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First published online 4 July 2007
doi: 10.1242/dev.02878

Development 134, 2807-2813 (2007)
Published by The Company of Biologists 2007
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The Drosophila HMG-domain proteins SoxNeuro and Dichaete direct trichome formation via the activation of shavenbaby and the restriction of Wingless pathway activity

Paul M. Overton1,*, William Chia1,2 and Marita Buescher1,2,{dagger}

1 Medical Research Council Centre for Developmental Neurobiology, King's College London, 4th Floor New Hunt's House, Guy's Hospital, London SE1 1UL, UK.
2 Temasek Life Sciences Laboratory, 1 Research Link, National University of Singapore, Singapore 117604, Republic of Singapore.


Figure 1
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  		Fig. 1. Cuticle phenotype and expression pattern of SoxN. (A-B') SoxN mutants have an aberrant cuticle phenotype. (A) Wild type (wt); cuticle of first instar larva. (A') Wild type; single denticle belt. (B) SoxNGA1192 mutant; cuticle of first instar larva. (B') SoxNGA1192 mutant; single denticle belt. Numbers (1-6) represent the six rows of denticles. (C-J) Expression pattern of SoxN and its regulation by signaling pathway activities. (C-E) Wild-type embryos double-stained with anti-SoxN (black) and anti-En (brown). (C) Stage 8; SoxN protein expression is ubiquitous. (D) Stage 9; SoxN expression decays in a narrow stripe just anterior to the En domain. (E) Stage 13; SoxN expression is restricted to segmental stripes, each with a width of six cells. (F) Stage 15 enGal4-UAS-taulacZ embryos double-stained with anti-SoxN (red) and anti-ß-galactosidase (green, detecting En expression). The anterior-most row of SoxN-expressing cells (1) corresponds to the posterior row of En-expressing cells. Arrows, notice that SoxN expression in the two posterior-most rows (5,6) is higher than in the anterior four rows (1-4). (G) Staining of stage-15 wgcx4 mutant embryos with anti-SoxN; notice that SoxN expression is derepressed as compared with wild type. (H) Staining of stage-15 wgcx4 embryos with an svb-specific RNA probe; notice that the expression patterns of SoxN and svb in wgcx4 embryos are highly similar. (I) Staining of stage-15 armVP16Gal4-UAS-EgfrAct embryos with anti-SoxN; notice that SoxN expression expands posteriorly by two to three rows of cells. (J) Staining of stage-15 armVP16Gal4-UAS-EgfrDN embryos with anti-SoxN; notice that SoxN expression is reduced. Cartoon: schematic representation of the ventral epidermis with the expression patterns of the wg, en, svb and SoxN genes. SoxN and svb are co-expressed in six rows of cells, which differentiate to produce denticles. SoxN and svb expression is stimulated by Der- and repressed by Wg-pathway activities.

 

Figure 2
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  		Fig. 2. SoxN is necessary and sufficient to activate svb expression/direct denticle formation. (A,B) svb RNA expression in stage-14 wild-type (wt; A) and SoxNGA1192 (B) embryos. (C) Ectopic denticle formation in an enGal4-UAS-SoxN larva. The arrow indicates ectopic denticle formation in the anterior row of the En-expressing stripe. Rows are labeled 1-6. (D,E) Misexpression of SoxN with the armVP16Gal4 driver results in ectopic denticle formation (indicated by brackets; D) in those areas where ectopic expression of svb occurs (E); note that svb is expressed in broad stripes but is not ubiquitous. (F) Anti-GFP staining (green) of stage-12 armVP16Gal4-UAS-SoxN-YFP embryos; note that armVP16Gal4 drives SoxN expression in broad stripes but is not ubiquitous. The red staining indicates the ventral midline. (G-I) Misexpression of SoxN with the wgGal4 (wgGal4-UAS-SoxN-YFP) driver results in ectopic denticle formation. (G) Ectopic denticle formation in wgGal4-UAS-SoxN-YFP larvae. (H) Cuticle and anti-YFP staining of stage-17 wgGal4-UAS-SoxN-YFP embryos; note that SoxN directs ectopic denticle formation in a cell-autonomous manner. (I) Cuticle and anti-HA staining of stage-17 wgGal4-UAS-SoxN-UAS-wg embryos; note that concomitant misexpression of SoxN and wg results in ectopic denticle formation in a cell-autonomous manner.

 

Figure 3
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  		Fig. 3. SoxN acts downstream of Der- and Wg-pathway activities to direct denticle formation. (A) Cuticle of a wg null mutant larva. (B) Cuticle phenotype of a wg;SoxNU6-35 double-mutant larva; notice that removal of SoxN function largely rescues the wg mutant phenotype. (C-E) Removal of SoxN function rescues the pan mutant phenotype. (C) pan2 mutant cuticle. (D) pan2;SoxNU6-35/+ cuticle. (E) pan2;SoxNU6-35 cuticle. (F) Cuticle of an scaGal4-UAS-EgfrAct larva; note the ectopic denticles posterior to the wild-type row-6 denticles (indicated by arrows). (G) Cuticle of an scaGal4-UAS-EgfrAct;SoxNGA1192 larva; note that ectopic denticle formation is largely suppressed.

 

Figure 4
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  		Fig. 4. svb function is necessary for the maintenance of epidermal SoxN and Dichaete expression. (A-H) Anti-SoxN stainings (A-D) and anti-Dichaete (anti-D) stainings (E-H) of wild-type (WT) and svb1 mutant embryos. Wild-type (A) and svb1 mutant (B) stage-13 embryos; note that there is no appreciable difference between wild-type and svb1 mutant embryos, indicating that svb is not required for the establishment of the SoxN expression domain. Wild-type (C) and svb1 mutant (D) stage-16 embryos. (D) Notice the severe reduction of SoxN expression, indicating a role for svb in the maintenance of SoxN expression. Wild-type (E) and svb1 mutant (F) stage-13 embryos; notice that there is no appreciable difference between wild-type and svb1 mutant embryos, indicating that svb is not required for the establishment of the Dichaete expression domain. Wild-type (G) and svb1 mutant (H) stage-16 embryos. (H) Notice the reduction in Dichaete expression, indicating a role for svb in the maintenance of Dichaete expression.

 

Figure 5
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  		Fig. 5. Dichaete is co-expressed and co-regulated with SoxN and is required for denticle formation. (A) Double-staining of wild-type (wt) stage-15 embryos with anti-SoxN (green) and anti-Dichaete (anti-D; red); notice the complete overlap between SoxN and Dichaete expression (yellow). (B) Staining of stage-15 wgcx4 mutant embryos with anti-D; notice that Dichaete is ectopically expressed. (C) Staining of stage-15 armVP16Gal4-UAS-EgfrDN embryos with anti-D; notice the reduction in Dichaete expression. (D) Wild-type cuticle. (E) SoxNU6-35 mutant cuticle. (F) SoxNU6-35;D/+ mutant cuticle.

 

Figure 6
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  		Fig. 6. SoxN and Dichaete negatively regulate Wg pathway activity. (A-D) Anti-En stainings of late-stage-10 embryos. Insets show higher magnifications of the En expression around the ventral midline. Black lines indicate the ventral midline. (A) Wild type (wt). (B) armVP16Gal4-UAS-SoxN; notice the reduction in En expression. (C) SoxNU6-35; notice the increase in En expression. (D) SoxNU6-35;D/+; notice that the increase in En expression is stronger than in SoxN single mutants. Cartoon: regulatory interactions between the Der- and Wg-pathway activities, SoxN, Dichaete (denoted by D) and svb. SoxN and Dichaete expression is stimulated by Der- and repressed by Wg-pathway activity. SoxN and Dichaete negatively regulate Wg pathway activity and activate the expression of svb. svb is not required for the establishment of the epidermal SoxN and Dichaete expression domains. However, svb is necessary for the maintenance of SoxN and Dichaete expression during the late stages of embryogenesis.

 




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