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Fig. 2. PtdIns(4,5)P₂ depletion inhibits sperm tail formation and causes defects in microtubule organization. (A-D) Phase-contrast micrographs of whole Drosophila testes. (A) A wild-type testis has many rope-like cysts of elongating spermatids (arrow). (B) Expression of SigD causes the accumulation of ovoid spermatid cysts (arrow). (C) Co-expression of Sktl with SigD restores spermatid elongation (arrow). (D) Expression of SigD^dead has no effect on sperm tail formation (arrow). (E-I) Confocal fluorescence micrographs of spermatids expressing β-tubulin-GFP (β-tub-GFP, green) and stained for DNA (blue). (E,H) Wild-type spermatids. (E) β-tub-GFP is incorporated into elongating axonemes and perinuclear forked microtubule (MT) arrays (arrow). (H) After individualization, β-tub-GFP is found in prominent perinuclear puncta (arrows) and axonemes (inset, magnified cyst). (F,G,I) SigD-expressing spermatids. (F) Early elongating cyst showing β-tub-GFP in axonemes (arrowhead) and large diffuse perinuclear MT arrays (arrow). (G) Cysts with tangled tails of β-tub-GFP (arrowhead). (I) Later-stage cysts show poorly aligned MTs (arrowhead) and puncta dissociated from nuclei (arrows) (inset, magnified cyst). Scale bars: 50 µm (A-D), 20 µm (E-I).

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