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First published online 30 November 2005
doi: 10.1242/dev.02173

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Requirement of abdominal-A and Abdominal-B in the developing genitalia of Drosophila breaks the posterior downregulation rule

David Foronda^*, Beatriz Estrada^*,, Luis de Navas and Ernesto Sánchez-Herrero

Centro de Biología Molecular Severo Ochoa (C.S.I.C.-U.A.M.), Universidad Autónoma de Madrid, Cantoblanco, 28049 Madrid, Spain.

View larger version (72K): [in a new window]   Fig. 1. Expression of Abd-B and cad in the embryonic genital disc. Anterior is to the top. (A-C) Double staining of a late stage 15 hdc-lacZ embryo with anti-ß-galactosidase (green, A, note the three clusters) and anti-Abd-B (red, B) antibodies. A merged image is shown in C. Note that the posterior cluster of genital disc cells is not marked with Abd-B (arrow). (D-F) Double staining of a stage 15 cad-Gal4/UAS-GFP; hdc-lacZ/+ embryo marked with anti-ß-galactosidase (red, D) and GFP (green, E). A merged image is shown in F. See that the posterior cluster (arrow) shows cad expression. (G-L) Embryonic genital disc cells of a UAS-myc-EGFPF/+; Abd-B-Gal4LDN/hdc-lacZ embryo, showing expression corresponding to the Abd-B m transcript (GFP expression, green, I-L) in the anterior lateral cells of the disc primordium, marked with an anti-ß-galactosidase antibody (blue, G,J,L). GFP-marked cells are a subset of those expressing Abd-B (red, H,K,L). Note that about four anterior-lateral cells of the disc primordium express GFP (arrows, J) and that some Abd-B-expressing cells are not marked with GFP (arrows, K). A merged image is shown in L. vc, ventral cord.

View larger version (90K): [in a new window]   Fig. 2. Abd-B is needed for the formation of the embryonic genital disc. Anterior is to the top. (A) Late stage 15 hdc-lacZ embryo showing the three clusters of cells that will form the genital disc. (B) Stage 15 Abd-BM5 hdc-lacZ homozygous embryo. The embryonic genital disc is formed by fewer cells in the mutant than in the wild type. (C) Similar stage embryo of the genotype Abd-BUab1/Abd-BM1 hdc-lacZ. The genital disc is disorganized and includes fewer cells than the wild type does. (D) Stage 15 Abd-BM1 hdc-lacZ homozygous embryo. Some ß-galactosidase-expressing cells are scattered (out of focus) and only the posterior cluster remains (arrow). (E,F) In Abd-BM1 hdc-lacZ homozygous embryos, Dll is ectopically activated in some posterior cells (arrows, E; red, F); hdc is marked in green in F. vc, ventral cord.

View larger version (114K): [in a new window]   Fig. 3. Expression and requirement of abd-A in the female internal genitalia. (A) Third instar female genital disc, stained with an anti-abd-A antibody. (B) Similar disc of an abd-A-lacZ larva stained with X-Gal. The pattern of expression reproduces that of the anti-abd-A antibody. (C,D) X-Gal staining of internal female genitalia from two late pupae. The seminal receptacle (sr), spermathecae (s), uterus (u) and oviducts (ov) are stained, whereas parovaria (p) are not. o, ovaries. (E) Female genital disc of an abd-Aiab3-277/DfR59 female larva stained with anti-abd-A antibody, showing a reduction in abd-A expression compared with wild type. (F) Female internal genitalia of an abd-Aiab3-277/Df 109 female. See that most elements of the internal genitalia and the ovaries have disappeared (compare with C). vp, vaginal plates (part of the external genitalia).

View larger version (43K): [in a new window]   Fig. 4. Abd-B m and Abd-B r expression in mature genital discs. (A) Scheme representing the Abd-B transcription unit (not drawn to scale). The RNA is not represented. Red or blue rectangles represent exons of Abd-B m (red) or of Abd-B r (blue) transcripts, and black boxes represent coding regions. (B,C) Drawings of the female (B) and male (C) genital discs (ventral views), indicating the A8 and A9 primordia. The A10 primordium is on the opposite side in both discs. (D,F) Female discs hybridized with probes detecting the Abd-B m transcript (D) or the Abd-B r RNAs (`common' probe; F). (E,G) Male discs hybridized with Abd-B m (E) and Abd-B r (G) probes. Arrowheads in D and F indicate the A9, and in E and G, the A8 segment. Note that the Abd-B m transcript is expressed in the A8 of female and male discs, with some weak expression in the male A9, and that the Abd-B r transcript is present in the male and female A9, with some signal also in the female A8 (arrows; the most central expression in this primordium, out of focus, is the signal from the dorsal A9). Note also the low or absent Abd-B r expression in the anterior female A9.

View larger version (86K): [in a new window]   Fig. 5. Abd-B m function in the genitalia and the female genital disc. (A) An Abd-BM5 mutant clone in the female right eighth hemi-tergite, marked with yellow, is converted into an anterior tergite (arrow). Compare with the left, wild-type, hemi-tergite (arrowhead). (B) Similar mutant clone showing transformation of the genitalia into a sternite (arrow). S7, seventh sternite. (C) Abd-BM3 clone, marked with yellow, showing transformation of the male A8 tergite (normally very small) into an anterior one (arrow). T7, seventh tergite; g, genitalia. (D-F) Abd-BM5 clones in the A8 of the female disc, marked by the absence of the lacZ marker (green, D), do not activate Dll (red, E; arrowheads in E and H mark the wild-type Dll expression). a, analia. The merged image is shown in F. (G-L) abd-AM1 mutant clones in the female A8, marked by the absence of the ß-galactosidase marker (G,J, green), do not eliminate Dll (H, red) or Abd-B (K, red) expression. Merged images are shown in I and L. (M-O) Abd-BM5 clones in the A8 of the female genital disc, marked by the absence of the lacZ marker (in green in M), do not change abd-A expression (red, N). A merged image is shown in O. (P-R) Abd-BD18 mutant clones in the A8 segment, marked by the absence of the ß-galactosidase marker (blue, P,R; clones are outlined), eliminate abd-A expression, detected by an anti-abd-A antibody (green, Q,R), and activate Dll protein expression (red, P,R). (S) Abd-BD18 abd-A+ clones, marked with GFP (in green), eliminate Abd-B (blue) and activate Dll (red) expression. The boxed clone is shown in detail in T-W. (X) Similar clones induced in the leg disc do not eliminate Dll, as shown by the co-expression of the GFP marker (green) and Dll (red), giving a yellow color (arrowheads). (Y) dpp-Gal4/UAS-Dll female disc showing elimination of abd-A expression in the dpp domain (arrowheads).

View larger version (68K): [in a new window]   Fig. 6. Abd-B expression in Abd-B m mutant embryos and clones, and Abd-B m expression and function in the female internal genitalia. (A) Wild-type stage 11 embryo showing Abd-B expression, detected with the lA2E9 monoclonal antibody, in PS12-PS14 (A7-A9). (B) Abd-BM5 homozygous embryo at a similar stage, stained with the same antibody. The Abd-B expression is restricted to PS14. (C-E) Abd-BM5 clones induced in female A8 do not eliminate Abd-B expression. Clones are marked by the absence of GFP marker (green, C), and Abd-B expression is detected with the anti-rabbit anti-Abd-B (red, D). A merged image is shown in E. (F) Abd-BM5 clones induced in the same primordium, marked by the absence of X-gal staining (light blue), do not ectopically activate Abd-B r expression (purple). (G,H) Detail of the regions boxed in F. Note the Abd-B r wild-type expression in the female A9 (arrowheads) and its absence, or very low expression, in the clones (arrows). (I) Female internal genitalia of an UAS-lacZ/+; Abd-B-Gal4LDN/+ late pupa. The spermathecae (s) and uterus (u) are stained, whereas parovaria (p) and oviducts (ov) are not. o, ovaries; vp, vaginal plates. (J) Detail of I, showing absence of staining in the parovaria. (K) Abd-B m mutant female showing disappearance of all the external and internal genitalia except the parovaria (p). h, hindgut; a, analia.

View larger version (74K): [in a new window]   Fig. 7. The Abd-B r function is required for the development of the male genitalia. (A-D) In the male genitalia, some Abd-BUab1 clones transform into leg tissue (A, boxed region is amplified in B; arrows point to bracted bristles) or into antenna (C, boxed region is amplified in D), ar, malformed arista; III, third antennal segment. (E-G) Some Abd-BUab1 clones in the A9 of the male disc, marked by the absence of the lacZ marker (red, E) eliminate Abd-B expression (green, F). Merged image in G. (H-J) Male disc bearing similar clones, marked by the absence of GFP (green, H), eliminate Abd-B expression (I, blue), and in some cells activate Dll (I,J, red, arrow). In some clones there is no Abd-B expression but no Dll activation either (arrowheads, I). (K) Drawing of a male genital disc showing the distribution of Abd-BUab1 clones that eliminate Abd-B expression (pink) and those that do not (green). The region where most `pink' clones accumulate includes, mainly, the presumptive penis apparatus domain. (L) Internal genitalia of an Abd-BUab1/Abd-Bx23-1 female. Note the presence of three spermathecae (s) in the highly abnormal internal genitalia. vp, vaginal plates.

View larger version (18K): [in a new window]   Fig. 8. Comparison of genetic interactions in embryonic epidermis and in the mature female genital disc, and organization of the A8 in the female genital disc. (A) Regulatory interactions between abd-A, Abd-B and Dll in the embryonic epidermis and in the mature genital discs. In the embryo, Abd-B represses abd-A in the A8, and abd-A represses Dll in the abdominal segments. In the third instar female genital disc, it seems that Abd-B maintains abd-A transcription, and that ectopic Dll can repress abd-A. (B) In the A8 of the female disc we can distinguish two regions (Estrada and Sánchez-Herrero, 2001) (this report). The anterior region (region I, purple) expresses buttonhead, abd-A and low levels of Abd-B, and can activate Dll in the absence of Abd-B. It represents the `appendage' part of the genitalia and corresponds mainly or exclusively to the internal genitalia. The lower region (region II) does not express abd-A or buttonhead, but expresses high levels of Abd-B. It corresponds to the `trunk' region of the female A8. In this region, Dll is not activated in the absence of Abd-B.