Fig. 4. Only Shot isoforms A and B, those that contain complete actin binding domains, are detectably expressed in the nervous system, and the long isoforms of Shot are present in axons.A, In situ hybridization with a cDNA probe containing 5′ sequence derived from the actin binding isoform A and the first half of the actin binding domain coding sequence shared also by isoform B reveals transcripts in the CNS (arrow) and muscle attachment cells (concave arrow) at stage 17 as well as earlier (data not shown). B, Whole-mountin situ hybridization to a stage 17 embryo with a cDNA probe containing exons specific for isoforms C and D reveals comparable levels of expression in muscle attachment cells (concave arrow), but no detectable expression in the CNS (arrow). Scale bar, 30 μm. C, Confocal (1 μm) section of a stage 14 wild-type embryo stained with polyclonal anti-Kak (Strumpf and Volk, 1998). This antibody recognizes epitopes in the rod domain present in the long isoforms of Shot/Kak. Axons in the anterior commissure (ac) and posterior commissure (pc), as well as motor axons (ma) and longitudinal axons, contain Shot/Kak. Scale bar, 10 μm.D, A confocal section showing Shot/Kak (red, arrowhead) protein in the sensory axons emanating from the dorsal cluster of sensory neurons in each abdominal hemisegment. The antiserum also recognizes Shot protein in epithelial cells and a nonspecific epitope in tracheal lumen. The epithelial and sensory axon staining disappears inshot mutants, but the tracheal lumen staining does not. Scale bar, 10 μm. E, The dorsal cluster sensory neurons and their axons in the same section revealed with mAb 22C10 (Fujita et al., 1982). F, Merge of D andE.