No abstract

Balanced fusion and fission are key for proper function and physiology of mitochondria 1,2 . Remodelling of the mitochondrial inner membrane (IM) is mediated by dynamin-like Mitochondrial genome maintenance 1 protein (Mgm1) in fungi or the related Optic atrophy protein 1 (OPA1) in animals [3][4][5] . Mgm1 is required for the preservation of mitochondrial DNA in yeast 6 , whereas mutations in the OPA1 gene in humans are a common cause for autosomal dominant optic atrophy, a genetic disorder affecting the optical nerve 7,8 . Mgm1 and OPA1 are present in mitochondria as a membrane-integral long (l) form and a short (s) form that is soluble in the intermembrane space. Yeast strains expressing temperaturesensitive mutants of Mgm1 9,10 or mammalian cells devoid of OPA1 display fragmented mitochondria 11,12 , suggesting an important role of Mgm1/OPA1 in IM fusion. Consistently, only the mitochondrial outer membrane (OM), but not the IM, fuses in the absence of functional Mgm1 13,14 . Mgm1 and OPA1 have also been shown to maintain proper cristae architecture 10,14 . For example, OPA1 prevents the release of pro-apoptotic factors by tightening cristae junctions 15 . Finally, s-OPA1 localises to mitochondrial constriction sites, where it presumably promotes mitochondrial fission 16 . How Mgm1/OPA1 perform their diverse functions in membrane fusion, scission, and cristae organisation is at present unknown. Here, we present crystal and electron cryo-tomography (cryo-ET) structures of Chaetomium thermophilum Mgm1. Mgm1 consists of a GTPase domain, a bundle signalling element (BSE) domain, a stalk, and a paddle domain containing a membrane binding site. Biochemical and cell-based experiments demonstrate that the Mgm1 stalk mediates assembly of bent tetramers into helical filaments. Cryo-ET of Mgm1-decorated lipid tubes and fluorescence microscopy experiments on reconstituted membrane tubes indicate how the tetramers assemble on positively or negatively curved membranes. Our findings convey how Mgm1/OPA1 filaments dynamically remodel the mitochondrial IM.We purified and crystallised a truncated s-Mgm1 isoform from the thermophilic fungus Chaetomium thermophilum (from here on Mgm1) (Fig. 1a, Extended Data Fig. 1a, Supplementary Data Fig. 1). Crystals of this construct grown in the absence of nucleotides diffracted to 3.6 Å resolution. The structure was solved by single anomalous dispersion (Extended Data Fig. 1b, c, Extended Data Table 1).The structure of Mgm1 contains four domains: A G domain, a bundle signalling element (BSE) domain, a stalk, and a paddle (Fig. 1a, b). The G domain closely resembles that of human dynamin (Extended Data Fig. 2). An interface across the nucleotide-binding site responsible for G domain dimerisation in the dynamin superfamily (the 'G interface') is highly conserved in Mgm1 (Extended Data Fig. 1e). The adjacent BSE domain consists of three helices derived from different regions of Mgm1 (Fig. 1a, b). The BSE domain contacts the G domain, as in the closed conformation of dynamin [17][18][19] . The M...

The majority of preproteins destined for mitochondria carry N-terminal presequences. The presequence translocase of the inner mitochondrial membrane (TIM23 complex) plays a central role in protein sorting. Preproteins are either translocated through the TIM23 complex into the matrix or are laterally released into the inner membrane. We report that the small hydrophobic protein Mgr2 controls the lateral release of preproteins. Mgr2 interacts with preproteins in transit through the TIM23 complex. Overexpression of Mgr2 delays preprotein release, whereas a lack of Mgr2 promotes preprotein sorting into the inner membrane. Preproteins with a defective inner membrane sorting signal are translocated into the matrix in wild-type mitochondria but are released into the inner membrane in Mgr2-deficient mitochondria. We conclude that Mgr2 functions as a lateral gatekeeper of the mitochondrial presequence translocase, providing quality control for the membrane sorting of preproteins.

Mitochondria are multifunctional metabolic factories and integrative signaling organelles of eukaryotic cells. The structural basis for their numerous functions is a complex and dynamic double-membrane architecture. The outer membrane connects mitochondria to the cytosol and other organelles. The inner membrane is composed of a boundary region and highly folded cristae membranes. The evolutionarily conserved mitochondrial contact site and cristae organizing system (MICOS) connects the two inner membrane domains via formation and stabilization of crista junction structures. Moreover, MICOS establishes contact sites between inner and outer mitochondrial membranes by interacting with outer membrane protein complexes. MICOS deficiency leads to a grossly altered inner membrane architecture resulting in far-reaching functional perturbations in mitochondria. Consequently, mutations affecting the function of MICOS are responsible for a diverse spectrum of human diseases. In this article, we summarize recent insights and concepts on the role of MICOS in the organization of mitochondrial membranes. This article is part of a Special Issue entitled: Membrane Contact Sites edited by Christian Ungermann and Benoit Kornmann.

Asgard archaea are considered to be the closest known relatives of eukaryotes. Their genomes contain hundreds of eukaryotic signature proteins (ESPs), which inspired hypotheses on the evolution of the eukaryotic cell1–3. A role of ESPs in the formation of an elaborate cytoskeleton and complex cellular structures has been postulated4–6, but never visualized. Here we describe a highly enriched culture of ‘Candidatus Lokiarchaeum ossiferum’, a member of the Asgard phylum, which thrives anaerobically at 20 °C on organic carbon sources. It divides every 7–14 days, reaches cell densities of up to 5 × 107 cells per ml and has a significantly larger genome compared with the single previously cultivated Asgard strain7. ESPs represent 5% of its protein-coding genes, including four actin homologues. We imaged the enrichment culture using cryo-electron tomography, identifying ‘Ca. L. ossiferum’ cells on the basis of characteristic expansion segments of their ribosomes. Cells exhibited coccoid cell bodies and a network of branched protrusions with frequent constrictions. The cell envelope consists of a single membrane and complex surface structures. A long-range cytoskeleton extends throughout the cell bodies, protrusions and constrictions. The twisted double-stranded architecture of the filaments is consistent with F-actin. Immunostaining indicates that the filaments comprise Lokiactin—one of the most highly conserved ESPs in Asgard archaea. We propose that a complex actin-based cytoskeleton predated the emergence of the first eukaryotes and was a crucial feature in the evolution of the Asgard phylum by scaffolding elaborate cellular structures.