The Genetic Control of Morphogenesis 
tron about 1 5 kb downstream of the transcription 
start site. These analyses of Scr are supported by a 
grant from the National Institutes of Health. 
The location of these three elements relative to 
the identified enhancer elements is intriguing. 
Two flank the enhancers, while the third is lo- 
cated in the midst of the regulatory elements. Our 
current hypothesis is that the transvection ele- 
=^-ments serve to define a chromatin domain con- 
trolling an on/off state for the locus and that the 
resident enhancers can only have their effects in 
the on state. We are currently extending our anal- 
ysis to define more precisely the boundaries and 
sequences associated with the above-identified 
fragments. We also hope to identify further the 
cellular factors that act through these elements. 
The labial Gene 
Using enhancer sniffers, we have identified a 
majority of the sequences needed for the normal 
expression of the lab gene. These are found up- 
stream, either within 3 .6 kb of the transcription 
start site or within the major 1 4-kb intron. Using a 
minigene construct that contains all of the up- 
stream sequences but lacks the intronic ele- 
ments, we have succeeded in rescuing the lethal- 
ity and morphological anomalies associated with 
deletion of the lab gene in the embryo. Thus it 
would appear that the regulatory elements found 
within the intron are not necessary to lab's nor- 
mal embryonic functions. 
Although the lab minigene rescues the embry- 
onic defects associated with /aft-deficient geno- 
types, there is no apparent rescue of adults. We 
have shown that this failure does not result from 
the minigene's inability to be expressed in adult 
tissues, but rather because its protein product is 
inappropriately and ectopically accumulated. 
This abnormal expression pattern only takes 
place in lab mutant animals and is not detected in 
normal minigene-bearing hosts. 
We have found that the normal lab gene actu- 
ally encodes two polypeptides, one of which is 
six amino acids longer than the other. The mini- 
gene is only capable of directing the synthesis of 
the shorter protein. It would appear, therefore, 
that animals capable of making the long form 
show correct adult expression, while short-form 
animals cannot, and that normal adult expression 
involves autogenous negative regulation. 
Our earlier studies revealed that lab expres- 
sion in the embryo also involved autogenous regu- 
lation. In that case, however, the effect was posi- 
tive. That is, lab protein served to keep the lab 
gene turned on, not to turn it off. Moreover, the 
short-form protein was capable of performing 
this function. 
We have now demonstrated that the sequences 
at the lab locus necessary for the positive and 
negative loops are, like the required proteins, 
different. The positive sequences are upstream of 
the transcription initiation site, while the nega- 
tive targets are in the transcribed portion of the 
gene in the first exon. We are currently investi- 
gating the possibility that the two loops are af- 
fected by the direct interaction of the lab protein 
isoforms with the two alternate target sites in the 
lab gene itself. 
The proboscipedia Gene 
As in the case of lab, we have used a minigene 
derived from the pb locus and have obtained full 
rescue of the adult homeotic mutant phenotype. 
Moreover, fragments extracted from one of the 
introns and placed upstream of the pb promoter 
function to direct a normal spatiotemporal pat- 
tern of expression. Thus sequences both up- and 
downstream of the transcription start site are re- 
quired for normal pb expression. 
However, tests of the "enhancer" sequences 
on a heterologous promoter element in the 
sniffer constructs demonstrate no activity. On the 
other hand, minigene constructs in which the en- 
hancer elements are deleted can be directed by 
novel "enhancer" elements derived from other 
loci. Therefore it appears that there are specific 
promoter/enhancer interactions at this locus and 
that the specificity appears to lie primarily with 
the "enhancer" elements. 
We have also found a second regulatory ele- 
ment at the pb locus upstream of the transcrip- 
tion start site. However, this element, unlike the 
positive enhancer above, appears to be negative 
and serves to prevent ectopic expression of the 
gene. Moreover, like the Scr locus, it seems to 
have a pairing-sensitive component. We are now 
beginning a dissection of this system, which 
offers an opportunity to make direct comparisons 
between two similar regulatory schemes in the 
ANT-C. Our work on the proboscipedia gene is 
supported by a grant from the National Institutes 
of Health. 
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