PREFACE Mitochondria are remarkably dynamic organelles that migrate, divide and fuse. Cycles of mitochondrial fission and fusion ensure metabolite and mitochondrial DNA (mtDNA) mixing and dictate organelle shape, number and bioenergetic functionality. There is mounting evidence that mitochondrial dysfunction is an early and causal event in neurodegeneration. Mutations in mitochondrial fusion GTPases (mitofusin-2 and optic atrophy-1), neurotoxins and oxidative stress all disrupt the cable-like morphology of functional mitochondria. This results in impaired bioenergetics and mitochondrial migration and can trigger neurodegeneration. These findings suggest potential new treatment avenues for neurodegenerative diseases.

Huntington disease (HD) is an inherited and incurable neurodegenerative disorder caused by an abnormal polyglutamine (polyQ) expansion in huntingtin (HTT). PolyQ length determines disease onset and severity with a longer expansion causing earlier onset. The mechanisms of mutant HTT-mediated neurotoxicity remain unclear; however, mitochondrial dysfunction is a key event in HD pathogenesis1,2. Here we tested whether mutant HTT impairs the mitochondrial fission/fusion balance and thereby causes neuronal injury. We show that mutant HTT triggers mitochondrial fragmentation in neurons and fibroblasts of HD individuals in vitro and HD mice in vivo before the presence of neurological deficits and HTT aggregates. Interestingly, mutant HTT abnormally interacts with the mitochondrial fission GTPase dynamin-related protein 1 (DRP1) in HD mice and individuals which in turn stimulates its enzymatic activity. Importantly, mutant HTT-mediated mitochondrial fragmentation, defects in anterograde and retrograde mitochondrial transport, and neuronal cell death are all rescued by reducing DRP1 GTPase activity with the dominant-negative DRP1K38A mutant. Thus, DRP1 might represent a new therapeutic target to combat neurodegeneration in HD.

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The molecular bases underlying the pathogenesis of neurodegenerative diseases are gradually being disclosed. One problem that investigators face is distinguishing primary from secondary events. Rare, inherited mutations causing familial forms of these disorders have provided important insights into the molecular networks implicated in disease pathogenesis. Increasing evidence indicates that accumulation of aberrant or misfolded proteins, protofibril formation, ubiquitin-proteasome system dysfunction, excitotoxic insult, oxidative and nitrosative stress, mitochondrial injury, synaptic failure, altered metal homeostasis and failure of axonal and dendritic transport represent unifying events in many slowly progressive neurodegenerative disorders.

Lethal factor (LF) is a protein (relative molecular mass 90,000) that is critical in the pathogenesis of anthrax. It is a highly specific protease that cleaves members of the mitogen-activated protein kinase kinase (MAPKK) family near to their amino termini, leading to the inhibition of one or more signalling pathways. Here we describe the crystal structure of LF and its complex with the N terminus of MAPKK-2. LF comprises four domains: domain I binds the membrane-translocating component of anthrax toxin, the protective antigen (PA); domains II, III and IV together create a long deep groove that holds the 16-residue N-terminal tail of MAPKK-2 before cleavage. Domain II resembles the ADP-ribosylating toxin from Bacillus cereus, but the active site has been mutated and recruited to augment substrate recognition. Domain III is inserted into domain II, and seems to have arisen from a repeated duplication of a structural element of domain II. Domain IV is distantly related to the zinc metalloprotease family, and contains the catalytic centre; it also resembles domain I. The structure thus reveals a protein that has evolved through a process of gene duplication, mutation and fusion, into an enzyme with high and unusual specificity.

Summary In response to many apoptotic stimuli, oligomerization of Bax is essential for mitochondrial outer membrane permeabilization and the ensuing release of cytochrome c. These events are accompanied by mitochondrial fission that appears to require Drp1, a large GTPase of the dynamin superfamily. Loss of Drp1 leads to decreased cytochrome c release by a mechanism that is poorly understood. Here we show that Drp1 stimulates tBid-induced Bax oligomerization and cytochrome c release by promoting tethering and hemifusion of membranes in vitro. This function of Drp1 is independent of its GTPase activity and relies on arginine 247 and the presence of cardiolipin in membranes. In cells, overexpression of Drp1 R247A/E delays Bax oligomerization and cell death. Our findings reveal a novel function of Drp1 and provide a new insight into the mechanism of Bax oligomerization.

Mutations in OPA1, a dynamin-related GTPase involved in mitochondrial fusion, cristae organization and control of apoptosis, have been linked to non-syndromic optic neuropathy transmitted as an autosomal-dominant trait (DOA). We here report on eight patients from six independent families showing that mutations in the OPA1 gene can also be responsible for a syndromic form of DOA associated with sensorineural deafness, ataxia, axonal sensory-motor polyneuropathy, chronic progressive external ophthalmoplegia and mitochondrial myopathy with cytochrome c oxidase negative and Ragged Red Fibres. Most remarkably, we demonstrate that these patients all harboured multiple deletions of mitochondrial DNA (mtDNA) in their skeletal muscle, thus revealing an unrecognized role of the OPA1 protein in mtDNA stability. The five OPA1 mutations associated with these DOA 'plus' phenotypes were all mis-sense point mutations affecting highly conserved amino acid positions and the nuclear genes previously known to induce mtDNA multiple deletions such as POLG1, PEO1 (Twinkle) and SLC25A4 (ANT1) were ruled out. Our results show that certain OPA1 mutations exert a dominant negative effect responsible for multi-systemic disease, closely related to classical mitochondrial cytopathies, by a mechanism involving mtDNA instability.

The Death Inducing Signaling Complex (DISC) formed by Fas receptor, FADD and caspase-8 is a pivotal trigger of apoptosis1-3. The Fas/FADD DISC represents a receptor platform, which once assembled initiates the induction of programmed cell death. A highly oligomeric network of homotypic protein interactions comprised of the death domains (DD) of Fas and FADD is at the center of DISC formation4, 5. Thus characterising the mechanistic basis for the Fas/FADD interaction is paramount for understanding DISC signaling but has remained enigmatic largely due to a lack of structural data. We have successfully formed and isolated the Fas/FADD DD complex and here we report the 2.7 Å crystal structure. The complex shows a tetrameric arrangement of four FADD DDs bound to four Fas DDs. We show that an opening of the Fas DD exposes the FADD binding site and simultaneously generates a Fas/Fas bridge. The result is a regulatory Fas/FADD complex bridge governed by weak protein:protein interactions revealing a model where the complex functions as a mechanistic switch. This switch prevents accidental DISC assembly, yet allows for highly processive DISC formation and clustering upon a sufficient stimulus. Thus besides depicting a previously unknown mode of death domain interactions, these results further uncover a mechanism for receptor signaling solely by oligomerization and clustering events.