently linked
glycosaminoglycan (GAG) sidechains, which are thought to mediate the proteoglycan-ligand interaction. UDP-glucose
dehydrogenase is an enzyme required for the synthesis of GAGs. Through germline replacement, I have generated a
maternal zygotic Ugdh mutant (MZ jek) to determine the requirement for proteoglycans in the developing embryo. In
accordance with mouse and Drosophila ugdh mutants, Fgf signalling is reduced in MZ jek. Intriguingly, MZ jek possess
left-right patterning abnormalities, characterized by bilateral lefty2 expression in the lateral plate mesoderm. The midline
appears to be intact in the mutant embryos, when examined by insitu hybridization for shh , no tail , and lefty1 gene
expression. Kupffer's Vesicle formation and ciliogenesis proceed normally in MZ jek , generating the necessary
stereotypical counterclockwise flow required for symmetry breaking. Examination of calcium signalling in the wildtype
embryos reveals unilateral phosphorylation of CamKII on the left side of the KV, a kinase that becomes activated in
response to increases in intracellular calcium levels. Interestingly, in mutant embryos P-CamKII is found on both sides of
the KV, possibly representing bilateral calcium influxes and subsequent improper Nodal pathway activation.
Program/Abstract # 455
The role of HSPG in anterior-posterior axis formation.
Yamamoto, Masamichi, Gunma University, maebashi-city, Gunma, Japan
Nodal and Bmp signaling play a central role in establishing the anterior-posterior axis of embryo. These growth factors act
spatially and temporally in embryo. Heparan sulfate proteoglycans modulate activity of growth factors such as TGF-β,
Nodal, Bmp, Fgf, Wnt and Shh during various developmental stage by controlling extracellular stabilization, movement
and retention both on the cell surface and on the extracellular matrix. Here, we investigated the candidate of heparin sulfate
proteoglycans regulating the Nodal and Bmp signal in anterior-posterior axis formation. We found two new candidate
genes in regulating of anterior-posterior axis.