Fig. 5. Phenotypes of unc-53 and their promoter-specific rescue. (A) The Egl phenotype was scored by counting the number of progeny from individual worms of different genotypes and is represented as an average with standard deviation for each strain. For wild-type, n152 and e2432 homozygous, and n152/mnDf87 heterozygous worms, the progeny from 16, 29, 20 and 23 individual worms, respectively, were counted. Transgenic strains, all in an unc-53(n152) background, containing the cosmid T28D2 or the unc-53 minigene expressed under the control of the AB, A or B promoter are represented as n152;T28D2, n152;pAB::unc-53, n152;pA::unc-53 and n152;pB::unc-53, respectively. For each of these, the progeny of 12, 23, 23 and 19 individuals from two, five, two and five independent strains, respectively, were counted. Each independent strain of a given genotype gave similar results. Both unc-53 alleles show a strong Egl phenotype, indistinguishable from that of n152/mnDf87 heterozygotes. The unc-53(n152) phenotype was largely rescued by the cosmid T28D2, by pAB::unc-53 and by pB::unc-53, but not by pA::unc-53. (B) Quantification of ALN and PLN axonal outgrowth defects. The worm was divided into three regions (1-3) with reference to the vulva and the anus. The stop point of ALN axons was determined using a pA::GFP reporter carried by wild-type (n=100), n152 homozygous (n=50) and two independent strains of n152;pA::unc-53 (n=100) worms. The percentage of axons stopping at each of the three positions was then calculated. The PLN defect was scored similarly for 70, 44 and 80 worms, respectively. The two n152;pA::unc-53 strains gave comparable results. The unc-53(n152) phenotype was largely rescued by pA::unc-53. (C) Quantification of the excretory canal outgrowth defect. The worm was divided into four regions (1-4) between the vulva and the anus. The stop point of canals was determined by DIC microscopy for wild type (n=100), n152 (n=100), n152/mnDf87 (n=52), n152;T28D2 (n=48), n152;pAB::unc-53 (n=50), n152;pA::unc-53 (n=62) and n152;pB::unc-53 (n=57). For the transgenic strains, two independent lines for each were examined. The percentage of canals stopping at each of the four positions was then calculated. unc-53(n152) was associated with a stereotyped arrest of the canal at the level of the vulva, that was almost as penetrant as n152/mnDf87 heterozygotes. The phenotype was largely rescued by the cosmid T28D2. The pAB::unc-53 and pA::unc-53 constructs gave a degree of rescue, in contrast to pB::unc-53.

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