Fig. 2

Träge carries a tmod4 loss-of-function allele. (A) Linkage analysis of trg mutants through SNPtrack resulted in a peak at 31.96 Mb on chromosome (chr.) 16 (marked by green arrow). (B) Within the locus lies tmod4 that carries a nonsense mutation, indicated by a red arrowhead. (C) The triplet TTG that encodes a leucine residue in wild-type tmod4 is mutated to the stop codon TAG in tmod4^trg mutants (L132X). (D) Schematic of Tmod4 shows the tropomyosin and actin-capping domain (TM-cap) that comprises two tropomyosin-binding helices (orange) and one actin-binding helix (red) together with the actin-binding domain (LRR-cap) with its five leucine-rich repeats. The arrowhead marks the location of the L132X mutation. (E) Whole-mount in situ hybridization at 3 dpf shows an abundance of tmod4 transcript in trunk muscle that is strikingly reduced in tmod4^trg, suggesting nonsense-mediated decay. (F,F′) The weaker in situ signal in tmod4^trg is also noted on cross sections. Interestingly, tmod4 transcript is absent from the superficial slow muscle fibres (arrowheads), spinal cord (sc) and notochord (nc). Longitudinal sections show muscle-specific tmod4 transcript in the (G,G′) trunk and (H) head musculature. G′ shows an enlarged image of the region of interest in G. (I–J′) Administration of 200 μM of tmod4ATG(4+21) into wild-type embryos phenocopied tmod4^trg mutants by inducing a highly significant reduction in birefringence after normalization to control-injected embryos (P<0.01, n=3). (I) Quantification results, data are means ± s.e.m., **P<0.01. (J,J′) example images. contr., control; WT, wild type.