pap encodes the fly homolog of the human transcriptional mediator subunit TRAP240. (a) Molecular organization of the pap/dTRAP240 region (cytological position 78A1-3, chromosome 3L). The position of the recessive lethal P-lacW element insertion allele pap¹ is shown with the map of this region. (E) EcoRI; (B) BamHI; (S) SalI. The pap⁵³ allele deletes the entire coding portion of exon 1 (amino acids 1–23). The transcription unit is shown with boxes representing exons; ATG is the presumptive initiator codon, filled boxes indicate protein coding region, and broken lines indicate introns (the precise site of transcription initiation is not known). (b) Protein sequence comparisons of TRAP240 homologs from Drosophila (d), human (h), and Caenorhabditis elegans (c). Because of the length of the proteins, only the most conserved regions (the N and C termini) are shown. Identical amino acids are shaded in black and conservative changes in gray. The Genbank accession numbers for the fly and human proteins are AF227214 and AF117754, respectively. The putative C. elegans homolog is derived from EMBL accession number Q93442, with aligned sequence beginning at a conserved methionine.

Expression and activity of the pap/dTRAP240 locus. (a,b,d,e) Localization of PAP in imaginal discs from third-instar larvae using a monospecific rabbit polyclonal serum. Shown are a prothoracic leg imaginal disc (a) with enlargement (b), and a wing imaginal disc (d) with enlargement (e). PAP protein is expressed in most or all imaginal cells (a,d) and accumulates preferentially in the nucleus (b,e). Mutant clones at the distal extremity of the second tarsal segment in the T1 legs (c) exhibited an ectopic distal sex comb (dsc). Ectopic sex comb teeth (small circles) are yellow (pap⁻). A maximum of three ectopic teeth were observed in any pap⁻ clone. Within the normally placed sex comb (sc) at the distal end of the first tarsal segment of this sample, three mutant cells (marked with circles) are interspersed with pap⁺ cells. This sex comb is of normal size. Hence, the pap⁻ condition affected neither sex comb cell differentiation nor rotation for alignment. (f) Twin spot clonal analysis of pap function in imaginal disc development. Adjacent (twin) clones of pap^−/− (no GFP; black arrow) and pap^+/+ cells (two copies of ubiquitin–GFP; white arrow) are identified by expression of the cell-autonomous GFP marker in a third-instar larval wing imaginal disc. Note the similar sizes and distributions of the wild-type and mutant cells.

Identification and genomic localization of Drosophila melanogaster homologs to known human MED genes. These include dTRAP80, an apparent metazoan homolog of yeast SRB4. (a) Cytological positions of 21 predicted dMED loci are shown. These genes have been annotated by the Genome Annotation Database of Drosophila (GadFly; http://www.fruitfly.org/annot/) and are listed in Materials and Methods. Homology searches identified dTRAP80 as the apparent metazoan homolog of yeast SRB4 (see c). (b) Genomic organization of the dTRAP80 locus (cytological interval 90F1-2 of chromosome 3R) is shown. The dTRAP80 region map, (E) EcoRI; (N) NotI; (K) KpnI; (Bg) BglII, indicates the position of the recessive lethal dTRAP80¹ P-element insertion, previously referred to as l(3)s2956 (Spradling et al. 1999). The exon–intron organization is shown beneath with filled boxes for protein coding region and broken lines indicating introns. (c) Aligned protein sequences of TRAP80/Srb4 homologs from D. melanogaster (d), human (h), Schizosaccharomyces pombe (sp), and Saccharomyces cerevisiae (sc). Genbank accession numbers are AF244916, AF117657, CAB10081, and L12026, respectively. A Caenorhabditis elegans TRAP80 homolog was also detected (not shown; Genbank accession number CAB76740). Identical amino acids are shaded in black, and conservative changes (Taylor 1986) in gray. Positions of aligned amino acids predicted to adopt helical (H) or extended (E) secondary conformations (http://www.ibcp.fr/predict.html) are indicated above the alignment.

pap and dTRAP80 functions contribute to segmental identity specification. Labial palp identity is determined by the joint action of Scr and pb. (a,c) Loss-of-function mutations of pap/dTRAP240 and dTRAP80 affect labial palp development, favoring T1 leg development. (a) The mutant combination pb⁴/pb⁵ (where pb⁵ is a protein-null allele and pb⁴ is a hypomorph leading to a C-terminally truncated PB protein) transforms distal labial palps to antennal aristae (arista), whereas the proximal labium retains normal pseudotracheal rows (pst). (c) The pap¹ and dTRAP80¹ mutations are recessive in this tissue but enhance the pb^4/pb⁵ mutant phenotype. Shown are mouthparts of a pb⁴/pap¹ pb⁵ dTRAP80¹ adult female. Morphology is altered compared to a; pseudotracheae are replaced by apparent leg structures, and claws replace the aristae. (b,d) Scr acts with pb in the labium to promote labial (or antennal/labial) identity but transforms the labium to distal T1 leg in the absence of pb. Accumulation of SCR protein was examined in imaginal labial disc cells of pb⁴/pb⁵ mutants, giving rise to mixed antennal/labial identities (b) or pb⁴/pap¹ pb⁵ dTRAP80¹ mutants that give predominantly T1 leg tissue (d). Nuclear SCR protein was detected throughout the discs in both cases, as it was for wild type. SCR accumulation is not increased and, indeed, may be somewhat reduced (cf. b and d), in the discs giving rise to T1 legs. (e–h) Scr expression examined in clones of pap⁵³ homozygous cells. (e) Shown is a labial imaginal disc harboring a twin spot clone of +/+ (white, vertical arrow) or of pap⁵³/pap⁵³ cells (black, diagonal arrow). (f) Expression of the SCR homeodomain protein is seen in the same labial disc. The clone is outlined and indicated by the diagonal arrow. SCR accumulation is unaltered in the mutant cells. (g) This T1 leg imaginal disc harbors a prominent clone of pap⁵³/pap⁵³ cells (black, limit indicated by arrow). (h) SCR protein accumulation in the same T1 disc. The clone is outlined and its border indicated by the arrow. SCR accumulation is unchanged in the pap⁻ cells.