(A) In primary rodent cortical neurons, the S85C, F115C, P154S, and T622A disease-associated MATR3 mutants have the same granular nuclear distribution as MATR3(WT)-EGFP. (B) All four disease mutations display a subtle but significant increase in toxicity compared to MATR3(WT)-EGFP (comparing to MATR3(WT)-EGFP, n = 2920: MATR3(S85C)-EGFP, HR = 1.16, n = 2031, ***p=3.79×10−6; MATR3(F115C)-EGFP, HR = 1.14, n = 2144, ***p=5.57×10−5; MATR3(P154S)-EGFP, HR = 1.24, n = 2092, ***p=1.77×10−11; MATR3(T622A)-EGFP, HR = 1.14, n = 2137, ***p=6.02×10−5; Cox proportional hazards). (C–D) siRNA targeting the endogenous rat Matr3 reduced MATR3 antibody reactivity by approximately 65% (scrambled siRNA, n = 576; anti-Matr3 siRNA, n = 508; p<0.0001, two-tailed t-test). (E) Neurons transfected with anti-Matr3 siRNA displayed a higher risk of death compared to those transfected with scrambled siRNA (HR = 1.20; scrambled siRNA, n = 2507; anti-Matr3, n = 2623; ***p=2.05×10−8, Cox proportional hazards). Scale bars in (A), 10 μm; scale bars in (C), 20 μm.