A. Genetic mapping of the ia50 mutation. Three-point mapping with the indicated single nucleotide polymorphisms placed ia50 at +5.67 mu on chromosome V. Injection of a pool of 3 cosmids in the region (F20G2, F53C11, and F55B12) rescued the ia50 homozygous mutants, as assayed by Pnhr-57::GFP expression. This region includes the swan-1 gene. swan-1(ok267) is a deletion mutation. B.
ia50 mutants carry a point mutation at the 3′ end of intron 2 of swan-1. This results in failure to splice out intron 2 from the mRNA and introduces an early stop codon, illustrated by an asterisk. C. Depletion of swan-1 by bacterially-mediated RNAi increased expression of the Pnhr-57::GFP reporter, as assayed by protein blots. In control experiments, RNAi for the related swan-2 gene did not alter Pnhr-57::GFP levels. D. The swan-1(ok267) mutation increased expression of the Pnhr-57::GFP reporter in vhl-1(ok161) animals, and this phenotype was suppressed by the hif-1(ia04) strong loss-of-function mutation. Pnhr-57::GFP levels were assayed by protein blots, and the error bars represent standard errors. **: P<0.01.