Parkin mitochondria in the autophagosome

Narendra et al. (see p. 795 of this issue) have made an exciting new discovery that links the fields of mitochondrial quality control and the genetics of Parkinson's disease (PD). Through an elegant series of high-resolution imaging experiments, they are the first to provide evidence that the PARK2 gene product Parkin is selectively recruited to damaged or uncoupled mitochondria. This recruitment leads to the clearance of the organelles through the autophagosome, demonstrating a primary function for Parkin in the regulation of mitochondrial turnover. This work significantly increases our understanding of PD and provides a new framework for the development of therapeutic interventions

Bax/Bak promote sumoylation of DRP1 and its stable association with mitochondria during apoptotic cell death

Dynamin-related protein 1 (DRP1) plays an important role in mitochondrial fission at steady state and during apoptosis. Using fluorescence recovery after photobleaching, we demonstrate that in healthy cells, yellow fluorescent protein (YFP)–DRP1 recycles between the cytoplasm and mitochondria with a half-time of 50 s. Strikingly, during apoptotic cell death, YFP-DRP1 undergoes a transition from rapid recycling to stable membrane association. The rapid cycling phase that characterizes the early stages of apoptosis is independent of Bax/Bak. However, after Bax recruitment to the mitochondrial membranes but before the loss of mitochondrial membrane potential, YFP-DRP1 becomes locked on the membrane, resulting in undetectable fluorescence recovery. This second phase in DRP1 cycling is dependent on the presence of Bax/Bak but independent of hFis1 and mitochondrial fragmentation. Coincident with Bax activation, we detect a Bax/Bak-dependent stimulation of small ubiquitin-like modifier-1 conjugation to DRP1, a modification that correlates with the stable association of DRP1 with mitochondrial membranes. Altogether, these data demonstrate that the apoptotic machinery regulates the biochemical properties of DRP1 during cell death

Insertion of an uncharged polypeptide into the mitochondrial inner membrane does not require a trans-bilayer electrochemical potential: effects of positive charges

AbstractMitochondria with a ruptured outer membrane exhibited impaired import into this membrane of an outer membrane fusion protein containing the signal-anchor sequence of Mas70p. However, the Mas70p signal-anchor efficiently targeted and inserted the protein directly into exposed regions of the inner membrane. Import into the inner membrane was dependent on δψ and this dependence was due to the presence of the positively-charged amino acids located at positions 2, 7, and 9 of the signal-anchor. In contrast to wild-type signal-anchor, mutants lacking the positively-charged residues mediated import into the inner membrane in both the presence and absence of δψ. The results suggest two conclusions: (1) δψ-dependent import of the signal-anchor sequence was due exclusively to an effect of δψ on the positively-charged domain of the signal-anchor, rather than to an effect of δψ on a property of the inner membrane import machinery; (2) in the absence of δψ, the positively-charged domain of the signal-anchor prevented the otherwise import-competent signal-anchor from inserting into the membrane. This suggests that the positively-charged domain leads import across the inner membrane, and that dqi is required to vectorially clear this domain in order to allow the distal region of the signal-anchor to enter the translocation pathway. The implications of these findings on the mechanism of import into the mitochondrial inner membrane and matrix are discussed

Pathways From Food Insecurity to Intimate Partner Violence Perpetration Among Peri-Urban Men in South Africa.

INTRODUCTION: Although poverty is sometimes seen as a driver of intimate partner violence victimization, less is known about how it intersects with men's violence perpetration. Food insecurity is a sensitive marker of poverty that may have unique mechanisms leading to men's intimate partner violence perpetration given its association with gender roles and men "providing for the family." METHODS: Using cluster-based sampling, the team conducted an audio-assisted questionnaire in 2016 among men living in a peri-urban settlement near Johannesburg, South Africa. The aim was to examine the relationship between men's food insecurity and their use of past-year intimate partner violence, and to explore the pathways linking these two conditions. RESULTS: Among 2,006 currently partnered men, nearly half (48.4%) perpetrated intimate partner violence and more than half (61.4%) were food insecure. Food insecurity was associated with doubled odds of intimate partner violence (OR=2.15, 95% CI=1.73, 2.66). This association persisted after controlling for sociodemographics, relationship characteristics, and neighborhood clustering. In a structural equation model, food insecurity retained a direct relationship with men's violence perpetration and worked through indirect pathways of mental health and relationship quality. CONCLUSIONS: Addressing men's perpetration of intimate partner violence may require examination of broader structural challenges, such as food insecurity. Future interventions should consider livelihood strategies alongside relationship and mental health approaches

Vps35 Mediates Vesicle Transport between the Mitochondria and Peroxisomes

SummaryMitochondria-derived vesicles (MDVs) have been shown to transport cargo from the mitochondria to the peroxisomes [1]. Mitochondria and peroxisomes share common functions in the oxidation of fatty acids and the reduction of damaging peroxides [2, 3]. Their biogenesis is also linked through both the activation of master transcription factors such as PGC-1α [4, 5] and the common use of fission machinery, including DRP1, Mff, and hFis1 [6–9]. We have previously shown that MDVs are formed independently of the known mitochondrial fission GTPase Drp1 and are enriched for a mitochondrial small ubiquitin-like modifier (SUMO) E3 ligase called MAPL (mitochondrial-anchored protein ligase) [1]. Here, we demonstrate that the retromer complex, a known component of vesicle transport from the endosome to the Golgi apparatus [10–13], regulates the transport of MAPL from mitochondria to peroxisomes. An unbiased screen shows that Vps35 and Vps26 are found in complex with MAPL, and confocal imaging reveals Vps35 recruitment to mitochondrial vesicles. Silencing of Vps35 or Vps26A leads to a significant reduction in the delivery of MAPL to peroxisomes, placing the retromer within a novel intracellular trafficking route and providing insight into the formation of MAPL-positive MDVs

Analysis of the yeast arginine methyltransferase Hmt1p/Rmt1p and its in vivo function. Cofactor binding and substrate interactions

Many eukaryotic RNA-binding proteins are modified by methylation of arginine residues. The yeast Saccharomyces cerevisiae contains one major arginine methyltransferase, Hmt1p/Rmt1p, which is not essential for normal cell growth. However, cells missing HMT1 and also bearing mutations in the mRNA-binding proteins Np13p or Cbp80p can no longer survive, providing genetic backgrounds in which to study Hmt1p function. We now demonstrate that the catalytically active form of Hmt1p is required for its activity in vivo. Amino acid changes in the putative Hmt1p S-adenosyl-L-methionine-binding site were generated and shown to be unable to catalyze methylation of Np13p in vitro and in vivo or to restore growth to strains that require HMT1. In addition these mutations affect nucleocytoplasmic transport of Np13p. A cold- sensitive mutant of Hmt1p was generated and showed reduced methylation of Np13p, but not of other substrates, at 14 °C. These results define new aspects of Hmt1 and reveal the importance of its activity in vivo

Golgi derived PI(4)P-containing vesicle drives late steps of mitochondrial division

Mitochondrial plasticity is a key regulator of cell fate decisions. Mitochondrial division involves Dynamin-related protein-1 (Drp1) oligomerization, which constricts membranes at endoplasmic reticulum (ER) contact sites. The mechanisms driving the final steps of mitochondrial division are still unclear. Here, we found that microdomains of phosphatidylinositol 4-phosphate (PI(4)P) on Trans-Golgi network (TGN) vesicles were recruited to mitochondria-ER contact sites and could drive mitochondrial division downstream of Drp1. The loss of the small GTPase ADP-ribosylation factor 1 (Arf1) or its effector, phosphatidylinositol 4-kinase IIIβ (PI(4)KIIIβ) in different mammalian cell lines, prevented PI(4)P generation and led to a hyperfused and branched mitochondrial network, marked with extended mitochondrial constriction sites. Thus, recruitment of TGN-PI(4)P-containing vesicles at mitochondria-ER contact sites may trigger final events leading to mitochondrial scission.This work was supported by the Canadian Institutes of Health Research Operating Grants Program (CIHR grant 68833 to H.M.M.), the Medical Research Council (MRC grants MC_UU_00015/7 and RG89175 to J.P.), the Isaac Newton Trust (grant RG89529 to J.P.), and the Wellcome Trust Institutional Strategic Support Fund (grant RG89305 to J.P.). H.M.M. is a Canada Research Chair. S.N. and L.-C.T. are recipients of Daiichi Sankyo Foundation of Life Science and Ramon Areces postdoctoral fellowships, respectively. L.T. was supported by an MRC-funded graduate student fellowship. V.P was supported by the European Union’s Horizon 2020 research and innovation program (MITODYN-749926)

The dynamics of cardiolipin synthesis post-mitochondrial fusion

AbstractAlteration in mitochondrial fusion may regulate mitochondrial metabolism. Since the phospholipid cardiolipin (CL) is required for function of the mitochondrial respiratory chain, we examined the dynamics of CL synthesis in growing Hela cells immediately after and 12h post-fusion. Cells were transiently transfected with Mfn-2, to promote fusion, or Mfn-2 expressing an inactive GTPase for 24h and de novo CL biosynthesis was examined immediately after or 12h post-fusion. Western blot analysis confirmed elevated Mfn-2 expression and electron microscopic analysis revealed that Hela cell mitochondrial structure was normal immediately after and 12h post-fusion. Cells expressing Mfn-2 exhibited reduced CL de novo biosynthesis from [1,3-3H]glycerol immediately after fusion and this was due to a decrease in phosphatidylglycerol phosphate synthase (PGPS) activity and its mRNA expression. In contrast, 12h post-mitochondrial fusion cells expressing Mfn-2 exhibited increased CL de novo biosynthesis from [1,3-3H]glycerol and this was due to an increase in PGPS activity and its mRNA expression. Cells expressing Mfn-2 with an inactive GTPase activity did not exhibit alterations in CL de novo biosynthesis immediately after or 12h post-fusion. The Mfn-2 mediated alterations in CL de novo biosynthesis were not accompanied by alterations in CL or monolysoCL mass. [1-14C]Oleate incorporation into CL was elevated at 12h post-fusion indicating increased CL resynthesis. The reason for the increased CL resynthesis was an increased mRNA expression of tafazzin, a mitochondrial CL resynthesis enzyme. Ceramide-induced expression of PGPS in Hela cells or in CHO cells did not alter expression of Mfn-2 indicating that Mfn-2 expression is independent of altered CL synthesis mediated by elevated PGPS. In addition, Mfn-2 expression was not altered in Hela cells expressing phospholipid scramblase-3 or a disrupted scramblase indicating that proper CL localization within mitochondria is not essential for Mfn-2 expression. The results suggest that immediately post-mitochondrial fusion CL de novo biosynthesis is “slowed down” and then 12h post-fusion it is “upregulated”. The implications of this are discussed

Common bottlenose dolphin, Tursiops truncatus, seasonal habitat use and associations with habitat characteristics in Roanoke Sound, North Carolina

Understanding how habitat characteristics influence common bottlenose dolphin, Tursiops truncatus, distribution and behavior can be useful for conservation. The dolphin community in Roanoke Sound, North Carolina primarily exhibits seasonal residency and there is limited information on their habitat use. The objectives of this study were to increase habitat use knowledge and determine the relationship between habitat characteristics and dolphin distribution using standardized photographic-identification data (2009 – 2017). A hot spot (Getis-Ord Gi*) analysis showed dolphins frequently use the southern region containing the mouth of the estuary for feeding and traveling. Habitat characteristics were modeled with zero-altered gamma (ZAG), generalized linear (GLM), and generalized additive (GAM) models to predict dolphin group density. Models showed that groups were more likely to be present in areas with greater benthic slope variation and shallow areas closer to land, and that different habitat characteristics were associated with feed, social, and travel activities. This study suggests that Roanoke Sound provides a seasonal foraging area and travel corridor between the estuaries and coastal waters. This information contributes baseline knowledge of how habitat potentially influences dolphin distribution and behavior which can be useful for management and conservation, especially in areas where habitat changes and impacts need to be assessed

A novel cell-free mitochondrial fusion assay amenable for high-throughput screenings of fusion modulators

Abstract Background Mitochondria are highly dynamic organelles whose morphology and position within the cell is tightly coupled to metabolic function. There is a limited list of essential proteins that regulate mitochondrial morphology and the mechanisms that govern mitochondrial dynamics are poorly understood. However, recent evidence indicates that the core machinery that governs mitochondrial dynamics is linked within complex intracellular signalling cascades, including apoptotic pathways, cell cycle transitions and nuclear factor kappa B activation. Given the emerging importance of mitochondrial plasticity in cell signalling pathways and metabolism, it is essential that we develop tools to quantitatively analyse the processes of fission and fusion. In terms of mitochondrial fusion, the field currently relies upon on semi-quantitative assays which, even under optimal conditions, are labour-intensive, low-throughput and require complex imaging techniques. Results In order to overcome these technical limitations, we have developed a new, highly quantitative cell-free assay for mitochondrial fusion in mammalian cells. This assay system has allowed us to establish the energetic requirements for mitochondrial fusion. In addition, our data reveal a dependence on active protein phosphorylation for mitochondrial fusion, confirming emerging evidence that mitochondrial fusion is tightly integrated within the global cellular response to signaling events. Indeed, we have shown that cytosol derived from cells stimulated with different triggers either enhance or inhibit the cell-free fusion reaction. Conclusions The adaptation of this system to high-throughput analysis will provide an unprecedented opportunity to identify and characterize novel regulatory factors. In addition, it provides a framework for a detailed mechanistic analysis of the process of mitochondrial fusion and the various axis of regulation that impinge upon this process in a wide range of cellular conditions. See Commentary: http://www.biomedcentral.com/1741-7007/8/9