06-P034

Determining the developmental basis of idiopathic clubfoot

Elizabeth Kilby, Neil Vargesson

University of Aberdeen, Aberdeen, United Kingdom

Idiopathic Congenital Talipes Equinovarus, or clubfoot, is a

common developmental disorder of the foot, affecting at least 2

in every 1000 live births in Scotland. The defect is characterised

by a twisting of the foot and loss of calf muscle. Diagnosis is usually

made on postnatal examination and treatment; usually a series of

foot manipulations, takes place within the first year. Treatment can

be particularly painful and is not always successful as the defect

can recur leading to life-long disability in some cases. Very little

is known of the aetiology of clubfoot despite it being such a com-

mon problem and therefore our lab set out to elucidate the develop-

mental processes that occur during limb development which result

in clubfoot. The chick limb is a well established model system used

to study many developmental processes, signalling events and has

been used to model several human limb disorders. We have devel-

oped a chick model of human clubfoot using a neuromuscular

paralysing agent, resulting in a range of clubfoot conditions. Our

lab is currently using this chick model to determine the develop-

mental basis of the disorder. Initial phenotypic analyses of the

chick clubfoot limb correlate well with those observed in human

clubfoot; both exhibit muscle loss, tendon problems and bony

abnormalities. We will use this chick model of human clubfoot to

test our hypothesis that clubfoot may result from a failure of hind-

limb rotation and elucidate the developmental and molecular basis

of this congenital defect.

doi:10.1016/j.mod.2009.06.260

06-P035

Additional PEA3 binding sites in the long range limb regulator

disrupt posterior restriction of SHH causing polydactyly

Iain Williamson, Laura Lettice, John Wiltshire, Alison Hill,

Carlo DeAngelis, Paul Devenney, Robert Hill

MRC Human Genetics Unit, Edinburgh, United Kingdom

The congenital abnormality preaxial polydactyly (PPD) is

caused by the ectopic expression of the signalling protein Sonic

Hedgehog (SHH) in the developing limb. Targets of FGF signalling,

Pea3/Erm, have been shown to play a vital role in restricting Shh

to the posterior limb bud. The ZRS, a cis-regulator of Shh expres-

sion, has several conserved sub-domains containing binding

motifs recognized by PEA3. Single nucleotide mutations in the

ZRS allow the enhancer to induce Shh in the anterior as well as

posterior limb bud. Point mutations recently identified in two

unrelated human families with PPD generate additional PEA3

binding motifs. We report a direct interaction of PEA3 with the

ZRS and demonstrate that the addition of a PEA3 binding site is

sufficient to generate a loss of posterior restriction and ectopic

expression of Shh in the anterior mesenchyme. These data reveal

a direct interplay between SHH and FGF signalling.

doi:10.1016/j.mod.2009.06.261

06-P036

Dissecting the embryonic requirement of the Notch pathway

gene, Hes1, in the context of DiGeorge syndrome

Irinna Papangeli1, Kelly Lammerts Van Bueren1, Kerra Pearce1,

Catherine Roberts1, Dorota Szumska2, Shoumo Bhattacharya2,

Peter J. Scambler1

1Institute of Child Health, London, United Kingdom2University of Oxford, Oxford, United Kingdom

DiGeorge syndrome is the most common microdeletion syn-

drome in humans frequently caused by a 3 Mb deletion of chro-

mosome 22. The phenotype is characterized by craniofacial,

cardiovascular, thymus, parathyroid and psychiatric defects.

TBX1 localises within the commonly deleted region on 22q11

and is considered the major genetic determinant of the syn-

drome. Deletion of Tbx1 in animal models results in pharyngeal

and cardiovascular defects, highlighting its role in the develop-

ment of these structures. We have determined potential Tbx1 tar-

get genes through microarray screens, including Hes1, an effector

of the Notch pathway. Hes1 is a bHLH factor acting as a transcrip-

tional repressor regulating multiple processes. Homozygous

knockdown of the gene results in a range of defects and is embry-

onic lethal. We have shown that Hes1 is expressed throughout the

pharyngeal apparatus including the Tbx1 expression domains

and is downregulated in Tbx1/ compared to stage matched wild-

type embryos. Additionally, Hes1 null embryos exhibit craniofa-

cial and cardiovascular anomalies, partially recapitulating the

Tbx1 null phenotype. Conditional mutagenesis of Hes1 in the neu-

ral crest results in moderate thymic defects, while concurrent

deletion in the ectoderm and crest populations induces signifi-

cant thymic defects. We are currently investigating cell autono-

mous pathways that may be affecting cell proliferation or

apoptosis in the pharyngeal apparatus.

doi:10.1016/j.mod.2009.06.262

06-P037

Dubowitz syndrome maps to chromosome 12 within an interval

of 70 genes

Jing Tian1, Mohammad Shboul1, Barry Merriman2,

Hane Lee2, Hanan Hamamy3, Bruno Reversade1

1Institute of Medical Biology, A*STAR, Singapore, Singapore2Department of Human Genetics, David Geffen School of Medicine,

University of California, Los Angeles, Los Angeles, CA, United States3National Center for Diabetes, Endocrinology and Genetics, Amman,

Jordan

Dubowitz syndrome (DS) is a rare genetic disorder character-

ized by microcephaly, facial anomalies, eczema, high-pitched

hoarse voice, growth retardation and primordial short stature.

DS is found in less than 200 people around the world. While some

cases are sporadic most have an autosomal recessive mode of

inheritance. Identical-by-descent homozygosity mapping per-

formed on a DS family consisting of three affected girls with

and two unaffected siblings identified a candidate region of

22.9 Mb on chromosome 12. By re-sequencing this entire locus

consisting of 71 genes, we aim to identify the causative allele,

S130 M E C H A N I S M S O F D E V E L O P M E N T 1 2 6 ( 2 0 0 9 ) S 1 2 0 – S 1 3 6