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)

Whole-genome sequencing reveals host factors underlying critical COVID-19

Critical COVID-19 is caused by immune-mediated inflammatory lung injury. Host genetic variation influences the development of illness requiring critical care1 or hospitalization2–4 after infection with SARS-CoV-2. The GenOMICC (Genetics of Mortality in Critical Care) study enables the comparison of genomes from individuals who are critically ill with those of population controls to find underlying disease mechanisms. Here we use whole-genome sequencing in 7,491 critically ill individuals compared with 48,400 controls to discover and replicate 23 independent variants that significantly predispose to critical COVID-19. We identify 16 new independent associations, including variants within genes that are involved in interferon signalling (IL10RB and PLSCR1), leucocyte differentiation (BCL11A) and blood-type antigen secretor status (FUT2). Using transcriptome-wide association and colocalization to infer the effect of gene expression on disease severity, we find evidence that implicates multiple genes—including reduced expression of a membrane flippase (ATP11A), and increased expression of a mucin (MUC1)—in critical disease. Mendelian randomization provides evidence in support of causal roles for myeloid cell adhesion molecules (SELE, ICAM5 and CD209) and the coagulation factor F8, all of which are potentially druggable targets. Our results are broadly consistent with a multi-component model of COVID-19 pathophysiology, in which at least two distinct mechanisms can predispose to life-threatening disease: failure to control viral replication; or an enhanced tendency towards pulmonary inflammation and intravascular coagulation. We show that comparison between cases of critical illness and population controls is highly efficient for the detection of therapeutically relevant mechanisms of disease

Whole-genome sequencing reveals host factors underlying critical COVID-19

Critical COVID-19 is caused by immune-mediated inflammatory lung injury. Host genetic variation influences the development of illness requiring critical care1 or hospitalization2,3,4 after infection with SARS-CoV-2. The GenOMICC (Genetics of Mortality in Critical Care) study enables the comparison of genomes from individuals who are critically ill with those of population controls to find underlying disease mechanisms. Here we use whole-genome sequencing in 7,491 critically ill individuals compared with 48,400 controls to discover and replicate 23 independent variants that significantly predispose to critical COVID-19. We identify 16 new independent associations, including variants within genes that are involved in interferon signalling (IL10RB and PLSCR1), leucocyte differentiation (BCL11A) and blood-type antigen secretor status (FUT2). Using transcriptome-wide association and colocalization to infer the effect of gene expression on disease severity, we find evidence that implicates multiple genes—including reduced expression of a membrane flippase (ATP11A), and increased expression of a mucin (MUC1)—in critical disease. Mendelian randomization provides evidence in support of causal roles for myeloid cell adhesion molecules (SELE, ICAM5 and CD209) and the coagulation factor F8, all of which are potentially druggable targets. Our results are broadly consistent with a multi-component model of COVID-19 pathophysiology, in which at least two distinct mechanisms can predispose to life-threatening disease: failure to control viral replication; or an enhanced tendency towards pulmonary inflammation and intravascular coagulation. We show that comparison between cases of critical illness and population controls is highly efficient for the detection of therapeutically relevant mechanisms of disease

TRAP1 rescues PINK1 loss-of-function phenotypes

PTEN-induced kinase 1 (PINK1) is a serine/threonine kinase that is localized to mitochondria. It protects cells from oxidative stress by suppressing mitochondrial cytochrome c release, thereby preventing cell death. Mutations in Pink1 cause early-onset Parkinson's disease (PD). Consistently, mitochondrial function is impaired in Pink1-linked PD patients and model systems. Previously, in vitro analysis implied that the protective effects of PINK1 depend on phosphorylation of the downstream factor, TNF receptor-associated protein 1 (TRAP1). Furthermore, TRAP1 has been shown to mitigate α-Synuclein-induced toxicity, linking α-Synuclein directly to mitochondrial dysfunction. These data suggest that TRAP1 seems to mediate protective effects on mitochondrial function in pathways that are affected in PD. Here we investigated the potential of TRAP1 to rescue dysfunction induced by either PINK1 or Parkin deficiency in vivo and in vitro. We show that overexpression of human TRAP1 is able to mitigate Pink1 but not parkin loss-of-function phenotypes in Drosophila. In addition, detrimental effects observed after RNAi-mediated silencing of complex I subunits were rescued by TRAP1 in Drosophila. Moreover, TRAP1 was able to rescue mitochondrial fragmentation and dysfunction upon siRNA-induced silencing of Pink1 but not parkin in human neuronal SH-SY5Y cells. Thus, our data suggest a functional role of TRAP1 in maintaining mitochondrial integrity downstream of PINK1 and complex I deficits but parallel to or upstream of Parkin

<i>ND42</i> or <i>sicily</i> overexpression can rescue <i>pink1</i> mutant phenotypes in flies.

<p>Overexpression of two <i>ND42</i> transgenes, <i>ND42</i> and <i>ND42-HA</i>, in a wild type background has no effect on climbing (A) or flight behavior (B). In <i>pink1^B9</i> mutants, climbing (C) and flight ability (D), normalized to control, is significantly rescued by overexpression <i>ND42</i> or <i>sicily</i>. Histograms indicate mean ± s.e.m. (E) Transmission electron microscopy of flight muscle shows partial rescue of mitochondrial disruption. Scale bar  = 1 µm. Overexpression was driven by the ubiquitous driver <i>da-GAL4</i>. Control genotype is <i>da-GAL4</i>/+. Number of animals tested, n>50. * <i>P</i><0.05, *** <i>P</i><0.001, **** <i>P</i><0.0001, One-way ANOVA with Bonferroni correction. Comparisons are with control (A, B) or <i>pink1^B9</i> mutants (C, D).</p

Overexpression of <i>parkin</i> can rescue behavioral phenotypes in <i>pink1</i> mutants but not complex I deficiency.

<p>Analysis of (A) climbing and (B) flight ability in <i>pink1^B9</i> mutants overexpressing <i>parkin</i>. Charts show (C) the ratio of complex I to citrate synthase (CS) activity, and (D) relative ATP levels, normalized to control. Histograms indicate mean ± s.e.m. Overexpression was driven by the ubiquitous driver <i>da-GAL4</i>. Control genotype is <i>da-GAL4</i>/+. * <i>P<</i>0.05, ** <i>P<</i>0.01, *** <i>P<</i>0.001, **** <i>P</i><0.0001, One-way ANOVA with Bonferroni correction. Comparisons are with <i>pink1^B9</i> mutants.</p

A cell based RNAi screen to identify phenocopiers and suppressors of <i>pink1</i> RNAi-induced mitochondrial hyperfusion.

<p>(A) Schematic of the RNAi screen protocol (see <a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1004815#s4" target="_blank">Methods</a> for details). (B) Representative images of <i>Drosophila</i> S2R+ cells for mitochondrial morphology following dsRNA treatment of the indicated genes, stained with MitoTracker Red and imaged live (top row). Fluorescence images are converted to binary (B&W) and inverted to clarify the mitochondrial morphology (bottom row). Numbers represent the designated ‘morphology score’: 1, fragmented; 2, wild type; 3, fused/tubular; 4, hyperfused/clumped. (C) Comparison of morphology score of screen library amplicons in WT and <i>pink1</i> RNAi backgrounds. Solid-line box depicts those amplicons that phenocopy <i>pink1</i> RNAi (box limits: mean ± s.d. <i>pink1</i> control). Dashed-line box depicts amplicons which suppress <i>pink1</i> RNAi-induced fusion back to WT morphology (box limits: mean ± s.d. <i>DsRed</i> control). Controls for fragmentation (<i>Marf)</i> and fusion (<i>Drp1</i>) are shown. Scale bar  = 10 µm.</p

List of genes that suppress <i>pink1</i> dsRNA induced mitochondrial fusion.

<p>List of genes that suppress <i>pink1</i> dsRNA induced mitochondrial fusion.</p

List of genes that phenocopy <i>pink1</i> dsRNA induced mitochondrial fusion.

<p>List of genes that phenocopy <i>pink1</i> dsRNA induced mitochondrial fusion.</p

<i>ND42</i> or <i>sicily</i> overexpression does not rescue <i>parkin</i> mutant phenotypes in flies.

<p>In <i>park²⁵</i> mutants, climbing (A) and flight ability (B), normalized to control, is not rescued by <i>ND42</i> or <i>sicily</i> overexpression. Histograms indicate mean ± s.e.m. (C) Transmission electron microscopy of flight muscle shows widespread disruption of mitochondrial integrity. Scale bar  = 1 µm. Overexpression was driven by the ubiquitous driver <i>da-GAL4</i>. Control genotype is <i>da-GAL4</i>/+. Number of animals tested, n>50. * <i>P</i><0.05, *** <i>P</i><0.001, **** <i>P</i><0.0001, One-way ANOVA with Bonferroni correction. Comparisons are with <i>pink1^B9</i> mutants.</p