No abstract

The recognition that cytosolic mtDNA activates cGAS-STING innate immune signaling has unlocked novel disease mechanisms. Here, an uncharacterized variant predicted to affect TOP1MT function, P193L, was discovered in a family with multiple early-onset autoimmune diseases, including Systemic Lupus Erythematosus (SLE). Although there was no previous genetic association between TOP1MT and autoimmune disease, the role of TOP1MT as a regulator of mtDNA led us to investigate whether TOP1MT could mediate the release of mtDNA to the cytosol, where it could then activate the cGAS-STING innate immune pathway known to be activated in SLE and other autoimmune diseases. Through analysis of cells with reduced TOP1MT expression, we show that loss of TOP1MT results in release of mtDNA to the cytosol, which activates the cGAS-STING pathway. We also characterized the P193L variant for its ability to rescue several TOP1MT functions when expressed in TOP1MT knockout cells. We show that the P193L variant is not fully functional, as its re-expression at high levels was unable to rescue mitochondrial respiration deficits, and only showed partial rescue for other functions, including repletion of mtDNA replication following depletion, nucleoid size, steady state mtDNA transcripts levels, and mitochondrial morphology. Additionally, expression of P193L at endogenous levels was unable to rescue mtDNA release-mediated cGAS-STING signaling. Overall, we report a link between TOP1MT and mtDNA release leading to cGAS-STING activation. Moreover, we show that the P193L variant has partial loss of function that may contribute to autoimmune disease susceptibility via cGAS-STING mediated activation of the innate immune system.

Systemic lupus erythematosus (SLE) is an autoimmune condition where the underlying dysfunction is often unknown. Here, we identify a novel link between the mitochondrial topoisomerase TOP1MT and SLE, as a novel variant predicted to affect TOP1MT function, P193L, was identified in a family with SLE and other autoimmune diseases. Although there was no previous genetic association between TOP1MT and SLE, the role of TOP1MT as a regulator of mtDNA led us to investigate whether TOP1MT could mediate the release of mtDNA to the cytosol, where it could then activate the cGAS-STING innate immune pathway. This pathway increases the expression of type-I interferons and is known to be activated in some patients with SLE. Through analysis of cells lacking TOP1MT, or re-expressing the P193L variant, we established a new association by which TOP1MT dysfunction leads to increased release of mtDNA to the cytosol, causing activation of the cGAS-STING pathway. We also characterized the P193L variant for its ability to rescue several TOP1MT functions in TOP1MT knockout cells. While re-expression of the P193L variant was able to rescue steady state mtDNA copy number, most functions investigated only showed partial rescue, including repletion of mtDNA replication following depletion, nucleoid size, steady state mtDNA transcripts levels, and mitochondrial morphology. Meanwhile, the P193L variant failed to rescue decreased mitochondrial respiration. Overall, these findings support the notion that P193L variant is not fully functional and likely influences susceptibility to SLE via cGAS-STING mediated activation of the innate immune system.