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Fig. S1
Characterization of lsm8 mutants. Related to Figure 1
(A) Representative sequence traces indicating the G>T transition at nucleotide position 8611002 (Zv9) on chromosome 4; conceptual translation of the nucleotide sequence is indicated in three-letter code (stop codon is marked by *).
(B) Deduced protein structure of wild-type and predicted mutant (E72X) proteins.
(C) Phenotypic rescue of lsm8 mutation by injection of wild-type zebrafish (Dr) lsm8 mRNA as determined by the extent of thymopoietic activity (as measured by the ratio of hybridization signals obtained for rag1 and growth hormone [gh]); error bars represent S.E.M.; the significance level of the difference is indicated (t-test; two-tailed). The number (n) of embryos analyzed is indicated.
(D) Functionally competent thymic rudiment in lsm8 mutants. At 5 dpf, expression of the thymopoietic marker foxn1 in mutant embryos is indistinguishable from that in wild-type embryos (circles; left panels). To examine the receptive capacity of thymic rudiments, purified ikaros-positive hematopoietic kidney marrow cells from adult wild-type ikaros:eGFP transgenic fish were injected into the sinus venosus of wild-type and heterozygous embryos (collectively designated as +/± genotypes) and lsm8 mutant embryos at 2 dpf. 72 hours later, the numbers of green cells in the rudiment were counted; a total of 40 lsm8+/± and 10 lsm8-/- embryos were successfully injected; transplantation failed in 47 (54%) and 28 (74%) embryos, respectively. Scale bar, 10 μm.
(E) Characterization of early hematopoiesis. Whole mount RNA in situ hybridization was carried out with the indicated probes at various time points. runx1, 36 hpf. cmyb, 36 hpf; gata1, 24 hpf; l-plastin, 24 hpf. Shown are overviews (left panels) and magnifications (right panels). Scale bars, 50 μm.