CELLULAR INTERACTIONS IN EMBRYOGENESIS 
RoELAND NussE, Ph.D., Associatc Investigator 
Research in the laboratory of Dr. Nusse is focused 
on the function of growth factors in early embryo- 
genesis, in particular on a group of genes whose 
original discovery was related to their oncogenic 
properties. The prototype member of this group, 
Wnt l, is an oncogene in mouse mammary cancer; 
normal expression of this gene is limited to early 
development. Wnt-1 is part of a large family of re- 
lated genes. All members of the mouse Wnt family 
encode secreted proteins closely related to the Dro- 
sophila segment polarity gene wingless. Dr. Nusse's 
group studies the function of this family of genes in 
both mammals and Drosophila. 
Expression of Mouse Wnt Genes During 
Embryogenesis 
Dr. Patricia Salinas, a fellow from the Leukemia 
Society, has further analyzed the expression of 
Wnt- 3 in the developing brain. In 9. 5 -day mouse 
embryos, Wnt-3 is expressed in a restricted area of 
the diencephalon before any morphological signs of 
segmentation appear. Around embryonic day 11.5, 
Wnt-3 expression becomes restricted to the D2 
neuromere (dorsal thalamus) . A mouse homologue 
of the Drosophila Distal-less gene is expressed in a 
nonoverlapping area immediately anterior to and 
abutting the VTnf-J-expressing domain, corre- 
sponding to the Dl compartment (ventral thala- 
mus). In addition, Wnt-3 is expressed in the mid- 
brain-hindbrain region. In the adult mouse, Wnt-3 is 
expressed in subsets of neural cells derived from the 
original areas of expression in the diencephalon. 
These results suggest that Wnt-3 provides positional 
information for the formation of segmental bound- 
aries and defines the regional identity of compart- 
ments within the central nervous system (CNS) . The 
continued expression of Wnt-3 in the adult mouse 
brain suggests a distinct role in the mature CNS. 
In the brain of adult mice, Wnt-3 expression is 
found in the dorsal thalamus. In addition, the gene is 
highly expressed in Purkinje cells of the cerebel- 
lum, which are located immediately adjacent to the 
granular layer. In the cerebellum from mice mutant 
at the weaver locus, where the migration of granular 
cells is disturbed, Wnt-3 expression in the Purkinje 
cells decays prematurely. 
The wingless Gene Family in Drosophila 
The segment polarity gene wingless is thought to 
interact with a group of genes with a similar pheno- 
type to control cell differentiation within individual 
segments of Drosophila embryos. Dr. Nusse's labo- 
ratory is particularly interested in the interaction of 
wingless with other segment polarity genes, one of 
which may encode the wingless receptor. At least 
one other segment polarity gene, engrailed, is 
under the control of wingless expression, and other 
segmentation genes are thought to mediate this 
control. 
One interest of the laboratory is to establish func- 
tional assays for wingless by rescuing mutants with 
wild-type DNA constructs and to do structure-func- 
tion analysis with site-directed mutants. Derek Less- 
ing, a graduate student, has recently established the 
position of the 5' end of the wingless transcriptional 
unit and has sequenced 2,500 base pairs of the 
wingless promoter. Marcel van den Heuvel and 
Cynthia Harryman have characterized the molecular 
nature of several existing wingless alleles. In all of 
the embryonic viable alleles analyzed, the encoded 
protein is unaltered. Embryonic lethal alleles, in 
contrast, have all undergone mutations in the pro- 
tein-encoding domain of the gene, including dele- 
tions, point mutations of conserved residues, and 
P-element inserts. A temperature-sensitive mutation 
was shown to be a substitution of a conserved cys- 
teine for a serine residue. Immunostaining of mu- 
tant embryos shows that the embryonic lethal alleles 
lead to either no wingless antigen or to an appar- 
ently nonsecreted form of the protein. Transfection 
of the mutant alleles into tissue culture cells fol- 
lowed by immunoprecipitation or immunostaining 
analysis supported these results. Wild-type protein 
is found on the cell surface or in the extracellular 
matrix, whereas mutant protein is retained in the 
endoplasmic reticulum (ER). 
Dr. Lee Fradkin has generated epitope-tagged 
wingless \2iv'\mts to examine protein secretion in S2 
cells and to purify the wingless protein from either 
eukaryotic or prokaryotic cells. In collaboration 
with Dr. Norbert Perrimon (HHMI, Harvard Medical 
School), Marcel van den Heuvel has examined the 
distribution of the wingless protein in embryos mu- 
tant for other segment polarity genes to identify 
gene products interacting with the wingless pro- 
tein. The results suggest that the porcupine gene 
product plays a role in the secretion of the wingless 
protein. Further molecular analysis of these genes 
and their products is in progress. 
To examine the consequences of ectopic wingless 
activity, Jasprien Noordermeer made transgenic flies 
that express wingless under the control of an hsp 70 
CELL BIOLOGY AND REGULATION 97 
