Fig 1

Fig 1

(A) Representative images of a wild-type zebrafish embryo, a homozygous mutant embryo carrying the previously described stalactite (stl) missense mutation in mttp, and a homozygous c655 mutant embryo at 3 days post fertilization (dpf); Scale = 500 μm. The dark/opaque yolk phenotype in embryos from c655 heterozygous in-crosses segregated with a Mendelian ratio consistent with a homozygous recessive mutation, mean +/- SD. For source data, see S4 File. (B) Euclidean distance mapping analysis plots produced by MMAPPR [51], showing the likely genomic region of the c655 mutation. Plot of the LOESS fit to the mapping scores (Euclidean Distance⁴) across all 25 chromosomes (top) and expanded view of chromosome 1(GRCz10: CM002885.1) (bottom). Single nucleotide variants (SNVs) present in this 11 MB region in c655 mutant embryos were assessed for their effect on annotated genes using the Ensembl Variant Effect Predictor [52], including using the Sorting Intolerant from Tolerant algorithm (SIFT) [53], to predict the impact of changes on protein-coding sequence (tolerated or deleterious). We extracted variants that alter the protein-coding sequence as candidates for the causal mutation (223 variants in 64 genes, of which 42 are missense variants predicted to be deleterious; S1 File). One of the SNVs linked to the c655 phenotype was a G>T missense mutation predicted to be deleterious in exon 18 of the microsomal triglyceride transfer protein gene (ENSDARG00000008637, Chr1:11,421,261 GRCz10, red arrow in B shows the position of the G>T missense mutation in mttp). (C) Representative image of a trans-heterozygous mttp^stl/c655 embryo; 3 dpf, scale = 500 μm. The dark/opaque yolk phenotype is present at expected ratios and genotyping confirms that only the embryos with opaque yolks are trans-heterozygous for the mttp alleles. (D) Depiction of the mttp gene structure highlighting the locations of the stl (L475P) (position 11431645 (GRCz10), transcript mtp-204 (ENSDART00000165753.2)) and c655 (G863V) missense alleles in exon 11 and 18, respectively. An additional SNV in mttp at position Chr1:11,421,300 GRCz10 (T>C) causing a missense mutation (M850T) was also identified in c655 mutants; however, this SNV was not predicted to be deleterious and has been previously noted in Ensembl.