Fig. S1. E414 is a loss-of-function allele of RacGAP50C. Stage 16 Drosophila embryos stained for Muscle myosin. (A-B′) Muscle patterning at low (A,B) and high (A′,B′) magnification in wild-type (A,A′) or E414/E414 (B,B′) embryos. Arrowheads in A′ mark muscle 21 (LT1). Arrowheads in B′-F mark muscles making inappropriate attachments. Arrow in B′ marks LT muscle that fails to migrate. (D) E414/Df(2R)CX1 embryo. (E) E414/RacGAP50C^DH15 embryo. Non-complementation identified E414 as a RacGAP50C allele. (F) Homozygous mutants of the strong loss-of-function allele, RacGAP50C^DH15, have defects similar to E414. (C) Schematic showing the wild-type muscle pattern of a single abdominal hemisegment from a Drosophila embryo, with name and number labels for muscles as referred to throughout this work. DA, dorsal acute; DO, dorsal oblique; LT, lateral transverse; VL, ventral lateral; VO, ventral oblique.

Fig. S2. Localization of RacGAP. (A,B) RacGAP (green) and DAPI (blue) staining in stage 14 (A) and 15 (B) embryos. RacGAP is localized throughout the muscle cytoplasm and is concentrated at the nuclear periphery (arrowheads) during the stages when myotube migration is occurring. In A, a fusing myoblast with diffuse RacGAP staining can be seen (asterisk). (C) Staining with a rat antibody against RacGAP (see Materials and methods). Arrowhead indicates a concentration of RacGAP near the nuclear periphery.

Fig. S3. The overall polarity of the muscle fiber is not affected in RacGAP^DH15 mutants. The transmembrane receptor Roundabout is localized to the ends of muscle fibers at stage 16 in both wild type (A) and RacGAP^DH15 mutants (B), demonstrating that the overall polarity of the muscle fiber is not affected.