Fig. 3. SPON-1 is synthesized in muscle and localizes to basement membranes. (A) Ventral view of an adult midbody showing that Pspon-1-GFP (juEx593) is expressed in body wall muscles (arrows, two muscle quadrants). (B) DIC optics showing the muscle quadrants (arrows) on the ventral side near the vulva. (C) Embryonic expression of Pspon-1-GFP from the comma stage onwards. (D-H) SPON-1::GFP localization visualized by anti-GFP immunostaining. (D) In 1.5-fold stage embryos SPON-1::GFP is localized to muscle quadrants (arrowhead). (E) In juEx1111 larvae, SPON-1::GFP localizes at muscle dense bodies (inset, large arrowhead) and at M lines (inset, small arrowhead). (F) Colocalization of SPON-1::GFP (anti-GFP) and the dense body component vinculin (MH24 immunostaining) in juEx734 adult muscles. (G,H) SPON-1 was detected on the pharynx surface (overexpressing line juEx698, arrow, G) and within coelomocytes (SPON-1::GFP, arrowheads, H) in juEx734 adults. (I) Functional SPON-1::GFP expressed under the control of the pharynx-specific myo-2 gene (juEx1302) localizes to muscle BMs (arrowhead) in 3-fold stage embryos (anti-GFP immunostaining). (J,K) Immunostaining with anti-SPON-1 antibodies of animals of genotype spon-1(ju402); juEx698[spon-1(+)]. (J) Dorsal view of a 1.5-fold stage embryo stained with anti-SPON-1 (red), and anti-Myotactin (MH46) and anti-AJM-1 (MH27) (both green), which mark muscle-epidermal attachments and epidermal adherens junctions, respectively. SPON-1 is in muscle cells, beneath Myotactin staining. (K) Larva stained with anti-SPON-1 (red) and anti-MHC (green) to show body muscles. SPON-1 expression was detected in or adjacent to body muscles (arrow) and the excretory canal (arrowhead). Scale bars: 10 µm.