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First published online 23 May 2007
doi: 10.1242/dev.003616

Development 134, 2387-2396 (2007)
Published by The Company of Biologists 2007
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Zfrp8, the Drosophila ortholog of PDCD2, functions in lymph gland development and controls cell proliferation

Svetlana Minakhina, Marina Druzhinina and Ruth Steward*

Waksman Institute, Department of Molecular Biology and Biochemistry, Cancer Institute of New Jersey, Rutgers University, 190 Frelinghuysen Road, Piscataway, NJ 08854-8020, USA.


Figure 1
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  		Fig. 1. The Zfrp8 lymph gland phenotype. (A,B) Wild-type (A) and Df(2R)SM206/Df(2R)SM206 (B) Drosophila lymph glands. Primary lobes are marked with I, secondary lobes with II. (C-F) Confocal crossections of 16- to 17-stage wild-type (C) and a Zfrp8-null (D) embryos stained with anti-Srp antibody (Srp) show the early increase in size of mutant lymph glands. Wild-type (E) and Zfrp8-null (F) larval lymph glands stained with anti-Hemese antibody (He). (G) Hemocyte counts in wild-type and in Zfrp8 and Df(2R)SM206 mutant larvae. Hemocytes were counted from at least ten third instar larvae of each genotype. Scale bars: 70 µm in A,B; 15 µm in C,F.

 

Figure 2
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  		Fig. 2. The mutant lymph gland contains differentiated hemocytes. Confocal images of wild-type (A,C,E) and Df(2R)SM206 (B,D,F) Drosophila lymph glands stained with (A,B) anti-Ser to locate PSCs, (C,D) P1 antibodies as a plasmatocyte marker, and (E,F) L1 antibodies to mark lamellocytes. Scale bars: 150 µm.

 

Figure 3
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  		Fig. 3. Drosophila lymph gland size is dependent on the dose of Zfrp8. Lymph glands from (A) wild-type, (B) Df(2R)SM206, (C) Df(2R)SM206/+, (D) Df(2R)SM206/Arkk11502, (E) Df(2R)SM206/+, pnrMD237/+, (F) Df(2R)SM206, pnrMD237/+, (G) Df(2R)SM206/+, Cdc27L7123/+ and (H) l(1)dd42/+, Df(2R)SM206/+ third instar wandering larvae stained with Hoechst 33258 for DNA. Scale bars: 70 µm.

 

Figure 4
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  		Fig. 4. Cell proliferation in wild-type and mutant Drosophila lymph glands. (A,B) Lymph glands from wild-type (A) and Df(2R)SM206 (B) mid-third instar larvae stained for cells in S phase. Confocal section from the middle of the organ shows BrdU incorporation in cells of primary (I) and secondary (II) lobes. (C-E) Mitotic cells in the lymph glands from wild-type (C), Zfrp8M-1-1 (D) and Df(2R)SM206 (E) mid-third instar larvae. Projection through 10 µm section of the organ shows cells with condensed DNA, stained with anti-H3P antibodies. Arrows point to areas with high mitotic activity. Scale bars: 70 µm.

 

Figure 5
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  		Fig. 5. Distribuiton of CycA and CycB in Zfrp8 mutants. Comparative analysis of CycA and CycB distribution in (A,C,D,G,H) wild-type and (B,E,F,I,J) Df(2R)SM206 Drosophila lymph glands and brains. The combined confocal fields (A,B) show larval brain with adjacent discs and lymph glands (partial) stained with anti-CycB, anti-H3P and Hoechst 33258. The schematics depict the tissues shown in the panels beneath. Br, brain; D, imaginal discs; I,II, primary and secondary lobes, respectively, of lymph glands (LG). High magnifications of brain (C,G,E,I) and lymph gland secondary lobes (D,H,F,J) show differences in CycB (C-F) and CycA (G-J) distribution. (Df(2R)SM206 and Zfrp8M-1-1 give similar results.) Scale bars: 70 µm in A,B; 18 µm in C,J.

 

Figure 6
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  		Fig. 6. Zfrp8 and centrosome organization. Confocal projection through one layer of lymph gland cells from squashed (A,B) and intact (C-F) lymph glands of wild-type (A,C,E) and Zfrp8 mutant (B,D,F) Drosophila. (Df(2R)SM206 and Zfrp8M-1-1 give similar results.) Cells were stained with anti-{gamma}-Tubulin (A-E), anti-Dgrip91 (C,D) and anti-Dgrip84 (F) to visualize centrosomes. Confocal sections of lymph glands (E,F) show Zfrp8-HA adjacent to centrosmes (arrows). DNA is stained with Hoechst 33258. Scale bar: 5 µm.

 

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© The Company of Biologists Ltd 2007