Supplementary Material for
Altering Glypican-1 levels modulates canonical Wnt signaling during
trigeminal placode development
Celia E. Shiau1,2, Na Hu1, and Marianne Bronner-Fraser1,*
1. Division of Biology 139-74, California Institute of Technology
Pasadena, CA 91125, USA
2. Present address: Department of Developmental Biology, Stanford University School
of Medicine, Stanford, CA 94305, USA
* Author for correspondence
Supplementary Figure 1
Measurement of ganglia sizes shows dramatic reduction after expression of full-length
but not mutant forms of GPC1.
(A) Graph showing the area of trigeminal ganglia is not significantly different between control
and mutant GPC1 electroporated embryos through the different stages of analysis (stages 15 –
19). Separate plots were made for FL-GPC1 electroporated ganglia also shown in Fig. 2 that
were either normally or reduced sized in order to show the dramatic area reduction in “reduced”
phenotype cases. Bars show s.e.m. (B) Histogram showing frequency of the two types of
phenotypes detected over all analyzed stages 15–19: “reduction” means overall ganglion size is
reduced and “disorganized” means aberrant integration of placodal cells but with generally
normal ganglion size. GPC1 over-expression by FL-GPC1 caused a significant percentage of
reduced ganglia (accounting for both “reduced” and “severely reduced” phenotypes as shown in
Fig. 2), while GPC1 inhibition by GPC1-GPI led to a high percentage of disorganized ganglia.
The reduced ganglia phenotype is largely not detected with expression of any mutant GPC1
construct.
Supplementary Fig. 1
Supplementary Figure 2
Subcellular localization of FL-GPC1 and GPC1-GPI proteins by myc-tag detection.
Chick embryos transfected by in ovo electroporation targeting the trigeminal placodal tissue with
either the cytopcig-FL-GPC1-6xmyc or cytopcig- GPC1-GPI-6xmyc construct were selected for
high cytoplasmic GFP reporter expression and collected for section analysis at stage 14.
Sections through the trigeminal region show (A-D) GFP positive transfected ectodermal and
placodal cells expressing FL-GPC1-6xmyc protein as shown by the myc tag in red. (B-D)
Higher magnification of the dotted box region in A showing expression of FL-GPC1 myc fusion
protein localized to both the cytoplasm and the cell membrane (arrows). (E-H) By contrast,
GPC1-GPI-6xmyc protein expression is more highly concentrated at or near the placodal cell
membrane than FL-GPC1 (arrows), and in some cells is undetectable in the cytoplasm (top
arrow).
Supplementary Fig. 2