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Fig. S1. Cv-d structure, mutants and RNAi. (A) Predicted alterations in new EMS-induced cv-d mutants. cv-d1 (Fig. 1E) has an in-frame deletion in exon 5 that is predicted to remove K485-N565 from the Vg domain. Sequencing of cv-d1 RT-PCR products revealed that the mutant also fails to excise the downstream intron 7, despite the lack of any mutations in the splice sites or intron; this is predicted to add 71 erroneous amino acids and a stop codon after N758 within the DUF1943 domain. cv-d13 has a four nucleotide deletion after the codon for T193 that is predicted to add 16 erroneous amino acids and a stop codon. (B) Anti-V5-stained western blot showing the size of the Cv-d-V5 secreted from S2 cells. (C) Almost complete loss of PCV in adult tub-gal4 UAS-cv-d-RNAi wing (using UAS-cv-d-RNAi #9622R-1 from NIG-Fly). (D) Range of PCV phenotypes in various mutant, RNAi and rescued mutant adults. The penetrance of the cv-d1 PCV defect is increased by placing it over a deletion, whereas that of cv-d13 is not (not shown). (E) Pairwise BLAST scores between selected insect lipoproteins (known or predicted) from Drosophila melanogaster (Dros.), Aedes aegypti (Aedes, mosquito), Apis mellifera (Apis, honeybee), Tenebrio molitor (Teneb., mealworm beetle), Tribolium castaneum (Tribo., red flour beetle) and Acyrthosiphon pisum (Acyr., pea aphid). High BLAST scores group proteins into four families: Cv-d/160MEP, yolk Vtg, Apolipophorin (Apo)-related and Apolipophorins.
Fig. S2. cv-d13 only slightly affects wing size and does not detectably alter BMP signaling in late third instar wing discs. (A-C) Comparison of wild type (A) and cv-d13 (B) adult wings, superimposed in C. The size is only slightly smaller, and the positions of L2 or L5 did not shift closer to the anterior-posterior compartment boundary (typical of reduced Dpp signaling from the anterior-posterior compartment boundary of third instar wing discs) (Blair, 2007). (D,E) Wing pouch regions of wild type (A) and cv-d13 (B) late third instar wing discs, showing largely identical anti-pMad staining.
Fig. S3. Strong cv-d-V5 overexpression of UAS-cv-d-V5 with hh-gal4 disrupts the developing wing margin. (A) Adult wing showing scalloping of posterior wing margin after posterior expression of cv-d-V5. (B) Late third instar wing imaginal disc showing reduction of wing margin-specific anti-Wingless (Wg) staining (green), as well as reduction of posterior anti-Senseless (Sens) staining (purple) in flanking, Wg-dependent margin bristle precursors. Bar shows approximate domain of posterior hh-gal4 expression.
