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Small GTPases Jul 2020Mutant RAS isoforms are the most common oncogenes affecting human cancers. After decades of effort in developing drugs targeting oncogenic RAS-driven cancers, we are... (Review)
Review
Mutant RAS isoforms are the most common oncogenes affecting human cancers. After decades of effort in developing drugs targeting oncogenic RAS-driven cancers, we are still charting an unclear path. Despite recent developments exemplified by KRAS (G12C) inhibitors, direct targeting of mutant RAS remains a difficult endeavor. Inhibiting RAS function by targeting its post-translational prenylation processing has remained an important approach, especially with recent progress on the study of isoprenylcysteine carboxylmethyltransferase (ICMT), the unique enzyme for the last step of prenylation processing of RAS isoforms and other substrates. Inhibition of ICMT has shown efficacy both and in RAS-mutant cancer models. We will discuss the roles of RAS family of proteins in human cancers and the impact of post-prenylation carboxylmethylation on RAS driven tumorigenesis. In addition, we will review what is known of the molecular and cellular impact of ICMT inhibition on cancer cells that underlie its anti-proliferative and pro-apoptosis efficacy.
Topics: Animals; Humans; Methylation; Protein Prenylation; ras Proteins
PubMed: 29261009
DOI: 10.1080/21541248.2017.1415637 -
Biochemical Pharmacology Dec 2015Post-translational prenylation involves incorporation of 15-(farnesyl) or 20-(geranylgeranyl) carbon derivatives of mevalonic acid into highly conserved C-terminal...
Post-translational prenylation involves incorporation of 15-(farnesyl) or 20-(geranylgeranyl) carbon derivatives of mevalonic acid into highly conserved C-terminal cysteines of proteins. The farnesyl transferase (FTase) and the geranylgeranyl transferase (GGTase) mediate incorporation of farnesyl and geranylgeranyl groups, respectively. At least 300 proteins are prenylated in the human genome; the majority of these are implicated in cellular processes including growth, differentiation, cytoskeletal function and vesicle trafficking. From a functional standpoint, isoprenylation is requisite for targeting of modified proteins to relevant cellular compartments for regulation of effector proteins. Pharmacological and molecular biological studies have provided compelling evidence for key roles of this signaling pathway in physiological insulin secretion in normal rodent and human islets. Recent evidence indicates that inhibition of prenylation results in mislocalization of unprenylated proteins, and surprisingly, they remain in active (GTP-bound) conformation. Sustained activation of G proteins has been reported in mice lacking GGTase, suggesting alternate mechanisms for the activation of non-prenylated G proteins. These findings further raise an interesting question if mislocalized, non-prenylated and functionally active G proteins cause cellular pathology since aberrant protein prenylation has been implicated in the onset of cardiovascular disease and diabetes. Herein, we overview the existing evidence to implicate prenylation in islet function and potential defects in this signaling pathways in the diabetic β-cell. We will also identify critical knowledge gaps that need to be addressed for the development of therapeutics to halt defects in these signaling steps in β cells in models of impaired insulin secretion, metabolic stress and diabetes.
Topics: Diabetes Mellitus; GTP-Binding Proteins; Glucose; Humans; Islets of Langerhans; Protein Prenylation; Protein Processing, Post-Translational; Signal Transduction
PubMed: 26215874
DOI: 10.1016/j.bcp.2015.07.004 -
Methods in Molecular Biology (Clifton,... 2019Protein prenylation, found in eukaryotes, is a posttranslational modification in which one or two isoprenoid groups are added to the C terminus of selected proteins...
Protein prenylation, found in eukaryotes, is a posttranslational modification in which one or two isoprenoid groups are added to the C terminus of selected proteins using either a farnesyl group or a geranylgeranyl group. Prenylation facilitates protein localization mainly to the plasma membrane where the prenylated proteins, including small GTPases, mediate signal transduction pathways. Changes in the level of prenylated proteins may serve a critical function in a variety of diseases. Metabolic labeling using modified isoprenoid probes followed by enrichment and proteomic analysis allows the identities and levels of prenylated proteins to be investigated. In this protocol, we illustrate how the conditions for metabolic labeling are optimized to maximize probe incorporation in HeLa cells through a combination of in-gel fluorescence and densitometric analysis.
Topics: HeLa Cells; Humans; Protein Prenylation; Proteomics; Staining and Labeling; Terpenes
PubMed: 31152393
DOI: 10.1007/978-1-4939-9532-5_3 -
International Journal of Molecular... Dec 2019The members of Rho family of GTPases, RhoA and Rac1 regulate endothelial cytoskeleton dynamics and hence barrier integrity. The spatial activities of these GTPases are...
The members of Rho family of GTPases, RhoA and Rac1 regulate endothelial cytoskeleton dynamics and hence barrier integrity. The spatial activities of these GTPases are regulated by post-translational prenylation. In the present study, we investigated the effect of prenylation inhibition on the endothelial cytoskeleton and barrier properties. The study was carried out in human umbilical vein endothelial cells (HUVEC) and protein prenylation is manipulated with various pharmacological inhibitors. Inhibition of either complete prenylation using statins or specifically geranylgeranylation but not farnesylation has a biphasic effect on HUVEC cytoskeleton and permeability. Short-term treatment inhibits the spatial activity of RhoA/Rho kinase (Rock) to actin cytoskeleton resulting in adherens junctions (AJ) stabilization and ameliorates thrombin-induced barrier disruption whereas long-term inhibition results in collapse of endothelial cytoskeleton leading to increased basal permeability. These effects are reversed by supplementing the cells with geranylgeranyl but not farnesyl pyrophosphate. Moreover, long-term inhibition of protein prenylation results in basal hyper activation of RhoA/Rock signaling that is antagonized by a specific Rock inhibitor or an activation of cAMP signaling. In conclusion, inhibition of geranylgeranylation in endothelial cells (ECs) exerts biphasic effect on endothelial barrier properties. Short-term inhibition stabilizes AJs and hence barrier function whereas long-term treatment results in disruption of barrier properties.
Topics: Cell Membrane Permeability; Cytoskeleton; Endothelial Cells; Endothelium; GTP Phosphohydrolases; Humans; Intercellular Junctions; Models, Biological; Protein Prenylation; rho-Associated Kinases
PubMed: 31861297
DOI: 10.3390/ijms21010002 -
ACS Chemical Biology Oct 2022Photoswitchable lipids have emerged as attractive tools for the optical control of lipid bioactivity, metabolism, and biophysical properties. Their design is typically...
Photoswitchable lipids have emerged as attractive tools for the optical control of lipid bioactivity, metabolism, and biophysical properties. Their design is typically based on the incorporation of an azobenzene photoswitch into the hydrophobic lipid tail, which can be switched between its - and -form using two different wavelengths of light. While glycero- and sphingolipids have been successfully designed to be photoswitchable, isoprenoid lipids have not yet been investigated. Herein, we describe the development of photoswitchable analogs of an isoprenoid lipid and systematically assess their potential for the optical control of various steps in the isoprenylation processing pathway of CaaX proteins in . One photoswitchable analog of farnesyl diphosphate () allowed effective optical control of substrate prenylation by farnesyltransferase. The subsequent steps of isoprenylation processing (proteolysis by either Ste24 or Rce1 and carboxyl methylation by Ste14) were less affected by photoisomerization of the group introduced into the lipid moiety of the substrate a-factor, a mating pheromone from yeast. We assessed both proteolysis and methylation of the a-factor analogs and the bioactivity of a fully processed a-factor analog containing the photoswitch, exogenously added to cognate yeast cells. Combined, these data describe the first successful conversion of an isoprenoid lipid into a photolipid and suggest the utility of this approach for the optical control of protein prenylation.
Topics: Saccharomyces cerevisiae; Terpenes; Farnesyltranstransferase; Peptides; Protein Prenylation; Pheromones; Lipids; Sphingolipids; Membrane Proteins; Metalloendopeptidases; Saccharomyces cerevisiae Proteins
PubMed: 36194691
DOI: 10.1021/acschembio.2c00645 -
Human & Experimental Toxicology 2022The resistance of glioblastoma to chemotherapy remains a significant clinical problem. Targeting alternative pathways such as protein prenylation is known to be...
The resistance of glioblastoma to chemotherapy remains a significant clinical problem. Targeting alternative pathways such as protein prenylation is known to be effective against many cancers. Fluvastatin is a potent competitive inhibitor of 3-hydroxy-3-methylglutaryl- CoA (HMG-CoA) reductase, thereby inhibits prenylation. We demonstrate that fluvastatin alone effectively inhibits proliferation and induces apoptosis in multiple human glioblastoma cell lines. The combination index analysis shows that fluvastatin acts synergistically with common chemotherapy drugs for glioblastoma: temozolomide and irinotecan. We further show that fluvastatin acts on glioblastoma through inhibiting prenylation-dependent Ras activation. The combination of fluvastatin and low dose temozolomide resulted in remarkable inhibition of glioblastoma tumor in mice throughout the whole treatment duration without causing toxicity. Such combinatorial effects provide the basis for utilizing these FDA-approved drugs as a potential clinical approach in overcoming resistance and improving glioblastoma treatment.
Topics: Animals; Coenzyme A; Drug Resistance, Neoplasm; Fatty Acids, Monounsaturated; Fluvastatin; Glioblastoma; Humans; Hydroxymethylglutaryl-CoA Reductase Inhibitors; Irinotecan; Mice; Oxidoreductases; Protein Prenylation; Temozolomide
PubMed: 36171180
DOI: 10.1177/09603271221125934 -
Chemical Reviews Feb 2018Protein lipidation, including cysteine prenylation, N-terminal glycine myristoylation, cysteine palmitoylation, and serine and lysine fatty acylation, occurs in many...
Protein lipidation, including cysteine prenylation, N-terminal glycine myristoylation, cysteine palmitoylation, and serine and lysine fatty acylation, occurs in many proteins in eukaryotic cells and regulates numerous biological pathways, such as membrane trafficking, protein secretion, signal transduction, and apoptosis. We provide a comprehensive review of protein lipidation, including descriptions of proteins known to be modified and the functions of the modifications, the enzymes that control them, and the tools and technologies developed to study them. We also highlight key questions about protein lipidation that remain to be answered, the challenges associated with answering such questions, and possible solutions to overcome these challenges.
Topics: Alkyl and Aryl Transferases; Animals; Humans; Kinetics; Myristic Acids; Palmitic Acids; Protein Interaction Maps; Protein Prenylation; Proteins; Substrate Specificity
PubMed: 29292991
DOI: 10.1021/acs.chemrev.6b00750 -
Current Protocols in Chemical Biology Sep 2018Protein prenylation involves the attachment of a farnesyl or geranylgeranyl group onto a cysteine residue located near the C-terminus of a protein, recognized via a...
Protein prenylation involves the attachment of a farnesyl or geranylgeranyl group onto a cysteine residue located near the C-terminus of a protein, recognized via a specific prenylation motif, and results in the formation of a thioether bond. To identify putative prenylated proteins and investigate changes in their levels of expression, metabolic labeling and subsequent bioorthogonal labeling has become one of the methods of choice. In that strategy, synthetic analogues of biosynthetic precursors for post-translational modification bearing bioorthogonal functionality are added to the growth medium from which they enter cells and become incorporated into proteins. Subsequently, the cells are lysed and proteins bearing the analogues are then covalently modified using selective chemical reagents that react via bioorthogonal processes, allowing a variety of probes for visualization or enrichment to be attached for subsequent analysis. Here, we describe protocols for synthesizing several different isoprenoid analogues and describe how they are metabolically incorporated into mammalian cells, and the incorporation into prenylated proteins visualized via in-gel fluorescence analysis. © 2018 by John Wiley & Sons, Inc.
Topics: Alkynes; Molecular Structure; Protein Prenylation; Proteins; Terpenes
PubMed: 30058775
DOI: 10.1002/cpch.46 -
Cell Metabolism Apr 2019Recent research focusing on brown adipose tissue (BAT) function emphasizes its importance in systemic metabolic homeostasis. We show here that genetic and...
Recent research focusing on brown adipose tissue (BAT) function emphasizes its importance in systemic metabolic homeostasis. We show here that genetic and pharmacological inhibition of the mevalonate pathway leads to reduced human and mouse brown adipocyte function in vitro and impaired adipose tissue browning in vivo. A retrospective analysis of a large patient cohort suggests an inverse correlation between statin use and active BAT in humans, while we show in a prospective clinical trial that fluvastatin reduces thermogenic gene expression in human BAT. We identify geranylgeranyl pyrophosphate as the key mevalonate pathway intermediate driving adipocyte browning in vitro and in vivo, whose effects are mediated by geranylgeranyltransferases (GGTases), enzymes catalyzing geranylgeranylation of small GTP-binding proteins, thereby regulating YAP1/TAZ signaling through F-actin modulation. Conversely, adipocyte-specific ablation of GGTase I leads to impaired adipocyte browning, reduced energy expenditure, and glucose intolerance under obesogenic conditions, highlighting the importance of this pathway in modulating brown adipocyte functionality and systemic metabolism.
Topics: Adipocytes, Brown; Adolescent; Adult; Animals; Cell Differentiation; Cell Proliferation; Cells, Cultured; Humans; Male; Mevalonic Acid; Mice; Mice, Inbred Strains; Middle Aged; Protein Prenylation; Uncoupling Protein 1; Young Adult
PubMed: 30581121
DOI: 10.1016/j.cmet.2018.11.017 -
Molecular Biology of the Cell May 2003The majority of Rab proteins are posttranslationally modified with two geranylgeranyl lipid moieties that enable their stable association with membranes. In this study,...
The majority of Rab proteins are posttranslationally modified with two geranylgeranyl lipid moieties that enable their stable association with membranes. In this study, we present evidence to demonstrate that there is a specific lipid requirement for Rab protein localization and function. Substitution of different prenyl anchors on Rab GTPases does not lead to correct function. In the case of YPT1 and SEC4, two essential Rab genes in Saccharomyces cerevisiae, alternative lipid tails cannot support life when present as the sole source of YPT1 and SEC4. Furthermore, our data suggest that double geranyl-geranyl groups are required for Rab proteins to correctly localize to their characteristic organelle membrane. We have identified a factor, Yip1p that specifically binds the di-geranylgeranylated Rab and does not interact with mono-prenylated Rab proteins. This is the first demonstration that the double prenylation modification of Rab proteins is an important feature in the function of this small GTPase family and adds specific prenylation to the already known determinants of Rab localization.
Topics: Membrane Proteins; Mutation; Protein Prenylation; Saccharomyces cerevisiae; Saccharomyces cerevisiae Proteins; Vesicular Transport Proteins; rab GTP-Binding Proteins
PubMed: 12802060
DOI: 10.1091/mbc.e02-11-0707