Figure 5.

Polyglutamine Moves MOAG-2/LIR-3 Cytosol Where It Turns into a Positive Regulator of Protein Aggregation

(A and B) Number of body bends in N2 wild-type worms and worms overexpressing MOAG-2/LIR-3 (OE) (A) and Q40 wild-type and Q40 worms overexpressing MOAG-2/LIR-3 (Q40;OE) (B). The average of three biological replicates is represented. Data are represented as mean ± SEM and significance was calculated using a two-tailed unpaired Student’s t test. ^∗∗p < 0.01; ^∗∗∗p < 0.001. See also Figure S5A.

(C) Thioflavin T reaction profiles of 4 μM HttQ48 or HttQ23 solutions in the absence or presence of equimolar concentrations of MOAG-2/LIR-3 (black) or the modifier of aggregation MOAG-4 (blue). The concentration of MOAG-2/LIR-3 in whole-organism lysates has been determined to be between 0.3 and 8.5 ppm (http://pax-db.org). The average of four replicates is represented and error bars indicate mean ± SD.

(D) Western blot of subcellular fractionation of MOAG-3/LIR-3, which was detected using α-FLAG antibody. Q40-YFP, LMN-1 (nuclear marker), and α-tubulin (cytosolic marker) were used as controls. See Figure S5C.

(E) Western blot analysis of MOAG-2/LIR-3 SDS solubility from wild-type (WT) and MOAG-2/LIR-3-overexpressing (OE) worms in N2 and polyglutamine worms (Q40). Q40-YFP and β-actin expression were included as controls.

In (D) and (E), the results were generated from L4-stage animals and a representative experiment of three biological replicates is shown.

(F) Model proposing how MOAG-2/LIR-3 drives polyglutamine aggregation. MOAG-2/LIR-3 normally regulates the transcription of small ncRNAs in the nucleus. In the presence of polyglutamine, MOAG-2/LIR-3 is moved to the cytosol, where it exposes an ectopic aggregation-promoting activity.