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Cell Nov 2018Mammalian SWI/SNF (mSWI/SNF) ATP-dependent chromatin remodeling complexes are multi-subunit molecular machines that play vital roles in regulating genomic architecture...
Mammalian SWI/SNF (mSWI/SNF) ATP-dependent chromatin remodeling complexes are multi-subunit molecular machines that play vital roles in regulating genomic architecture and are frequently disrupted in human cancer and developmental disorders. To date, the modular organization and pathways of assembly of these chromatin regulators remain unknown, presenting a major barrier to structural and functional determination. Here, we elucidate the architecture and assembly pathway across three classes of mSWI/SNF complexes-canonical BRG1/BRM-associated factor (BAF), polybromo-associated BAF (PBAF), and newly defined ncBAF complexes-and define the requirement of each subunit for complex formation and stability. Using affinity purification of endogenous complexes from mammalian and Drosophila cells coupled with cross-linking mass spectrometry (CX-MS) and mutagenesis, we uncover three distinct and evolutionarily conserved modules, their organization, and the temporal incorporation of these modules into each complete mSWI/SNF complex class. Finally, we map human disease-associated mutations within subunits and modules, defining specific topological regions that are affected upon subunit perturbation.
Topics: Animals; Chromatin; Chromatin Assembly and Disassembly; Chromosomal Proteins, Non-Histone; Drosophila; Gene Knockout Techniques; HEK293 Cells; Humans; Mass Spectrometry; Mutagenesis; Protein Subunits; Recombinant Proteins; Transcription Factors
PubMed: 30343899
DOI: 10.1016/j.cell.2018.09.032 -
Neuropharmacology Oct 2020Nicotine is a highly addictive drug found in tobacco that drives its continued use despite the harmful consequences. The initiation of nicotine abuse involves the... (Review)
Review
Nicotine is a highly addictive drug found in tobacco that drives its continued use despite the harmful consequences. The initiation of nicotine abuse involves the mesolimbic dopamine system, which contributes to the rewarding sensory stimuli and associative learning processes in the beginning stages of addiction. Nicotine binds to neuronal nicotinic acetylcholine receptors (nAChRs), which come in a diverse collection of subtypes. The nAChRs that contain the α4 and β2 subunits, often in combination with the α6 subunit, are particularly important for nicotine's ability to increase midbrain dopamine neuron firing rates and phasic burst firing. Chronic nicotine exposure results in numerous neuroadaptations, including the upregulation of particular nAChR subtypes associated with long-term desensitization of the receptors. When nicotine is no longer present, for example during attempts to quit smoking, a withdrawal syndrome develops. The expression of physical withdrawal symptoms depends mainly on the α2, α3, α5, and β4 nicotinic subunits in the epithalamic habenular complex and its target regions. Thus, nicotine affects diverse neural systems and an array of nAChR subtypes to mediate the overall addiction process. This article is part of the special issue on 'Contemporary Advances in Nicotine Neuropharmacology'.
Topics: Animals; Brain; Humans; Nicotine; Nicotinic Agonists; Nicotinic Antagonists; Protein Subunits; Receptors, Nicotinic; Tobacco Use Disorder
PubMed: 32738308
DOI: 10.1016/j.neuropharm.2020.108256 -
Nature Jul 2022Mechanistic target of rapamycin complex 1 (mTORC1) controls growth by regulating anabolic and catabolic processes in response to environmental cues, including nutrients....
Mechanistic target of rapamycin complex 1 (mTORC1) controls growth by regulating anabolic and catabolic processes in response to environmental cues, including nutrients. Amino acids signal to mTORC1 through the Rag GTPases, which are regulated by several protein complexes, including GATOR1 and GATOR2. GATOR2, which has five components (WDR24, MIOS, WDR59, SEH1L and SEC13), is required for amino acids to activate mTORC1 and interacts with the leucine and arginine sensors SESN2 and CASTOR1, respectively. Despite this central role in nutrient sensing, GATOR2 remains mysterious as its subunit stoichiometry, biochemical function and structure are unknown. Here we used cryo-electron microscopy to determine the three-dimensional structure of the human GATOR2 complex. We found that GATOR2 adopts a large (1.1 MDa), two-fold symmetric, cage-like architecture, supported by an octagonal scaffold and decorated with eight pairs of WD40 β-propellers. The scaffold contains two WDR24, four MIOS and two WDR59 subunits circularized via two distinct types of junction involving non-catalytic RING domains and α-solenoids. Integration of SEH1L and SEC13 into the scaffold through β-propeller blade donation stabilizes the GATOR2 complex and reveals an evolutionary relationship to the nuclear pore and membrane-coating complexes. The scaffold orients the WD40 β-propeller dimers, which mediate interactions with SESN2, CASTOR1 and GATOR1. Our work reveals the structure of an essential component of the nutrient-sensing machinery and provides a foundation for understanding the function of GATOR2 within the mTORC1 pathway.
Topics: Humans; Amino Acids; Arginine; Carrier Proteins; Cryoelectron Microscopy; Leucine; Mechanistic Target of Rapamycin Complex 1; Multiprotein Complexes; Nutrients; Protein Domains; Protein Subunits; Proteins
PubMed: 35831510
DOI: 10.1038/s41586-022-04939-z -
Cellular and Molecular Life Sciences :... Dec 2014In eukaryotic cells, proteasomes are highly conserved protease complexes and eliminate unwanted proteins which are marked by poly-ubiquitin chains for degradation. The... (Review)
Review
In eukaryotic cells, proteasomes are highly conserved protease complexes and eliminate unwanted proteins which are marked by poly-ubiquitin chains for degradation. The 26S proteasome consists of the proteolytic core particle, the 20S proteasome, and the 19S regulatory particle, which are composed of 14 and 19 different subunits, respectively. Proteasomes are the second-most abundant protein complexes and are continuously assembled from inactive precursor complexes in proliferating cells. The modular concept of proteasome assembly was recognized in prokaryotic ancestors and applies to eukaryotic successors. The efficiency and fidelity of eukaryotic proteasome assembly is achieved by several proteasome-dedicated chaperones that initiate subunit incorporation and control the quality of proteasome assemblies by transiently interacting with proteasome precursors. It is important to understand the mechanism of proteasome assembly as the proteasome has key functions in the turnover of short-lived proteins regulating diverse biological processes.
Topics: Animals; Humans; Models, Biological; Models, Molecular; Proteasome Endopeptidase Complex; Protein Binding; Protein Conformation; Protein Subunits
PubMed: 25107634
DOI: 10.1007/s00018-014-1699-8 -
Biomolecules Jan 2019The phosphatidylinositol 3-kinase (PI3K) pathway plays a central role in the regulation of cell signaling, proliferation, survival, migration and vesicle trafficking in... (Review)
Review
The phosphatidylinositol 3-kinase (PI3K) pathway plays a central role in the regulation of cell signaling, proliferation, survival, migration and vesicle trafficking in normal cells and is frequently deregulated in many cancers. The p85α protein is the most characterized regulatory subunit of the class IA PI3Ks, best known for its regulation of the p110-PI3K catalytic subunit. In this review, we will discuss the impact of p85α mutations or alterations in expression levels on the proteins p85α is known to bind and regulate. We will focus on alterations within the N-terminal half of p85α that primarily regulate Rab5 and some members of the Rho-family of GTPases, as well as those that regulate PTEN (phosphatase and tensin homologue deleted on chromosome 10), the enzyme that directly counteracts PI3K signaling. We highlight recent data, mapping the interaction surfaces of the PTEN⁻p85α breakpoint cluster region homology (BH) domain, which sheds new light on key residues in both proteins. As a multifunctional protein that binds and regulates many different proteins, p85α mutations at different sites have different impacts in cancer and would necessarily require distinct treatment strategies to be effective.
Topics: Class Ia Phosphatidylinositol 3-Kinase; Humans; Mutation; Neoplasms; PTEN Phosphohydrolase; Protein Domains; Protein Subunits; rab5 GTP-Binding Proteins
PubMed: 30650664
DOI: 10.3390/biom9010029 -
Frontiers in Immunology 2023Tuberculosis (TB), also known as the "White Plague", is caused by (). Before the COVID-19 epidemic, TB had the highest mortality rate of any single infectious disease.... (Review)
Review
Tuberculosis (TB), also known as the "White Plague", is caused by (). Before the COVID-19 epidemic, TB had the highest mortality rate of any single infectious disease. Vaccination is considered one of the most effective strategies for controlling TB. Despite the limitations of the Bacille Calmette-Guérin (BCG) vaccine in terms of protection against TB among adults, it is currently the only licensed TB vaccine. Recently, with the evolution of bioinformatics and structural biology techniques to screen and optimize protective antigens of , the tremendous potential of protein subunit vaccines is being exploited. Multistage subunit vaccines obtained by fusing immunodominant antigens from different stages of TB infection are being used both to prevent and to treat TB. Additionally, the development of novel adjuvants is compensating for weaknesses of immunogenicity, which is conducive to the flourishing of subunit vaccines. With advances in the development of animal models, preclinical vaccine protection assessments are becoming increasingly accurate. This review summarizes progress in the research of protein subunit TB vaccines during the past decades to facilitate the further optimization of protein subunit vaccines that may eradicate TB.
Topics: Animals; Protein Subunits; COVID-19; Vaccines, Subunit; Tuberculosis; Tuberculosis Vaccines; BCG Vaccine
PubMed: 37654500
DOI: 10.3389/fimmu.2023.1238586 -
Frontiers in Bioscience (Landmark... Jan 2017V-ATPases are ATP-driven proton pumps present in both intracellular and cell surface membranes of eukaryotes that function in many normal and disease processes.... (Review)
Review
V-ATPases are ATP-driven proton pumps present in both intracellular and cell surface membranes of eukaryotes that function in many normal and disease processes. V-ATPases are large, multi-subunit complexes composed of a peripheral domain (V) that hydrolyzes ATP and a membrane integral domain (V) that translocates protons. Because of the diversity of their functions, V-ATPase activity is controlled by a number of mechanisms. Regulated assembly of the V and V domains rapidly modulates V-ATPase activity in response to a variety of cues, including nutrient availability, growth factor stimulation and cellular differentiation. Considerable information has recently emerged concerning the cellular signaling pathways controlling regulated assembly. Acid secretion by epithelial cells in the kidney and epididymus is controlled by regulated trafficking of V-ATPases to the cell surface. Isoforms of subunit a of the V domain both control trafficking of V-ATPases to distinct cellular membranes and confer properties to the resultant complexes that help account for differences in pH between cellular compartments. Finally, differential expression of genes encoding V-ATPases subunits occurs in a number of contexts, including cancer.
Topics: Animals; Humans; Insect Proteins; Mammals; Models, Molecular; Protein Multimerization; Protein Subunits; Protein Transport; Saccharomyces cerevisiae Proteins; Vacuolar Proton-Translocating ATPases
PubMed: 27814636
DOI: 10.2741/4506 -
The Biochemical Journal Dec 2018Reversible phosphorylation of proteins is a post-translational modification that regulates all aspect of life through the antagonistic action of kinases and... (Review)
Review
Reversible phosphorylation of proteins is a post-translational modification that regulates all aspect of life through the antagonistic action of kinases and phosphatases. Protein kinases are well characterized, but protein phosphatases have been relatively neglected. Protein phosphatase 1 (PP1) catalyzes the dephosphorylation of a major fraction of phospho-serines and phospho-threonines in cells and thereby controls a broad range of cellular processes. In this review, I will discuss how phosphatases were discovered, how the view that they were unselective emerged and how recent findings have revealed their exquisite selectivity. Unlike kinases, PP1 phosphatases are obligatory heteromers composed of a catalytic subunit bound to one (or two) non-catalytic subunit(s). Based on an in-depth study of two holophosphatases, I propose the following: selective dephosphorylation depends on the assembly of two components, the catalytic subunit and the non-catalytic subunit, which serves as a high-affinity substrate receptor. Because functional complementation of the two modules is required to produce a selective holophosphatase, one can consider that they are split enzymes. The non-catalytic subunit was often referred to as a regulatory subunit, but it is, in fact, an essential component of the holoenzyme. In this model, a phosphatase and its array of mostly orphan substrate receptors constitute the split protein phosphatase system. The set of potentially generalizable principles outlined in this review may facilitate the study of these poorly understood enzymes and the identification of their physiological substrates.
Topics: Animals; Enzyme Inhibitors; Humans; Phosphoprotein Phosphatases; Phosphorylation; Protein Multimerization; Protein Phosphatase 1; Protein Phosphatase 2; Protein Subunits; Substrate Specificity
PubMed: 30523060
DOI: 10.1042/BCJ20170726 -
Current Opinion in Structural Biology Oct 2016With the convergence of breakthroughs in structural biology, specifically breaking the resolution barriers in cryo-electron microscopy and with continuing developments... (Review)
Review
With the convergence of breakthroughs in structural biology, specifically breaking the resolution barriers in cryo-electron microscopy and with continuing developments in crystallography, novel interfaces with other biophysical methods are emerging. Here we consider how mass spectrometry can inform these techniques by providing unambiguous definition of subunit stoichiometry. Moreover recent developments that increase mass spectral resolution enable molecular details to be ascribed to unassigned density within high-resolution maps of membrane and soluble protein complexes. Importantly we also show how developments in mass spectrometry can define optimal solution conditions to guide downstream structure determination, particularly of challenging biomolecules that refuse to crystallise.
Topics: Biology; Crystallography; DNA; Humans; Mass Spectrometry; Protein Subunits; RNA
PubMed: 27721169
DOI: 10.1016/j.sbi.2016.09.008 -
Biomolecules Apr 2020Asparagine-linked glycosylation, also known as -linked glycosylation is an essential and highly conserved post-translational protein modification that occurs in all... (Review)
Review
Asparagine-linked glycosylation, also known as -linked glycosylation is an essential and highly conserved post-translational protein modification that occurs in all three domains of life. This modification is essential for specific molecular recognition, protein folding, sorting in the endoplasmic reticulum, cell-cell communication, and stability. Defects in -linked glycosylation results in a class of inherited diseases known as congenital disorders of glycosylation (CDG). -linked glycosylation occurs in the endoplasmic reticulum (ER) lumen by a membrane associated enzyme complex called the oligosaccharyltransferase (OST). In the central step of this reaction, an oligosaccharide group is transferred from a lipid-linked dolichol pyrophosphate donor to the acceptor substrate, the side chain of a specific asparagine residue of a newly synthesized protein. The prokaryotic OST enzyme consists of a single polypeptide chain, also known as single subunit OST or ssOST. In contrast, the eukaryotic OST is a complex of multiple non-identical subunits. In this review, we will discuss the biochemical and structural characterization of the prokaryotic, yeast, and mammalian OST enzymes. This review explains the most recent high-resolution structures of OST determined thus far and the mechanistic implication of -linked glycosylation throughout all domains of life. It has been shown that the ssOST enzyme, AglB protein of the archaeon , and the PglB protein of the bacterium are structurally and functionally similar to the catalytic Stt3 subunit of the eukaryotic OST enzyme complex. Yeast OST enzyme complex contains a single Stt3 subunit, whereas the human OST complex is formed with either STT3A or STT3B, two paralogues of Stt3. Both human OST complexes, OST-A (with STT3A) and OST-B (containing STT3B), are involved in the -linked glycosylation of proteins in the ER. The cryo-EM structures of both human OST-A and OST-B complexes were reported recently. An acceptor peptide and a donor substrate (dolichylphosphate) were observed to be bound to the OST-B complex whereas only dolichylphosphate was bound to the OST-A complex suggesting disparate affinities of two OST complexes for the acceptor substrates. However, we still lack an understanding of the independent role of each eukaryotic OST subunit in -linked glycosylation or in the stabilization of the enzyme complex. Discerning the role of each subunit through structure and function studies will potentially reveal the mechanistic details of -linked glycosylation in higher organisms. Thus, getting an insight into the requirement of multiple non-identical subunits in the -linked glycosylation process in eukaryotes poses an important future goal.
Topics: Amino Acid Sequence; Animals; Catalytic Domain; Glycosylation; Hexosyltransferases; Humans; Isoenzymes; Membrane Proteins; Models, Molecular; Protein Subunits
PubMed: 32316603
DOI: 10.3390/biom10040624