eprintid: 16546
rev_number: 15
eprint_status: archive
userid: 1026
dir: disk0/00/01/65/46
datestamp: 2014-03-28 12:39:30
lastmod: 2014-03-31 07:27:41
status_changed: 2014-03-28 12:39:30
type: doctoralThesis
metadata_visibility: show
creators_name: Kumar, Sumit 
title: Molecular Dissection of Mouse Wnt3a-Frizzled8 Interaction Reveals Essential and Modulatory Determinants of Wnt Signaling Activity
subjects: 500
divisions: 140001
adv_faculty: af-14
keywords: English
abstract: 
Wnt/β-catenin signaling plays a critical role in animal development and adult tissue 
homeostasis, including regulation of cell fate decisions, axial pattering, organogenesis
and stem cell maintenance. Deregulation of Wnt signaling causes many human
diseases, including cancer and osteoporosis. Wnt proteins constitute a large family of
growth factors characterized by conserved cysteine and a lipid modification at their
N-terminal half. Upon transport through a specialized secretary route they act as
morphogens in variety of tissues by forming concentration gradient in the
extracellular space. The post-translational addition of palmitate or palmitoic acid to
Wnts renders them highly hydrophobic and is believed to control their membrane
localization, it is unknown, however, how Wnts are recruited from the membrane to
signaling complexes and how their signaling range is regulated.
Wnts bind to Frizzled (Fz) seven transmembrane receptors and LRP5/6 co-receptors,
which leads to stabilization of the transcription coactivation of β-catenin and
activation of target gene. The structural basis of Wnt signaling by receptor binding
has long been elusive. Only recently, the crystal structure of the Xenopus Wnt8-
Frizzled8-CRD complex was solved, but the significance of the interaction sites for
signaling has not been assessed. In the present thesis, by using a structure-based
mutagenesis approach, I present an extensive structure-function analysis of mouse
Wnt3a by in vitro and in vivo evaluation. Evidence is provided for an essential role of
Serine 209, Glycine 210 (site 1) and Tryptophan 333 (site 2) in Fz binding.
Importantly, I discovered that Valine 337 in the site 2 binding loop is critical for
signaling without contributing to binding. Mutations in the presumptive second CRDbinding
site (site 3) partly abolished Wnt binding, suggesting Fz dimerization as a
necessary step in signaling. Intriguingly, most site 3 mutations increased Wnt
signaling, probably by inhibiting Wnt-CRD oligomerization. In accordance, it was
observed that increasing amounts of soluble Fz8-CRD protein modulated Wnt3a
signaling in a biphasic manner. Based on these finding, a model was developed, in
which a concentration-dependent switch in Wnt-CRD complex formation from an
inactive aggregation state to an activated, high mobility state, represents a modulatory
mechanism in Wnt signaling gradients.
date: 2014
publisher: University of Heidelberg
id_scheme: DOI
id_number: 10.11588/heidok.00016546
ppn_swb: 1654265829
own_urn: urn:nbn:de:bsz:16-heidok-165460
date_accepted: 2014-01-13
advisor: HASH(0x564e1c39ec80)
language: eng
bibsort: KUMARSUMITMOLECULARD2014
full_text_status: public
place_of_pub: Heidelberg, Germany
citation:   Kumar, Sumit   (2014) Molecular Dissection of Mouse Wnt3a-Frizzled8 Interaction Reveals Essential and Modulatory Determinants of Wnt Signaling Activity.  [Dissertation]     
document_url: https://archiv.ub.uni-heidelberg.de/volltextserver/16546/1/PhD%20thesis-Sumit%20Kumar.pdf