Fig. 2

tor mutant embryos are rescued by injection of pnrc2 mRNA and phenocopied by frame-shifting mutation of the pnrc2 locus. Injection of 100 pg pnrc2 mRNA has no effect in wild-type sibling embryos (n=15/15 non-mutant siblings) (A, A’ vs B, B’) but restores striped her1 expression in tor mutant embryos (n=10/10 mutants) (C, C’ vs D, D’; Table 2). In these experiments, rescued tor^b644 mutant embryos were distinguished from wild-type siblings by lack of expression of pou3f1, a gene located in the tor^b644 deficiency interval. Asterisks (*) mark the neural pou3f1 expression domain. Arrows mark the her1 expression domain (A-D), magnified in dorsal view to the right of each embryo (A’-D’). Using CRISPR-based mutagenesis, we induced a 17 bp deletion within the pnrc2 coding sequence, creating an early frameshift allele, designated pnrc2^oz22 (E). Predicted mutant protein sequence is based on sequenced genomic DNA (E). The pnrc2^oz22 allele fails to complement the her1 accumulation phenotype of the tor^b644 deletion allele (n=4/15 embryos from a heterozygote intercross with tor-like her1 accumulation) (F, G). Similar to tor^b644 mutants, pnrc2^oz22 mutant embryos lack a striped her1 expression pattern (H, H').