Multiple BMP signalling components regulate nuclear growth in SCs.

A. Schematic of the paired male accessory glands (arrows), which pump their contents into the ejaculatory duct (arrowhead) during mating. Left inset shows the epithelial secretory monolayer containing secondary cells (SCs; green) and main cells (MCs), all of which are binucleate. Right inset is a section through the gland revealing the large lumen (asterisk). B. SC (circled) expressing a gene trap for the BMP type I receptor tkv, and stained with an antibody against the BMP type II receptor Wit. These receptors are present both on the plasma membrane (arrowhead) and co-localise in intracellular compartments (arrow). DAPI marks nuclei blue in SCs (asterisks) and surrounding non-expressing MCs. C-F. Accessory glands (AGs) from 6-day-old controls (C) and males expressing RNAis targeting tkv (D) or Mad (F) or expressing the BMP signalling antagonist Dad (E) in adult SCs under the control of esgF/O^ts after temperature shift at eclosion. AGs were dissected, fixed and imaged by confocal microscopy. Glands were stained with DAPI (blue) to mark nuclei and an anti-Fas3 antibody (yellow) to mark cell boundaries. SCs express nuclear GFP, which is also present in the cytosol. Pairs of nuclei from binucleate SCs and MCs are indicated by green and red arrows respectively. G. Bar chart showing SC nuclear size relative to that of MC neighbours for glands expressing different transgenes in SCs under esgF/O^ts control, normalized to the ratio for controls. Note that all manipulations that decrease BMP signalling significantly reduce SC nuclear size. Genotypes for images are: w; PBac[544.SVS-1]tkv^CPTI002487 (B); w; esg-GAL4 tub-GAL80^ts UAS-FLP; UAS-GFP_nls actin>FRT>CD2>FRT>GAL4 combined with no other transgene (C); P[TRiP.JF01485]attP2 (III) (D); P[w⁺ UAS-Dad] (II) (E); P[TRiP.JF01263]attP2 (III) (F). ***P<0.001, Kruskal-Wallis test, n = 10. Scale bar for (B) is 10 μm, for all other images it is 20 μm.