Molecular Genetics of Limb Development 
in Drosophila 
Stephen M. Cohen, Ph.D. — Assistant Investigator 
Dr. Cohen is also Assistant Professor in the Department of Cell Biology and in the Institute for Molecular 
Genetics at Baylor College of Medicine. He obtained his Ph.D. degree at Princeton University, working 
with Malcolm Steinberg. He did postdoctoral work with Harvey Lodish at the Whitehead Institute 
and with Herbert Jackie at the Max Planck Institute for Developmental Biology, Tiibingen, 
and at the University of Munich. 
MY laboratory is interested in the processes 
by which genetic information can be used 
to organize the three-dimensional pattern of the 
body during embryonic development. The partic- 
ular problem on which we have focused involves 
the organization of the limbs in the fruit fly Dro- 
sophila. We would like to understand the genetic 
and molecular mechanisms that the fly embryo 
uses to determine where the limbs will develop 
with respect to the rest of the body pattern. We 
are investigating how the appropriate embryonic 
cells become committed to develop as limb pre- 
cursors and subsequently how these cells cooper- 
atively organize the spatial pattern of the leg. 
Specification of Limb Cell Identity 
Using classical genetics, we have identified the 
gene Distal-less, which seems to function as a 
critical genetic switch that initiates limb develop- 
ment in the embryo. Embryos that lack Distal- less 
gene function do not develop larval or adult 
limbs. These observations indicate that Distal- 
less activity is critically required but do not tell 
us what the gene is actually doing. Using molecu- 
lar probes, we can visualize the RNA product of 
the Distal-less gene in the embryo and in the de- 
veloping adult limbs. All of the cells that will go 
on to form limb structures express Distal-less. An 
additional genetic experiment indicates that 
these cells need the Distal- less gene product to 
be limb cells. If the gene is removed from a group 
(or clone) of cells that were already committed 
to develop into limb structures, these cells in- 
stead become body wall cells. Distal-less there- 
fore specifies their identity as limb cells. Distal- 
less is a member of a family of important 
regulatory genes that encode a sequence-specific 
DNA-binding motif called the homeodomain. 
Distal-less therefore can be presumed to define 
cell identity by regulating the expression of other 
genes. We wish to identify the transcriptional tar- 
gets through which Distal-less acts. 
How Does the Embryo Position Its Limbs? 
The critical first event in limb development is 
determining the position at which the legs will 
develop. We are using a combination of genetic 
and molecular methods to identify the source of 
the instructions specifying the identity of the leg 
cells and the nature of the response that this sig- 
nal elicits. In the Drosophila embryo, the legs 
begin as clusters of cells. These clusters are estab- 
lished at a well-defined location within each tho- 
racic segment, with respect to both anterior- 
posterior and dorsal-ventral position. One way to 
mark a unique point is to draw two intersecting 
lines. This rather simple solution appears to de- 
scribe what the embryo does to identify the cell 
clusters. Thus the spatial cues that the embryo 
uses to position its legs appear to be stripes of 
cells that act as sources of secreted intercellular 
signals. The earliest detectable consequence of 
this signaling process is that presumptive leg 
cells express the Distal-less gene. 
Our previous work identified the segmentally 
repeated stripes of cells expressing the wingless 
gene as one of the signaling centers responsible 
for initiating leg development. Stripes of cells ex- 
pressing wingless bisect the nascent leg primor- 
dia. The wingless gene encodes a secreted inter- 
cellular signaling molecule related to the 
vertebrate oncogene INT- 1 . The secreted wing- 
less protein is required to signal nearby cells to 
express Distal- less but is not sufficient to do so. 
An intersecting stripe of information is required 
to specify the cluster. The decapentaplegic gene 
is a good candidate to encode this second signal. 
A stripe of cells expressing decapentaplegic in- 
tersects the wingless stripe at precisely the posi- 
tion where the leg is formed. The decapentaple- 
gic gene encodes a product homologous to the 
transforming growth factor-/3 (TGF-|5) class of se- 
creted signaling molecules. A group of cells that 
lies near the intersection of these two stripes 
should receive both signals. Although we are not 
yet certain whether decapentaplegic plays a di- 
rect role in this process, the simultaneous receipt 
of the wingless and TGF-/3 signals should, in prin- 
ciple, be sufficient to specify the identity of the 
leg precursor cells. 
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