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Transcription activates repressed domains in the Drosophila bithorax complex

Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School, Boston, MA 02115, USA

View larger version (11K): [in a new window]   Fig. 1. Map of the central region of the bithorax complex, showing the insertions associated with the Uab mutations. The horizontal line diagrams the DNA sequence, marked in kb according to the published sequence coordinates (Martin et al., 1995). The horizontal bars marked A-J above the sequence line indicate restriction fragments used to prepare probes for RNA in situ hybridization. Below the sequence line, the two previously characterized transcription units in the region are shown. The two triangles at the top left indicate the P element insertions from which the Uab alleles were derived. The bent arrows above the triangles indicate the direction of transcription in these elements. The triangles below the sequence line indicate the insertions in various Uab alleles. The G1 and L1 alleles are head-to-head duplications of the starting elements, the M1 and R1 alleles are inversions of the starting elements, and the HH1 allele is an inversion with an internal deletion.

View larger version (89K): [in a new window]   Fig. 2. Segmental transformations of two Uab alleles. For each genotype, a dorsal view of the abdominal tergites is shown on the left, and a close-up of the ventral sternites is on the right. The cuticle of the second abdominal segment (A2) is marked with a bracket in each panel. All panels show female cuticles, except that the dorsal picture of UabHH1 shows a male. Wild type: the tergite of the first abdominal segment is narrower than in the second abdominal segment, and it lacks the band of pigment and large bristles at the posterior margin. On the ventral side, the first abdominal segment lacks a sternite. The sternite of the second abdominal segment has fewer bristles than that of the third abdominal segment, and these bristles all point directly posterior. The wild-type second abdominal sternite also includes a clear anterior patch, called the Wheeler's organ (marked W.O.). UabR1 homozygote: the A1 tergite is enlarged, and has pigmentation and bristles like that of the A2 segment. There is a clear sternite in the A1 segment, although it is not completely transformed to a more posterior type. UabHH1 homozygote. These animals rarely eclose as adults. The A1 tergite appears fully transformed to the character of the A2 or A3 segment. This individual also shows an extra half tergite anterior to A1. This tergite also has pigment and large bristles typical of the A2 tergite. The A1 sternite resembles that of A3. Note that the A2 sternite also resembles that of A3, as judged by bristle number and orientation, and by the lack of a Wheeler's organ.

View larger version (104K): [in a new window]   Fig. 3. RNA in situ hybridization to Uab embryos. (A) An embryo from UabHH1/MKRS parents, hybridized with a probe from fragment B to detect distally oriented transcripts. The embryo is at stage 11 (6 hours old), and is dissected to show the whole epidermis in one plane. The anterior end is at the top; the anterior limit of RNA expression is in PS6, as marked. (B) Same as A, except the embryo is stage 13 (10 hours old). (C) Same as A, except that the probe is from fragment I, again detecting distally oriented transcripts. The weak signals represent rare transcripts that have extended 50 kb from the UabHH1 insertion. (D) Same as A, except that the embryo was from UabR1/MKRS parents. There is less readthrough transcription than in UabHH1, presumably because of the intact poly(A) addition signals in the UabR1 insertion. Wild-type embryos show no distally directed RNA with probes from either the B or the I fragments.

View larger version (82K): [in a new window]   Fig. 4. Suppression of the Uab transformation by P cytotype. UabHH1/TM1 female. The transformation of the A1 tergite is dramatic, but not as complete as in the homozygote, as shown in Fig. 2. UabHH1/Harwich female. The A1 tergite is restored to the wild-type character by the presence of the Harwich chromosomes, which confer the P cytotype.

View larger version (48K): [in a new window]   Fig. 5. RNA transcripts from rearrangement Uab alleles. (A) A map of the BX-C showing the positions of the Uab rearrangement breakpoints. Fragment B (red box) was used to make a strand-specific probe to detect rightward transcripts (directed towards abd-A). (B) Stage 13 embryo from Uab1/apXa parents. There is abnormal transcription in PS6-12, plus transcription in the posterior end of the embryo, probably representing PS14. (C) Stage 13 embryo from T(1:3) Uab5/FM6xFM6 parents. Ectopic RNA appears throughout the central nervous system, and in lateral clusters of cells (probably in the peripheral nervous system) in the first through seventh abdominal segments.

View larger version (156K): [in a new window]   Fig. 6. UabHH1 homozygous embryo at stage 14 (11 hours) stained with antibody to ABD-A protein. The arrow marks a cell in the ventral nerve chord in PS6 which is expressing ABD-A. At most only a few cells in PS6 show ABD-A misexpression by late embryogenesis. ABD-A is reduced in the epidermis of PS7 (flanked by brackets), along the border between the first and second abdominal segments.

View larger version (51K): [in a new window]   Fig. 7. Ventral cuticles of third instar larvae. The denticle bands from the first and second abdominal segments are shown, with a wild-type larva on the left and a UabHH1 homozygous larva on the right. The segmental transformations are subtle. In the mutant, the A1 denticle band has slightly more and larger denticles than wild type, while the A2 denticle band has slightly fewer denticles.

View larger version (29K): [in a new window]   Fig. 8. Model for the segmental transformations of UabHH1. The ventral cuticle of a wild-type larva is illustrated on the left, with the positions of segments and parasegments marked. A map of the BX-C is drawn along the bottom, showing the nine regulatory regions responsible for the development of parasegments 5-13. The segmental domains of the BX-C have different structures in different parasegments, progressing from all repressed (black ovals) in PS4 to all active (gray ellipses) in PS13. In UabHH1, new transcripts (arrows) appear in PS6-PS12, and these transcripts span the iab-2 and iab-3 regions. Transcription in the mutant animals switches these regulatory regions to the active state (hatched ovals), and their activity transforms the morphology of PS6 and PS7 to that of PS8.

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