Lysosomal storage disease proteo/lipidomic profiling using nMOST links ferritinophagy with mitochondrial iron deficiencies in cells lacking NPC2.

Lysosomal storage diseases (LSDs) comprised ~50 monogenic diseases characterized by the accumulation of cellular material in lysosomes and associated defects in lysosomal function, but systematic molecular phenotyping is lacking. Here, we develop a nanoflow-based multi-omic single-shot technology (nMOST) workflow allowing simultaneously quantify HeLa cell proteomes and lipidomes from more than two dozen LSD mutants, revealing diverse molecular phenotypes. Defects in delivery of ferritin and its autophagic receptor NCOA4 to lysosomes (ferritinophagy) were pronounced in NPC2-/- cells, which correlated with increased lyso-phosphatidylcholine species and multi-lamellar membrane structures visualized by cryo-electron-tomography. Ferritinophagy defects correlated with loss of mitochondrial cristae, MICOS-complex components, and electron transport chain complexes rich in iron-sulfur cluster proteins. Strikingly, mitochondrial defects were alleviated when iron was provided through the transferrin system. This resource reveals how defects in lysosomal function can impact mitochondrial homeostasis in trans and highlights nMOST as a discovery tool for illuminating molecular phenotypes across LSDs.

Complex dynamics in a synchronized cell-free genetic clock.

Complex dynamics such as period doubling and chaos occur in a wide variety of non-linear dynamical systems. In the context of biological circadian clocks, such phenomena have been previously found in computational models, but their experimental study in biological systems has been challenging. Here, we present experimental evidence of period doubling in a forced cell-free genetic oscillator operated in a microfluidic reactor, where the system is periodically perturbed by modulating the concentration of one of the oscillator components. When the external driving matches the intrinsic period, we experimentally find period doubling and quadrupling in the oscillator dynamics. Our results closely match the predictions of a theoretical model, which also suggests conditions under which our system would display chaotic dynamics. We show that detuning of the external and intrinsic period leads to more stable entrainment, suggesting a simple design principle for synchronized synthetic and natural genetic clocks.