Molecular Genetics of Limb Development in Drosophila 
expressing a receptor for the wingless signal 
would allow for specification of clusters of cells 
at unique points with respect to the body pattern. 
Distal-less-Dcpcndctit Genes Involved in 
Limb Development 
One approach to identifying genes that act 
downstream of Distal-less in organizing the limb 
involves looking for other genes expressed in the 
embryonic limb primordia. We would expect a 
downstream target gene to depend on Distal-less 
gene function in order to be expressed in the 
limb primordia. We have begun to screen 
through the genome, using a method for visualiz- 
ing the pattern of gene expression rather than for 
destroying the genes by mutation. Although this 
work is at an early stage, we have identified a 
gene, named disconnected, that is expressed in 
the embryonic limb primordia and that depends 
on Distal-less. 
Proximal-Distal Pattern Formation 
As mentioned above, flies that lack Distal-less 
gene activity do not develop limbs. Flies in which 
Distal-less gene activity is merely impaired de- 
velop limbs that are foreshortened along their 
proximal-distal axis. A range of Distal-less muta- 
tions of varying severity have been isolated that 
cause a graded series of defects in limb develop- 
ment. The characteristics of different mutations 
tell us that the amount of activity of the gene is 
somehow important in controlling the range of 
structures that the limb can develop. Distal parts 
of the limb require more Distal-less gene activity 
than proximal parts in order to develop normally. 
These observations suggest that the gene may 
play an important role in organizing the proxi- 
mal-distal axis of the limbs. 
In order to understand how Distal- less exerts 
this function, we have begun to examine the spa- 
tial distribution of the gene product in develop- 
ing limbs of the adult fly. Distal-less RNA is ex- 
pressed in a concentration gradient across the 
developing limb. The distal-most region of the 
limb expresses a high level, intermediate regions 
express lower levels, and proximal regions ex- 
press little or none. These observations are partic- 
ularly intriguing in that we have observed a dif- 
ferential requirement for the activity of the gene 
in different regions of the leg primordium. Distal 
regions require more activity and express more 
transcript. We are trying to define better the char- 
acteristics of this gradient and hope to determine 
whether the observed graded distribution of the 
gene product is contributing meaningful infor- 
mation to the organization of the proximal-distal 
pattern of the developing limb. 
To assess the role of Distal-less in organizing 
pattern in the developing adult limb, we have 
sought genes that might act downstream of Dis- 
tal-less in this process. The Abnormal leg pat- 
tern (Alp) gene was identified as a prospective 
partner on the basis of a specific phenotypic in- 
teraction with Distal-less. The severity of the leg 
defects caused by dominant mutations of Alp is 
sensitive to changes in the amount of Distal-less 
gene activity. During the past year, we have gen- 
erated mutants that eliminate Alp function and 
have cloned and characterized the gene. It is ex- 
pressed in a narrow ring of cells that encircles the 
developing limb. This ring can be taken as a 
marker for a defined cell identity along the proxi- 
mal-distal axis of the leg. Expression of Alp in this 
ring depends on the Distal-less gene. 
We suggest that understanding pattern forma- 
tion in the developing limb of the fly will teach 
us about fundamental mechanisms that may be 
important during development of vertebrate em- 
bryos. The system of pattern formation that we 
are studying differs from early pattern formation 
in the precellular fly embryo in that pattern is 
being organized in a field of cells, rather than in 
an array of nuclei sharing a common cytoplasm. 
Cells in the developing limb must define their 
own identities and communicate this information 
to their neighbors. In this context we are particu- 
larly intrigued by the role that Distal- less plays as 
a pattern organizer at the genetic level and by its 
prospective role as a regulator of gene expression 
at the molecular level. 
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