Despite their importance as signaling hubs, the function of mitochondria-ER contact sites in mitochondrial quality control pathways remains unexplored. Here we describe a mechanism by which Mfn2, a mitochondria-ER tether, gates the autophagic turnover of mitochondria by PINK1 and parkin. Mitochondria-ER appositions are destroyed during mitophagy, and reducing mitochondria-ER contacts increases the rate of mitochondrial degradation. Mechanistically, parkin/PINK1 catalyze a rapid burst of Mfn2 phosphoubiquitination to trigger p97-dependent disassembly of Mfn2 complexes from the outer mitochondrial membrane, dissociating mitochondria from the ER. We additionally demonstrate that a major portion of the facilitatory effect of p97 on mitophagy is epistatic to Mfn2 and promotes the availability of other parkin substrates such as VDAC1. Finally, we reconstitute the action of these factors on Mfn2 and VDAC1 ubiquitination in a cell-free assay. We show that mitochondria-ER tethering suppresses mitophagy and describe a parkin-/PINK1-dependent mechanism that regulates the destruction of mitochondria-ER contact sites.

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Nepeta pogonosperma is an important medicinal plant with anti-inflammatory effects. An efficient and reliable transformation system for this plant was developed through optimization of several factors which affected the rate of Agrobacterium rhizogenes mediated transformation. Five bacterial strains, A4, ATCC15834, LBA9402, MSU440 and A13, two explant types, leaves and stems, and several co-cultivation media were examined. The maximum rate of hairy root induction was obtained from stem explants using MSU440 and ATCC15834 bacterial strains. A drastic increase in the frequency of transformation (91 %) was observed when MS medium lacking NH 4 NO 3 , KH 2 PO 4 , KNO 3 and CaCl 2 . Hairy root lines were confirmed by polymerase chain reaction (PCR) using primers of the rolB gene. According to Southern blot analysis, one T-DNA copy was inserted into each of the hairy root lines. In the present study, transgenic hairy roots have been obtained trough genetic transformation by A. rhizogenes harbouring two plasmids, the Ri plasmid and pBI121 binary vector harbouring gus reporter gene. Expression of the gus gene in transgenic hairy root was confirmed by histochemical GUS assay.

Ionisable amino‐lipid is a key component in lipid nanoparticles (LNPs), which plays a crucial role in the encapsulation of RNA molecules, allowing efficient cellular uptake and then releasing RNA from acidic endosomes. Herein, we present direct evidence for the remarkable structural transitions, with decreasing membrane curvature, including from inverse micellar, to inverse hexagonal, to two distinct inverse bicontinuous cubic, and finally to a lamellar phase for the two mainstream COVID‐19 vaccine ionisable ALC‐0315 and SM‐102 lipids, occurring upon gradual acidification as encountered in endosomes. The millisecond kinetic growth of the inverse cubic and hexagonal structures and the evolution of the ordered structural formation upon ionisable lipid‐RNA/DNA complexation are quantitatively revealed by in situ synchrotron radiation time‐resolved small angle X‐ray scattering coupled with rapid flow mixing. We found that the final self‐assembled structural identity, and the formation kinetics, were controlled by the ionisable lipid molecular structure, acidic bulk environment, lipid compositions, and nucleic acid molecular structure/size. The implicated link between the inverse membrane curvature of LNP and LNP endosomal escape helps future optimisation of ionisable lipids and LNP engineering for RNA and gene delivery.