scholarly journals Human Mitochondrial DNA-Protein Complexes Attach to a Cholesterol-Rich Membrane Structure

2015 ◽  
Vol 5 (1) ◽  
Author(s):  
Joachim M. Gerhold ◽  
Şirin Cansiz-Arda ◽  
Madis Lõhmus ◽  
Oskar Engberg ◽  
Aurelio Reyes ◽  
...  
Keyword(s):  
Membrane Structure ◽  
Protein Complexes ◽  
Cholesterol Homeostasis ◽  
Membrane Composition ◽  
Human Mitochondrial Dna ◽  
Associated Proteins ◽  
Mtdna Replication ◽  
Mitochondrial Cholesterol ◽  
Mitochondrial Junctions ◽  
Mitochondrial Lipids

Abstract The helicase Twinkle is indispensable for mtDNA replication in nucleoids. Previously, we showed that Twinkle is tightly membrane-associated even in the absence of mtDNA, which suggests that Twinkle is part of a membrane-attached replication platform. Here we show that this platform is a cholesterol-rich membrane structure. We fractionated mitochondrial membrane preparations on flotation gradients and show that membrane-associated nucleoids accumulate at the top of the gradient. This fraction was shown to be highly enriched in cholesterol, a lipid that is otherwise low abundant in mitochondria. In contrast, more common mitochondrial lipids and abundant inner-membrane associated proteins concentrated in the bottom-half of these gradients. Gene silencing of ATAD3, a protein with proposed functions related to nucleoid and mitochondrial cholesterol homeostasis, modified the distribution of cholesterol and nucleoids in the gradient in an identical fashion. Both cholesterol and ATAD3 were previously shown to be enriched in ER-mitochondrial junctions and we detect nucleoid components in biochemical isolates of these structures. Our data suggest an uncommon membrane composition that accommodates platforms for replicating mtDNA and reconcile apparently disparate functions of ATAD3. We suggest that mtDNA replication platforms are organized in connection with ER-mitochondrial junctions, facilitated by a specialized membrane architecture involving mitochondrial cholesterol.

scholarly journals Erratum: Corrigendum: Human Mitochondrial DNA-Protein Complexes Attach to a Cholesterol-Rich Membrane Structure

2015 ◽  
Vol 5 (1) ◽  
Author(s):  
Joachim M. Gerhold ◽  
Şirin Cansiz-Arda ◽  
Madis Lõhmus ◽  
Oskar Engberg ◽  
Aurelio Reyes ◽  
...  
Keyword(s):  
Mitochondrial Dna ◽  
Membrane Structure ◽  
Protein Complexes ◽  
Human Mitochondrial Dna

scholarly journals Platelet adhesion: Structural and functional diversity of short dystrophin and utrophins in the formation of dystrophinassociated-protein complexes related to actin dynamics

2005 ◽  
Vol 94 (12) ◽  
pp. 1203-1212 ◽  
Author(s):  
Doris Cerecedo ◽  
Dalila Martínez-Rojas ◽  
Oscar Chávez ◽  
Francisco Martínez-Pérez ◽  
Francisco García-Sierra ◽  
...  
Keyword(s):  
Actin Cytoskeleton ◽  
Platelet Adhesion ◽  
Protein Complexes ◽  
Human Platelets ◽  
Actin Dynamics ◽  
Actin Binding ◽  
Dynamic Cell ◽  
Associated Proteins ◽  
Coated Surfaces ◽  
Dynamic Structures

SummaryPlatelets are dynamic cell fragments that modify their shape during activation. Utrophin and dystrophins are minor actin-binding proteins present in muscle and non-muscle cytoskeleton. In the present study, we characterised the pattern of Dp71 isoforms and utrophin gene products by immunoblot in human platelets. Two new dystrophin isoforms were found, Dp71f and Dp71d, as well as the Up71 isoform and the dystrophin-associated proteins, α and β-dystrobrevins. Distribution of Dp71d/Dp71Δ110 m, Up400/Up71 and dystrophin-associated proteins in relation to the actin cytoskeleton was evaluated by confocal microscopy in both resting and platelets adhered on glass. Formation of two dystrophin-associated protein complexes (Dp71d/Dp71Δ110 m ~DAPC and Up400/Up71~DAPC) was demonstrated by co-immunoprecipitation and their distribution in relation to the actin cytoskeleton was characterised during platelet adhesion. The Dp71d/Dp71Δ110 m ~DAPC is maintained mainly at the granulomere and is associated with dynamic structures during activation by adhesion to thrombin-coated surfaces. Participation of both Dp71d/Dp71Δ110 m ~DAPC and Up400/Up71~DAPC in the biological roles of the platelets is discussed.

scholarly journals Current Analytical Strategies in Studying Chromatin-Associated-Proteome (Chromatome)

Molecules ◽  
2021 ◽  
Vol 26 (21) ◽  
pp. 6694
Author(s):  
Niamat Khan ◽  
Sidra Shahid ◽  
Abdul R. Asif
Keyword(s):  
Drug Targets ◽  
Protein Complexes ◽  
Dynamic Structure ◽  
Associated Diseases ◽  
Specific Complex ◽  
Analytical Strategies ◽  
Associated Proteins ◽  
Therapeutic Molecules ◽  
Unique Nature ◽  
Mass Spectrometric Identification

Chromatin is a dynamic structure comprising of DNA and proteins. Its unique nature not only help to pack the DNA tightly within the cell but also is pivotal in regulating gene expression DNA replication. Furthermore it also protects the DNA from being damaged. Various proteins are involved in making a specific complex within a chromatin and the knowledge about these interacting partners is helpful to enhance our understanding about the pathophysiology of various chromatin associated diseases. Moreover, it could also help us to identify new drug targets and design more effective remedies. Due to the existence of chromatin in different forms under various physiological conditions it is hard to develop a single strategy to study chromatin associated proteins under all conditions. In our current review, we tried to provide an overview and comparative analysis of the strategies currently adopted to capture the DNA bounded protein complexes and their mass spectrometric identification and quantification. Precise information about the protein partners and their function in the DNA-protein complexes is crucial to design new and more effective therapeutic molecules against chromatin associated diseases.

scholarly journals Systematic discovery of endogenous human ribonucleoprotein complexes

2018 ◽  
Author(s):  
Anna L. Mallam ◽  
Wisath Sae-Lee ◽  
Jeffrey M. Schaub ◽  
Fan Tu ◽  
Anna Battenhouse ◽  
...  
Keyword(s):  
Rna Binding ◽  
Rna Binding Proteins ◽  
Protein Complexes ◽  
Embryonic Stem ◽  
Human Protein ◽  
Disease States ◽  
Ribonucleoprotein Complexes ◽  
Associated Proteins ◽  
Domains Of Life ◽  
Factor C

AbstractRNA-binding proteins (RBPs) play essential roles in biology and are frequently associated with human disease. While recent studies have systematically identified individual RBPs, their higher order assembly intoRibonucleoprotein (RNP) complexes has not been systematically investigated. Here, we describe a proteomics method for systematic identification of RNP complexes in human cells. We identify 1,428 protein complexes that associate with RNA, indicating that over 20% of known human protein complexes contain RNA. To explore the role of RNA in the assembly of each complex, we identify complexes that dissociate, change composition, or form stable protein-only complexes in the absence of RNA. Importantly, these data also provide specific novel insights into the function of well-studied protein complexes not previously known to associate with RNA, including replication factor C (RFC) and cytokinetic centralspindlin complex. Finally, we use our method to systematically identify cell-type specific RNA-associated proteins in mouse embryonic stem cells. We distribute these data as a resource, rna.MAP (rna.proteincomplexes.org) which provides a comprehensive dataset for the study of RNA-associated protein complexes. Our system thus provides a novel methodology for further explorations across human tissues and disease states, as well as throughout all domains of life.SummaryAn exploration of human protein complexes in the presence and absence of RNA reveals endogenous ribonucleoprotein complexes

scholarly journals Regulation of Clathrin-Mediated Endocytosis

2018 ◽  
Vol 87 (1) ◽  
pp. 871-896 ◽  
Author(s):  
Marcel Mettlen ◽  
Ping-Hung Chen ◽  
Saipraveen Srinivasan ◽  
Gaudenz Danuser ◽  
Sandra L. Schmid
Keyword(s):  
Conformational Changes ◽  
Posttranslational Modifications ◽  
Mammalian Cells ◽  
Environmental Changes ◽  
Protein Complexes ◽  
Adaptor Protein ◽  
Endocytic Pathway ◽  
Coat Proteins ◽  
Membrane Composition ◽  
Coated Pits

Clathrin-mediated endocytosis (CME) is the major endocytic pathway in mammalian cells. It is responsible for the uptake of transmembrane receptors and transporters, for remodeling plasma membrane composition in response to environmental changes, and for regulating cell surface signaling. CME occurs via the assembly and maturation of clathrin-coated pits that concentrate cargo as they invaginate and pinch off to form clathrin-coated vesicles. In addition to the major coat proteins, clathrin triskelia and adaptor protein complexes, CME requires a myriad of endocytic accessory proteins and phosphatidylinositol lipids. CME is regulated at multiple steps—initiation, cargo selection, maturation, and fission—and is monitored by an endocytic checkpoint that induces disassembly of defective pits. Regulation occurs via posttranslational modifications, allosteric conformational changes, and isoform and splice-variant differences among components of the CME machinery, including the GTPase dynamin. This review summarizes recent findings on the regulation of CME and the evolution of this complex process.

scholarly journals DRP1 regulates endoplasmic reticulum sheets to control mitochondrial DNA replication and segregation

2021 ◽  
Author(s):  
Hema Saranya Ilamathi ◽  
Sara Benhammouda ◽  
Justine Desrochers-Goyette ◽  
Matthew A Lines ◽  
Marc Germain
Keyword(s):  
Endoplasmic Reticulum ◽  
Mitochondrial Dna ◽  
Protein Complexes ◽  
Mitochondrial Dynamics ◽  
Mitochondrial Fission ◽  
Mitochondrial Diseases ◽  
Contact Sites ◽  
Transport Chain ◽  
Essential Components ◽  
Mtdna Replication

Mitochondria are multi-faceted organelles crucial for cellular homeostasis that contain their own genome. Mitochondrial DNA (mtDNA) codes for several essential components of the electron transport chain, and mtDNA maintenance defects lead to mitochondrial diseases. mtDNA replication occurs at endoplasmic reticulum (ER)-mitochondria contact sites and is regulated by mitochondrial dynamics. Specifically, mitochondrial fusion is essential for mtDNA maintenance. In contrast, while loss of mitochondrial fission causes the aggregation of nucleoids (mtDNA-protein complexes), its role in nucleoid distribution remains unclear. Here, we show that the mitochondrial fission protein DRP1 regulates nucleoid segregation by altering ER sheets, the ER structure associated with protein synthesis. Specifically, DRP1 loss or mutation leads to altered ER sheets that physically interact with mitobulbs, mitochondrial structures containing aggregated nucleoids. Importantly, nucleoid distribution and mtDNA replication were rescued by expressing the ER sheet protein CLIMP63. Thus, our work identifies a novel mechanism by which DRP1 regulates mtDNA replication and distribution.

scholarly journals Occurrence of cross-resistance and β-lactam seesaw effect in glycopeptide-, lipopeptide- and lipoglycopeptide-resistant MRSA correlates with membrane phosphatidylglycerol levels

2020 ◽  
Vol 75 (5) ◽  
pp. 1182-1186 ◽  
Author(s):  
Kelly M Hines ◽  
Tianwei Shen ◽  
Nathaniel K Ashford ◽  
Adam Waalkes ◽  
Kelsi Penewit ◽  
...  
Keyword(s):  
Cell Wall ◽  
Susceptibility Testing ◽  
Membrane Lipid ◽  
Cross Resistance ◽  
Membrane Composition ◽  
Membrane Lipid Composition ◽  
Usa300 Strain ◽  
Associated Proteins ◽  
The Relationship

Abstract Background Glycopeptides (GPs), lipopeptides (LPs) and lipoglycopeptides (LGPs) are related antimicrobials important for the management of invasive MRSA infections. Cross-resistance among these antibiotics in MRSA is well documented, as is the observation that susceptibility of MRSA to β-lactams increases as susceptibility to GPs and LPs decreases (i.e. the seesaw effect). Efforts to understand the relationship between GP/LP/LGP cross-resistance and the seesaw effect have focused on the PBPs, but the role of lipid metabolism has not been investigated. Objectives Since the cell membrane is structurally and metabolically integrated with the cell wall and anchors associated proteins, including PBPs, we examined the relationship between membrane lipid composition and the phenomena of cross-resistance among GPs/LPs/LGPs and the β-lactam seesaw effect. Methods We selected for daptomycin, vancomycin and dalbavancin resistance using the USA300 strain JE2 and evaluated the resulting mutants by WGS, MS-based lipidomics and antimicrobial susceptibility testing to assess the relationship between membrane composition, cross-resistance, and the seesaw effect. Results We observed cross-resistance to GPs/LPs/LGPs among the selected strains and the seesaw effect against various β-lactams, depending on the PBP targets of the particular β-lactam. We found that modification of membrane composition occurs not only in daptomycin-selected strains, but also vancomycin- and dalbavancin-selected strains. Significantly, we observed that the abundance of most phosphatidylglycerols positively correlates with MICs of GPs/LPs/LGPs and negatively correlates with the MICs of β-lactams. Conclusions These studies demonstrate a major association between membrane remodelling, cross-resistance and the seesaw effect.

scholarly journals A novel landscape of nuclear human CDK2 substrates revealed by in situ phosphorylation

2020 ◽  
Vol 6 (16) ◽  
pp. eaaz9899
Author(s):  
Yong Chi ◽  
John H. Carter ◽  
Jherek Swanger ◽  
Alexander V. Mazin ◽  
Robert L. Moritz ◽  
...  
Keyword(s):  
Cell Division ◽  
Protein Complexes ◽  
Nuclear Architecture ◽  
Cyclin Dependent Kinase ◽  
Oncogenic Signaling ◽  
Cellular Processes ◽  
Validation Rate ◽  
Associated Proteins ◽  
Epigenetic Regulators

Cyclin-dependent kinase 2 (CDK2) controls cell division and is central to oncogenic signaling. We used an “in situ” approach to identify CDK2 substrates within nuclei isolated from cells expressing CDK2 engineered to use adenosine 5′-triphosphate analogs. We identified 117 candidate substrates, ~40% of which are known CDK substrates. Previously unknown candidates were validated to be CDK2 substrates, including LSD1, DOT1L, and Rad54. The identification of many chromatin-associated proteins may have been facilitated by labeling conditions that preserved nuclear architecture and physiologic CDK2 regulation by endogenous cyclins. Candidate substrates include proteins that regulate histone modifications, chromatin, transcription, and RNA/DNA metabolism. Many of these proteins also coexist in multi-protein complexes, including epigenetic regulators, that may provide new links between cell division and other cellular processes mediated by CDK2. In situ phosphorylation thus revealed candidate substrates with a high validation rate and should be readily applicable to other nuclear kinases.

scholarly journals Mammalian PRC1 Complexes: Compositional Complexity and Diverse Molecular Mechanisms

2020 ◽  
Vol 21 (22) ◽  
pp. 8594
Author(s):  
Zhuangzhuang Geng ◽  
Zhonghua Gao
Keyword(s):  
Molecular Mechanisms ◽  
Protein Complexes ◽  
Ring Finger ◽  
Transcription Activation ◽  
Polycomb Group ◽  
Histone H2a ◽  
Catalytic Core ◽  
Associated Proteins ◽  
Regulation Mechanisms ◽  
Polycomb Repressive Complexes

Polycomb group (PcG) proteins function as vital epigenetic regulators in various biological processes, including pluripotency, development, and carcinogenesis. PcG proteins form multicomponent complexes, and two major types of protein complexes have been identified in mammals to date, Polycomb Repressive Complexes 1 and 2 (PRC1 and PRC2). The PRC1 complexes are composed in a hierarchical manner in which the catalytic core, RING1A/B, exclusively interacts with one of six Polycomb group RING finger (PCGF) proteins. This association with specific PCGF proteins allows for PRC1 to be subdivided into six distinct groups, each with their own unique modes of action arising from the distinct set of associated proteins. Historically, PRC1 was considered to be a transcription repressor that deposited monoubiquitylation of histone H2A at lysine 119 (H2AK119ub1) and compacted local chromatin. More recently, there is increasing evidence that demonstrates the transcription activation role of PRC1. Moreover, studies on the higher-order chromatin structure have revealed a new function for PRC1 in mediating long-range interactions. This provides a different perspective regarding both the transcription activation and repression characteristics of PRC1. This review summarizes new advancements regarding the composition of mammalian PRC1 and accompanying explanations of how diverse PRC1-associated proteins participate in distinct transcription regulation mechanisms.

scholarly journals Reconstitution of functional mRNA-protein complexes in a rabbit reticulocyte cell-free translation system.

1985 ◽  
Vol 5 (2) ◽  
pp. 342-351 ◽  
Author(s):  
J R Greenberg ◽  
E Carroll
Keyword(s):  
Mammalian Cells ◽  
Uv Light ◽  
Protein Complexes ◽  
Micrococcal Nuclease ◽  
Translation System ◽  
Cytoplasmic Process ◽  
Free Translation ◽  
Associated Proteins ◽  
A Cell ◽  
And Function

A variety of evidence suggests that the cytoplasmic mRNA-associated proteins of eucaryotic cells are derived from the cytoplasm and function there, most likely in protein synthesis or some related process. Furthermore, the evidence suggests that protein-free mRNA added to a cell-free translation system should become associated with a set of proteins similar to those associated with mRNA in native polyribosomes. To test this hypothesis, we added deproteinized rabbit reticulocyte mRNA to a homologous cell-free translation system made dependent on exogenous mRNA by treatment with micrococcal nuclease. The resulting reconstituted complexes were irradiated with UV light to cross-link the proteins to mRNA, and the proteins were analyzed by gel electrophoresis. The proteins associated with polyribosomal mRNA in the reconstituted complexes were indistinguishable from those associated with polyribosomal mRNA in intact reticulocytes. Furthermore, reticulocyte mRNA-associated proteins were very similar to those of cultured mammalian cells. The composition of the complexes varied with the translational state of the mRNA; that is, certain proteins present in polyribosomal mRNA-protein complexes were absent or reduced in amount in 40S to 80S complexes and in complexes formed in the absence of translation. However, other proteins, including a 78-kilodalton protein associated with polyadenylate, were present irrespective of translational state, or else they were preferentially associated with untranslated mRNA. These findings are in agreement with previous data suggesting that proteins associated with cytoplasmic mRNA are derived from the cytoplasm and that they function in translation or some other cytoplasmic process, rather than transcription, RNA processing, or transport from the nucleus to the cytoplasm.

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