Congress Abstracts - Society for Developmental Biology

Program/Abstract # 16
“Paracrine endodermal signaling promotes tracheal cartilage development”
Debora Sinner, John Snowball, Richard Lang, Jeff Whitsett (Cincinnati Children's Med. Ctr., USA)
Tracheo-bronchomalacia is a condition in which the walls of the upper conducting airways are soft because of the lack of cartilage.
While the prevalence of the disease is high (1:6000/live births), the etiology of tracheomalacia is unknown. Our goal is to understand
how the trachea is patterned during normal development and in congenital airway malacia. We generated a mouse model where Wls,
which mediates Wnt ligand secretion, was deleted from endoderm of the developing respiratory tract using ShhCre mice. In these
embryos, the tracheal mesenchyme was mispatterned: Sox9 expression was not detected while the αSMA expression pattern was
expanded into the ventral region of the trachea. Cartilage was near absent however no tracheoesophageal fistula was present. In
contrast, expression pattern of conducting airway markers was unaltered in tracheal epithelium. We hypothesize that Wls modulates
Sox9 expression in tracheal mesenchyme. We crossed the Sox9 conditional KO mice to the mesenchymal driver Dermo1Cre and
obtained mutants in which the cartilage and tracheal length was reduced, recapitulating in part endodermal deletion of Wls as well as
Wnt5a null phenotype. Deletion of Wls in tracheal mesenchyme via Dermo1Cre or in chondroblast using Col2a1Cre, had not effect in
cartilage formation, suggesting that Wls via a paracrine mechanism acts upstream of Col2a1 to induce cartilage. In vivo activation of
canonical Wnt signaling did not rescue the lack of cartilage in embryos were Wls was deleted from pulmonary endoderm. We
conclude that paracrine signaling from the respiratory endoderm modulated by Wls is required for Sox9 expression and patterning of
the tracheal mesenchyme. Partially supported by NIH-NHLBI 1K01HL115447-01 to DS.
Program/Abstract # 17
TALE-mediated modulation of transcriptional enhancers
Justin Crocker, David Stern (HHMI Janelia, USA)
In eukaryotes, the pattern and level of gene expression is modulated by regions of DNA, called transcriptional enhancers, which
encode binding sites for transcription factors. The majority of animal genomes may encode transcriptional enhancers, but we currently
understand far less about the structure and function of enhancer regions. Our understanding of enhancer structure and function is
derived mainly from reporter gene assays. However, these assays often fail to correctly recapitulate native expression patterns. Here
we show that Transcription Activator-Like Effectors (TALEs) allow targeted repression and activation of enhancers in the Drosophila
genome. TALE-repressors (TALERs) targeted against the promoter of the even-skipped gene (eve) caused repression of native eve
expression in multiple cell types. Ubiquitously expressed TALERs with DNA binding specificity for each of the five “stripe”
transcriptional enhancers of eve generated repression specifically of the focal enhancer, without disrupting the activity of neighboring
enhancers. Ubiquitously expressed TALE-activators (TALEAs) targeted against individual enhancers caused increased expression
specifically in cells normally activated by the focal enhancer. Phenotypic effects of these manipulations in embryos and adults are
consistent with the modulations of eve expression caused by TALERs and TALEAs. TALEs thus provide a novel tool for detection
and functional modulation of transcriptional enhancers in their native genomic context. Our results strongly support a model for
combinatorial activation of independent, modular enhancers, and the view that repression acts in a dominant fashion to make silent
enhancers inaccessible to transcriptional activators.
Program/Abstract # 18
Reunion after more than 700 million years of separation: fish transcription factors meet sea urchin DNA
Jongmin Nam (Rutgers, USA)
Commonality in gene regulatory systems is a key aspect of eukaryotic development, as it has been manifested by many successful
trans-specific complementation of transcription factors. As transcription factors require specific DNA sequences to function properly,
it stands to reason that their target sequences are also conserved. This assumption has been supported by numerous comparative
studies between closely related species and has also served as a milestone for comparative genomics. However, there exists a
knowledge gap in the deep evolution of transcription factors and cis-regulatory sequences: numerous transcription factors between
different phyla share common sequence features and function, but the same cannot be said for cis-regulatory DNA sequences. This
raises an interesting question about the prevalence of inter-phyletic cis-regulatory conservation. As a first step towards addressing this,
we asked how many cis-regulatory modules (CRMs) that are active during early embryogenesis of the purple sea urchin are also active
in the zebrafish embryo. Our results show that a high fraction of sea urchin CRMs, 58/109, promoted reporter expression in early
zebrafish embryogenesis. When the same set of CRMs was tested in a more closely related species, the pencil sea urchin, a higher
fraction of CRMs, 74/109, was active, reflecting a closer evolutionary relationship between the two urchins than to zebrafish. Our
experimental and statistical analyses suggest that the majority of these inter-phyletic or inter-generic regulatory interactions are due to
conserved regulatory interactions rather than opportunistic interactions. Thus, inter-phyletic functional conservation beyond
"detectable" sequence conservation may be abundant.
Program/Abstract # 19
The weak shall lead the strong: The role of transcription factor binding affinity in morphogen gradient responses and
enhancer evolution
Scott Barolo (U Michigan, USA)
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