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bioRxiv · 10.1101/2022.11.02.514842

Intracellular calcium elevations drive the nucleation of FIP200- and ATG13-containing pre-autophagosomal structures that become omegasomes

Smith, M.·Schoenfelder, P.·Manifava, M.·Polson, H.·Tooze, S.·Roderick, L.·Ktistakis, N.
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Abstract

Ca2+ modulates autophagy at multiple steps including the induction and maturation of autophagosomes, but the magnitude and spatiotemporal properties of this calcium signal and its ultimate effect on the autophagic machinery are unclear. Focusing on the induction step leading to omegasome formation, we report that low but sustained elevations in cytosolic calcium levels induce omegasome formation but treatments that only transiently elevate calcium do not. The calcium-induced structures are early intermediates that mature into omegasomes but do not constitute full autophagosomes because they are partially devoid of late autophagy proteins ATG16 and LC3. In addition to omegasomes, all four components of the ULK complex (ULK1, FIP200, ATG13, ATG101) respond to calcium modulation: they translocate to early autophagy puncta in complete medium upon calcium elevation, and are inhibited from translocation during starvation by calcium chelation with BAPTA-2 AM. The principal early step affected by calcium lies downstream of mTORC1 inactivation and upstream of VPS34 activation, coinciding biochemically with phosphorylation of ATG13 at serine 318, which is known to require ULK1 activity. However, although the calcium-mediated step requires ATG9, FIP200 and ATG13, it does not require ULK1/2, suggesting that calcium does not directly regulate ULK1 activity but rather it regulates the mechanism by which the ULK complex components ATG13 and FIP200, together with ATG9, nucleate pre-autophagosomal precursors. This calcium-induced nucleation is sufficient to drive autophagy induction up to the omegasome step, but not beyond it.

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Smith, M., Schoenfelder, P., Manifava, M., Polson, H., Tooze, S., Roderick, L., Ktistakis, N.. 2022-11-02. Intracellular calcium elevations drive the nucleation of FIP200- and ATG13-containing pre-autophagosomal structures that become omegasomes. https://doi.org/10.1101/2022.11.02.514842

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