Low complexity (LC) head domains 92 and 108 residues in length are, respectively, required for assembly of neurofilament light (NFL) and desmin intermediate filaments (IFs). As studied in isolation, ...these IF head domains interconvert between states of conformational disorder and labile, β-strand-enriched polymers. Solid-state NMR (ss-NMR) spectroscopic studies of NFL and desmin head domain polymers reveal spectral patterns consistent with structural order. A combination of intein chemistry and segmental isotope labeling allowed preparation of fully assembled NFL and desmin IFs that could also be studied by ss-NMR. Assembled IFs revealed spectra overlapping with those observed for β-strand-enriched polymers formed from the isolated NFL and desmin head domains. Phosphorylation and disease-causing mutations reciprocally alter NFL and desmin head domain self-association yet commonly impede IF assembly. These observations show how facultative structural assembly of LC domains via labile, β-strand-enriched self-interactions may broadly influence cell morphology.
Yeast ataxin-2, also known as Pbp1, senses the activity state of mitochondria in order to regulate TORC1. A domain of Pbp1 required to adapt cells to mitochondrial activity is of low sequence ...complexity. The low-complexity (LC) domain of Pbp1 forms labile, cross-β polymers that facilitate phase transition of the protein into liquid-like or gel-like states. Phase transition for other LC domains is reliant upon widely distributed aromatic amino acids. In place of tyrosine or phenylalanine residues prototypically used for phase separation, Pbp1 contains 24 similarly disposed methionine residues. Here, we show that the Pbp1 methionine residues are sensitive to hydrogen peroxide (H2O2)-mediated oxidation in vitro and in living cells. Methionine oxidation melts Pbp1 liquid-like droplets in a manner reversed by methionine sulfoxide reductase enzymes. These observations explain how reversible formation of labile polymers by the Pbp1 LC domain enables the protein to function as a sensor of cellular redox state.
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•Phase separation of the yeast ataxin-2 methionine-rich LC domain is sensitive to H2O2•The dissolved droplets are specifically revived by methionine sulfoxide reductases•Methionine to tyrosine variants confer resistance to H2O2in vitro and in cells•Yeast ataxin-2 senses cellular redox state to modulate TORC1 activity
Many proteins that are able to undergo phase transitions contain repeats rich in hydrophobic residues. By contrast, yeast ataxin-2 condensates rely on an abundance of methionines, providing a means for redox-sensitive regulation of its material properties.
Acetate Dependence of Tumors Comerford, Sarah A.; Huang, Zhiguang; Du, Xinlin ...
Cell,
12/2014, Letnik:
159, Številka:
7
Journal Article
Recenzirano
Odprti dostop
Acetyl-CoA represents a central node of carbon metabolism that plays a key role in bioenergetics, cell proliferation, and the regulation of gene expression. Highly glycolytic or hypoxic tumors must ...produce sufficient quantities of this metabolite to support cell growth and survival under nutrient-limiting conditions. Here, we show that the nucleocytosolic acetyl-CoA synthetase enzyme, ACSS2, supplies a key source of acetyl-CoA for tumors by capturing acetate as a carbon source. Despite exhibiting no gross deficits in growth or development, adult mice lacking ACSS2 exhibit a significant reduction in tumor burden in two different models of hepatocellular carcinoma. ACSS2 is expressed in a large proportion of human tumors, and its activity is responsible for the majority of cellular acetate uptake into both lipids and histones. These observations may qualify ACSS2 as a targetable metabolic vulnerability of a wide spectrum of tumors.
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•The acetyl-CoA synthetase ACSS2 is the major enzyme responsible for acetate utilization•Mice lacking ACSS2 exhibit reduced tumor burdens in two mouse models of liver cancer•A significant number of human cancers express ACSS2•ACSS2 might represent a metabolic vulnerability specific to tumors
Mice lacking the acetyl-CoA synthetase enzyme ACSS2 exhibit reduced tumor burdens in two mouse models of liver cancer, suggesting that acetate is a critical metabolite for tumor survival and potentially a targetable cancer vulnerability.
Measurements of the abundance of common metabolites in cultured embryonic stem (ES) cells revealed an unusual state with respect to one-carbon metabolism. These findings led to the discovery of ...copious expression of the gene encoding threonine dehydrogenase (TDH) in ES cells. TDH-mediated catabolism of threonine takes place in mitochondria to generate glycine and acetyl-coenzyme A (CoA), with glycine facilitating one-carbon metabolism via the glycine cleavage system and acetyl-CoA feeding the tricarboxylic acid cycle. Culture media individually deprived of each of the 20 amino acids were applied to ES cells, leading to the discovery that ES cells are critically dependent on one amino acid--threonine. These observations show that ES cells exist in a high-flux backbone metabolic state comparable to that of rapidly growing bacterial cells.
Budding yeast grown under continuous, nutrient-limited conditions exhibit robust, highly periodic cycles in the form of respiratory bursts. Microarray studies reveal that over half of the yeast ...genome is expressed periodically during these metabolic cycles. Genes encoding proteins having a common function exhibit similar temporal expression patterns, and genes specifying functions associated with energy and metabolism tend to be expressed with exceptionally robust periodicity. Essential cellular and metabolic events occur in synchrony with the metabolic cycle, demonstrating that key processes in a simple eukaryotic cell are compartmentalized in time.
Human genetic studies have given evidence of familial, disease-causing mutations in the analogous amino acid residue shared by three related RNA binding proteins causative of three neurological ...diseases. Alteration of aspartic acid residue 290 of hnRNPA2 to valine is believed to predispose patients to multisystem proteinopathy. Mutation of aspartic acid 262 of hnRNPA1 to either valine or asparagine has been linked to either amyotrophic lateral sclerosis or multisystem proteinopathy. Mutation of aspartic acid 378 of hnRNPDL to either asparagine or histidine has been associated with limb girdle muscular dystrophy. All three of these aspartic acid residues map to evolutionarily conserved regions of low-complexity (LC) sequence that may function in states of either intrinsic disorder or labile self-association. Here, we present a combination of solid-state NMR spectroscopy with segmental isotope labeling and electron microscopy on the LC domain of the hnRNPA2 protein. We show that, for both the wild-type protein and the aspartic acid 290-to-valine mutant, labile polymers are formed in which the LC domain associates into an in-register cross-β conformation. Aspartic acid 290 is shown to be charged at physiological pH and immobilized within the polymer core. Polymers of the aspartic acid 290-to-valine mutant are thermodynamically more stable than wild-type polymers. These observations give evidence that removal of destabilizing electrostatic interactions may be responsible for the increased propensity of the mutated LC domains to self-associate in disease-causing conformations.
The coiled-coil domains of intermediate filament (IF) proteins are flanked by regions of low sequence complexity. Whereas IF coiled-coil domains assume dimeric and tetrameric conformations on their ...own, maturation of eight tetramers into cylindrical IFs is dependent on either “head” or “tail” domains of low sequence complexity. Here we confirm that the tail domain required for assembly of Drosophila Tm1-I/C IFs functions by forming labile cross-β interactions. These interactions are seen in polymers made from the tail domain alone, as well as in assembled IFs formed by the intact Tm1-I/C protein. The ability to visualize such interactions in situ within the context of a discrete cellular assembly lends support to the concept that equivalent interactions may be used in organizing other dynamic aspects of cell morphology.
•The low complexity (LC) domains of ataxin-2 and TDP43 are methionine rich.•Methionine oxidation blocks self-association and phase separation by both LC domains.•Reduction of oxidized methionines by ...Msr enzymes restores phase separation.•Complexes formed by the LC domains represent evolutionarily crafted redox sensors.
Eukaryotic cells express thousands of protein domains long believed to function in the absence of molecular order. These intrinsically disordered protein (IDP) domains are typified by gibberish-like repeats of only a limited number of amino acids that we refer to as domains of low sequence complexity. A decade ago, it was observed that these low complexity (LC) domains can undergo phase transition out of aqueous solution to form either liquid-like droplets or hydrogels. The self-associative interactions responsible for phase transition involve the formation of specific cross-β structures that are unusual in being labile to dissociation. Here we give evidence that the LC domains of two RNA binding proteins, ataxin-2 and TDP43, form cross-β interactions that specify biologically relevant redox sensors.
This perspective begins with a speculative consideration of the properties of the earliest proteins to appear during evolution. What did these primitive proteins look like, and how were they of ...benefit to early forms of life? I proceed to hypothesize that primitive proteins have been preserved through evolution and now serve diverse functions important to the dynamics of cell morphology and biological regulation. The primitive nature of these modern proteins is easy to spot. They are composed of a limited subset of the 20 amino acids used by traditionally evolved proteins and thus are of low sequence complexity. This chemical simplicity limits protein domains of low sequence complexity to forming only a crude and labile type of protein structure currently hidden from the computational powers of machine learning. I conclude by hypothesizing that this structural weakness represents the underlying virtue of proteins that, at least for the moment, constitute the dark matter of the proteome.
This review covers research findings reported over the past decade concerning the ability of low complexity (LC) domains to self-associate in a manner leading to their phase separation from aqueous ...solution. We focus our message upon the reductionist use of two forms of phase separation as biochemical assays to study how LC domains might function in living cells. Cells and their varied compartments represent extreme examples of material condensates. Over the past half century, biochemists, structural biologists, and molecular biologists have resolved the mechanisms driving innumerable forms of macromolecular condensation. In contrast, we remain largely ignorant as to how 10%-20% of our proteins actually work to assist in cell organization. This enigmatic 10%-20% of the proteome corresponds to gibberish-like LC sequences. We contend that many of these LC sequences move in and out of a structurally ordered, self-associated state as a means of offering a combination of organizational specificity and dynamic pliability to living cells. Finally, we speculate that ancient proteins may have behaved similarly, helping to condense, organize, and protect RNA early during evolution.
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