Figure 6.

Variants in hTNPO2 cause different amounts of toxicity compared to wild-type hTNPO2 during fly development and in the eye

(A) UAS-hTNPO2 fly lines were generated. Western immunoblots (WBs) confirmed hTNPO2 protein levels are similar between lines using a drug-inducible ubiquitous driver (da-GAL4^GS) to express transgenes and a human TNPO1/2 antibody. Normalized hTNPO2 band density from three independent westerns were quantified. Each dot represents one independent sample. The mean from 3 individual samples is shown.

(B) da-GAL4 driven ectopic expression of UAS-hTNPO2:WT^HA reduces Mendelian ratios compared to UAS control flies, demonstrating toxicity during development. Variants p.Gln28Arg and p.Asp156Asn are more toxic than hTNPO2:WT whereas p.Trp727Cys is less toxic. Each dot represents one independent cross with >100 animals scored. The mean from three independent crosses is shown.

(C and D) Ectopic expression of UAS-hTNPO2:WT^HA disrupts eye development using either ey-GAL4 (early development) or GMR-GAL4 (late development). Scale bars = 100 μm.

(C) With ey-GAL4>hTNPO2:WT^HA, eyes are smaller than controls and have a rough eye phenotype. p.Trp370Cys and p.Trp370Arg are less toxic.

(D) With GMR-GAL4>hTNPO2:WT^HA, eyes are moderately smaller and there is a mild rough eye phenotype compared to controls. p.Gln28Arg and p.Asp156Asn are more toxic.

Statistics: 1-way ANOVAs with Dunnett’s (A) or Tukey’s (B) multiple comparisons test. no significance (n.s.) ≥ 0.05, ^∗p < 0.05, ^∗∗p < 0.01, ^∗∗∗p < 0.001, ^∗∗∗∗p < 0.0001. Error bars denote SD “UAS Control” is UAS-(empty).