JCS021931 Supplementary Material

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• Supplemental Figure S1 (Adobe PDF) -

  Fig. S1. Molecular characterization of the SPARC mutation Df(3R)nm136. (A) Genomic organization of H2Av and SPARC genes. A SPARC mutant allele, Df(3R)nm136, was generated by the imprecise excision of a P element resulting in a 1936-bp deletion within the H2Av gene. Another deletion line, 72b, was generated by imprecise excision that does not result in embryonic lethality. Df(3R)Tl-P is a deficiency line used for complementation analysis. (B) Anti-SPARC polyclonal antibodies recognize a 43 kDa protein from wild-type embryonic extracts. SPARC protein is not detected in protein extracts from SPARC-mutant embryos. β-tubulin was used as a loading control.

• Supplemental Figure S2 (Adobe PDF) -

  Fig. S2. Tracheal integrity is compromised in SPARC-mutant embryos. (A) An early ES17 wild-type embryo immunostained with anti-Crumbs antibody to highlight the tracheal system. The dorsal longitudinal trunk (upper arrows) is a transparent tubule that spans the length of the embryo and the ganglionic branches (lower arrows) are connected to the dorsal trunk. (B) The ES17 SPARC-mutant embryo shows a collapsed and looped dorsal trunk (upper arrows), deformed ganglionic branches (lower arrows) and an overall loss of tracheal integrity. (C) An ES17 SPARC RNAi embryo that does not express SPARC in haemocytes (SrpHemo-GAL4/+; UAS-SPARC RNAi/+) shows collapsed dorsal trunks and tangled ganglionic branches. The overall tracheal morphology is less severe than that of SPARC mutant embryos. (D) An ES17 embryo with ubiquitous loss of SPARC expression (da-GAL4/UAS-SPARC RNAi) results in collapsed commissures (anti-DN-Cadherin immunostaining, green) and an elongated, uncondensed ventral cord. (E) Cuticle preparation of a DCg1⁴¹² mutant embryo. Dorsal trunks are collapsed (upper arrow) and lesions are present in the ventral cuticle. (F) An ES17 DCg1²³⁴ homozygous embryo immunostained with anti-Hunchback (green) and anti-Repo (red) antibodies to mark neural and glia nuclei, respectively. The VNC is uncondensed and is kinked. (G) Cuticle preparation of a SPARC mutant embryo expressing a full-length SPARC cDNA by a haemocyte driver (UAS-SPARC/y or +; gcm-GAL4/+; H2Av-GFP, Df(3R)nm136/ H2Av-GFP, Df(3R)nm136). No holes are present in the ventral cuticle. All confocal images are projections. Scale bar, 10 µm.