Accessory subunits are integral for assembly and function of human mitochondrial complex I
David A. Stroud,Elliot Surgenor,Luke E. Formosa,Luke E. Formosa,Boris Reljic,Ann E. Frazier,Marris G. Dibley,Laura D. Osellame,Tegan Stait,Traude H. Beilharz,David R. Thorburn,David R. Thorburn,Agus Salim,Michael T. Ryan +13 more
TLDR
It is shown that 25 subunits are strictly required for assembly of a functional complex and 1 subunit is essential for cell viability, and coupling gene-editing technology with proteomics represents a powerful tool for dissecting large multi-subunit complexes and enables the study of complex dysfunction at a cellular level.Abstract:
Complex I (NADH:ubiquinone oxidoreductase) is the first enzyme of the mitochondrial respiratory chain and is composed of 45 subunits in humans, making it one of the largest known multi-subunit membrane protein complexes. Complex I exists in supercomplex forms with respiratory chain complexes III and IV, which are together required for the generation of a transmembrane proton gradient used for the synthesis of ATP. Complex I is also a major source of damaging reactive oxygen species and its dysfunction is associated with mitochondrial disease, Parkinson's disease and ageing. Bacterial and human complex I share 14 core subunits that are essential for enzymatic function; however, the role and necessity of the remaining 31 human accessory subunits is unclear. The incorporation of accessory subunits into the complex increases the cellular energetic cost and has necessitated the involvement of numerous assembly factors for complex I biogenesis. Here we use gene editing to generate human knockout cell lines for each accessory subunit. We show that 25 subunits are strictly required for assembly of a functional complex and 1 subunit is essential for cell viability. Quantitative proteomic analysis of cell lines revealed that loss of each subunit affects the stability of other subunits residing in the same structural module. Analysis of proteomic changes after the loss of specific modules revealed that ATP5SL and DMAC1 are required for assembly of the distal portion of the complex I membrane arm. Our results demonstrate the broad importance of accessory subunits in the structure and function of human complex I. Coupling gene-editing technology with proteomics represents a powerful tool for dissecting large multi-subunit complexes and enables the study of complex dysfunction at a cellular level.read more
Citations
More filters
Journal ArticleDOI
A SARS-CoV-2 protein interaction map reveals targets for drug repurposing.
David E. Gordon,Gwendolyn M. Jang,Mehdi Bouhaddou,Jiewei Xu,Kirsten Obernier,Kris M. White,Matthew J. O’Meara,Veronica V. Rezelj,Jeffrey Z. Guo,Danielle L. Swaney,Tia A. Tummino,Ruth Hüttenhain,Robyn M. Kaake,Alicia L. Richards,Beril Tutuncuoglu,Helene Foussard,Jyoti Batra,Kelsey M. Haas,Maya Modak,Minkyu Kim,Paige Haas,Benjamin J. Polacco,Hannes Braberg,Jacqueline M. Fabius,Manon Eckhardt,Margaret Soucheray,Melanie J. Bennett,Merve Cakir,Michael McGregor,Qiongyu Li,Bjoern Meyer,Ferdinand Roesch,Thomas Vallet,Alice Mac Kain,Lisa Miorin,Elena Moreno,Zun Zar Chi Naing,Yuan Zhou,Shiming Peng,Ying Shi,Ziyang Zhang,Wenqi Shen,Ilsa T Kirby,James E. Melnyk,John S. Chorba,Kevin Lou,Shizhong Dai,Inigo Barrio-Hernandez,Danish Memon,Claudia Hernandez-Armenta,Jiankun Lyu,Christopher J.P. Mathy,Tina Perica,Kala Bharath Pilla,Sai J. Ganesan,Daniel J. Saltzberg,Rakesh Ramachandran,Xi Liu,Sara Brin Rosenthal,Lorenzo Calviello,Srivats Venkataramanan,Jose Liboy-Lugo,Yizhu Lin,Xi Ping Huang,Yongfeng Liu,Stephanie A. Wankowicz,Markus Bohn,Maliheh Safari,Fatima S. Ugur,Cassandra Koh,Nastaran Sadat Savar,Quang Dinh Tran,Djoshkun Shengjuler,Sabrina J. Fletcher,Michael C. O’Neal,Yiming Cai,Jason C.J. Chang,David J. Broadhurst,Saker Klippsten,Phillip P. Sharp,Nicole A. Wenzell,Duygu Kuzuoğlu-Öztürk,Hao-Yuan Wang,Raphael Trenker,Janet M. Young,Devin A. Cavero,Devin A. Cavero,Joseph Hiatt,Joseph Hiatt,Theodore L. Roth,Ujjwal Rathore,Ujjwal Rathore,Advait Subramanian,Julia Noack,Mathieu Hubert,Robert M. Stroud,Alan D. Frankel,Oren S. Rosenberg,Kliment A. Verba,David A. Agard,Melanie Ott,Michael Emerman,Natalia Jura,Mark von Zastrow,Eric Verdin,Eric Verdin,Alan Ashworth,Olivier Schwartz,Christophe d'Enfert,Shaeri Mukherjee,Matthew P. Jacobson,Harmit S. Malik,Danica Galonić Fujimori,Trey Ideker,Charles S. Craik,Stephen N. Floor,James S. Fraser,John D. Gross,Andrej Sali,Bryan L. Roth,Davide Ruggero,Jack Taunton,Tanja Kortemme,Pedro Beltrao,Marco Vignuzzi,Adolfo García-Sastre,Kevan M. Shokat,Brian K. Shoichet,Nevan J. Krogan +128 more
TL;DR: A human–SARS-CoV-2 protein interaction map highlights cellular processes that are hijacked by the virus and that can be targeted by existing drugs, including inhibitors of mRNA translation and predicted regulators of the sigma receptors.
Journal ArticleDOI
BAK/BAX macropores facilitate mitochondrial herniation and mtDNA efflux during apoptosis
Kate McArthur,Kate McArthur,Kate McArthur,Lachlan Whitehead,Lachlan Whitehead,John M. Heddleston,Lucy Li,Benjamin S. Padman,Viola Oorschot,Niall D. Geoghegan,Niall D. Geoghegan,Stephane Chappaz,Stephane Chappaz,Stephane Chappaz,Sophia Davidson,Hui San Chin,Rachael M. Lane,Marija Dramicanin,Marija Dramicanin,Tahnee L. Saunders,Canny Sugiana,Romina Lessene,Romina Lessene,Laura D. Osellame,Teng Leong Chew,Grant Dewson,Grant Dewson,Michael Lazarou,Georg Ramm,Guillaume Lessene,Guillaume Lessene,Michael T. Ryan,Kelly L. Rogers,Kelly L. Rogers,Mark F. van Delft,Mark F. van Delft,Benjamin T. Kile,Benjamin T. Kile,Benjamin T. Kile +38 more
TL;DR: This study provides a mechanistic description of mtDNA release from mitochondria during apoptosis, and suggests that mtDNA is found outside the mitochondria—and, indeed, outside the cell—in a wide range of circumstances.
Journal ArticleDOI
Architecture of Human Mitochondrial Respiratory Megacomplex I2III2IV2.
TL;DR: The structure of the human respiratory chain megacomplex with 140 subunits and a subset of associated cofactors is examined using cryo-electron microscopy to reveal the precise assignment of individual subunits of human CI and CIII and enables future in-depth analysis of the electron transport chain as a whole.
Journal ArticleDOI
The Enigma of the Respiratory Chain Supercomplex
TL;DR: Data and hypotheses on the structures, roles, and assembly of respiratory-chain supercomplexes are evaluated and a future research agenda is proposed to address unanswered questions.
Journal ArticleDOI
Clarifying the supercomplex: the higher-order organization of the mitochondrial electron transport chain
James A. Letts,Leonid A. Sazanov +1 more
TL;DR: The oxidative phosphorylation electron transport chain (OXPHOS-ETC) of the inner mitochondrial membrane is composed of five large protein complexes, named CI–CV, and recent work has shed light on the assembly and function of the SCs.
References
More filters
Journal ArticleDOI
NIH Image to ImageJ: 25 years of image analysis
TL;DR: The origins, challenges and solutions of NIH Image and ImageJ software are discussed, and how their history can serve to advise and inform other software projects.
Journal ArticleDOI
MaxQuant enables high peptide identification rates, individualized p.p.b.-range mass accuracies and proteome-wide protein quantification.
Jiirgen Cox,Matthias Mann +1 more
TL;DR: MaxQuant, an integrated suite of algorithms specifically developed for high-resolution, quantitative MS data, detects peaks, isotope clusters and stable amino acid isotope–labeled (SILAC) peptide pairs as three-dimensional objects in m/z, elution time and signal intensity space and achieves mass accuracy in the p.p.b. range.
Journal ArticleDOI
Significance analysis of microarrays applied to the ionizing radiation response
TL;DR: A method that assigns a score to each gene on the basis of change in gene expression relative to the standard deviation of repeated measurements is described, suggesting that this repair pathway for UV-damaged DNA might play a previously unrecognized role in repairing DNA damaged by ionizing radiation.
Journal ArticleDOI
Tricine-sodium dodecyl sulfate-polyacrylamide gel electrophoresis for the separation of proteins in the range from 1 to 100 kDa.
H. Schägger,G. von Jagow +1 more
TL;DR: A discontinuous sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) system for the separation of proteins in the range from 1 to 100 kDa is described, and the omission of glycine and urea prevents disturbances which might occur in the course of subsequent amino acid sequencing.
Journal ArticleDOI
Genome engineering using the CRISPR-Cas9 system
F. Ann Ran,Patrick D. Hsu,Jason Wright,Vineeta Agarwala,Vineeta Agarwala,David A. Scott,Feng Zhang +6 more
TL;DR: A set of tools for Cas9-mediated genome editing via nonhomologous end joining (NHEJ) or homology-directed repair (HDR) in mammalian cells, as well as generation of modified cell lines for downstream functional studies are described.
Related Papers (5)
The architecture of respiratory supercomplexes
Supercomplex Assembly Determines Electron Flux in the Mitochondrial Electron Transport Chain
Esther Lapuente-Brun,Esther Lapuente-Brun,Raquel Moreno-Loshuertos,Rebeca Acín-Pérez,Ana Latorre-Pellicer,Carmen Colás,Eduardo Balsa,Eduardo Balsa,Ester Perales-Clemente,Pedro M. Quirós,Enrique Calvo,M. A. C. Rodríguez-Hernández,Plácido Navas,Raquel Cruz,Angel Carracedo,Carlos López-Otín,Acisclo Pérez-Martos,Patricio Fernández-Silva,Erika Fernandez-Vizarra,José Antonio Enríquez,José Antonio Enríquez +20 more