scholarly journals Signal and Specificity of Protein Ubiquitination for Proteasomal Degradation

2016 ◽  
Author(s):  
Yandong Yin ◽  
Jin Yang
Keyword(s):  
Theoretical Models ◽  
Proteasomal Degradation ◽  
Substrate Affinity ◽  
Transfer Rates ◽  
Biochemical Processes ◽  
Protein Ubiquitination ◽  
Ubiquitin System ◽  
Dna Repair Protein ◽  
Signal Specificity ◽  
Substrate Discrimination

The eukaryotic ubiquitin system regulates essential cell events such as DNA repair, protein homeostasis, and signal transduction. Like many biochemical processes, ubiquitination must ensure signaling efficiency and in the meantime maintain substrate specificity. We examine this signal-specificity relationship by theoretical models of polyubiquitinations that tag proteins for the proteasomal degradation. Parsimonious models provide explicit formulas to key measurable quantities and offer guiding insights into the signal-specificity tradeoffs under varying structures and kinetics. Models with measured kinetics from two primary cell-cycle ligases (SCF and APC) explain mechanisms of chain initiation, elongation slowdown, chain-length dependence of E3-substrate affinity, and deubiquitinases. We find that substrate discrimination over ubiquitin transfer rates is consistently more efficient than over substrate-E3 ligase binding energy, regardless of circuit structure, parameter value, and dynamics. E3-associated substrate deubiquitination increases the discrimination over the former and in the meantime decreases the latter, further widening their difference. Both discrimination strategies might be simultaneously explored by an E3 system to effectively proofread substrates as we demonstrated by analyzing experimental data from the CD4-Vpu-SCF system. We also identify that sequential deubiquitination circuit may act as a specificity switch, by which a modest change in deubiquitination and/or processivity can greatly increase substrate discrimination without much compromise in degradation signal. This property may be utilized as a gatekeeper mechanism to direct a temporal polyubiquitination and thus degradation order of substrates with small biochemical differences.

scholarly journals Stochastic reaction networks in dynamic compartment populations

2020 ◽  
Vol 117 (37) ◽  
pp. 22674-22683
Author(s):  
Lorenzo Duso ◽  
Christoph Zechner
Keyword(s):  
Biological Systems ◽  
Reaction Dynamics ◽  
Theoretical Models ◽  
Counting Processes ◽  
Mathematical Framework ◽  
Biological Processes ◽  
Biochemical Processes ◽  
Subcellular Compartmentalization ◽  
Stochastic Reaction ◽  
Stochastic Reaction Networks

Compartmentalization of biochemical processes underlies all biological systems, from the organelle to the tissue scale. Theoretical models to study the interplay between noisy reaction dynamics and compartmentalization are sparse, and typically very challenging to analyze computationally. Recent studies have made progress toward addressing this problem in the context of specific biological systems, but a general and sufficiently effective approach remains lacking. In this work, we propose a mathematical framework based on counting processes that allows us to study dynamic compartment populations with arbitrary interactions and internal biochemistry. We derive an efficient description of the dynamics in terms of differential equations which capture the statistics of the population. We demonstrate the relevance of our approach by analyzing models inspired by different biological processes, including subcellular compartmentalization and tissue homeostasis.

The emerging complexity of protein ubiquitination

2009 ◽  
Vol 37 (5) ◽  
pp. 937-953 ◽  
Author(s):  
David Komander
Keyword(s):  
Protein Phosphorylation ◽  
Intracellular Signalling ◽  
Present Review ◽  
Regulatory Mechanisms ◽  
Protein Regulation ◽  
Ubiquitin Chain ◽  
Post Translational Modifications ◽  
Protein Ubiquitination ◽  
Ubiquitin System ◽  
Signalling Cascades

Protein ubiquitination and protein phosphorylation are two fundamental regulatory post-translational modifications controlling intracellular signalling events. However, the ubiquitin system is vastly more complex compared with phosphorylation. This is due to the ability of ubiquitin to form polymers, i.e. ubiquitin chains, of at least eight different linkages. The linkage type of the ubiquitin chain determines whether a modified protein is degraded by the proteasome or serves to attract proteins to initiate signalling cascades or be internalized. The present review focuses on the emerging complexity of the ubiquitin system. I review what is known about individual chain types, and highlight recent advances that explain how the ubiquitin system achieves its intrinsic specificity. There is much to be learnt from the better-studied phosphorylation system, and many key regulatory mechanisms underlying control by protein phosphorylation may be similarly employed within the ubiquitin system. For example, ubiquitination may have important allosteric roles in protein regulation that are currently not appreciated.

scholarly journals Homocysteine thiolactone affects protein ubiquitination in yeast.

2013 ◽  
Vol 60 (3) ◽  
Author(s):  
Ewa Bretes ◽  
Jarosław Zimny
Keyword(s):  
Proteasomal Degradation ◽  
Amino Group ◽  
Homocysteine Thiolactone ◽  
Positive Correlation ◽  
Ubiquitinated Proteins ◽  
Protein Ubiquitination ◽  
Yeast Strains ◽  
Adverse Result

The formation of homocysteine thiolactone (HcyTl) from homocysteine occurs in all examined so far organisms including bacteria, yeast, and humans. Protein N-homocysteinylation at the ε-amino group of lysine is an adverse result of HcyTl accumulation. Since tagging of proteins by ubiquitination before their proteasomal degradation takes place at the same residue, we wondered how N-homocysteinylation may affect the ubiquitination of proteins. We used different yeast strains carrying mutations in genes involved in the homocysteine metabolism. We found positive correlation between the concentration of endogenous HcyTl and the concentration of ubiquitinated proteins. This suggests that N-homocysteinylation of proteins apparently does not preclude but rather promotes their decomposition.

Quantitative analysis of biochemical processes in living cells at a single-molecule level: a case of olaparib-PARP1 (DNA repair protein) interactions

The Analyst ◽  
2021 ◽  
Author(s):  
Aneta Karpinska ◽  
Marta Pilz ◽  
Joanna Buczkowska ◽  
Paweł Żuk ◽  
Karolina Kucharska ◽  
...  
Keyword(s):  
Quantitative Analysis ◽  
Single Molecule ◽  
Protein Interactions ◽  
Quantitative Description ◽  
Living Cells ◽  
Single Molecule Level ◽  
Biochemical Processes ◽  
Repair Protein ◽  
Dna Repair Protein ◽  
Modern Instrumentation

Quantitative description of biochemical processes inside living cells and at single-molecule levels remains a challenge at the forefront of modern instrumentation and spectroscopy. This paper demonstrates such single-cell, single-molecule analyses...

Role of ubiquitin in proteasomal degradation of mutant α1-antitrypsin Z in the endoplasmic reticulum

2000 ◽  
Vol 278 (1) ◽  
pp. G39-G48 ◽  
Author(s):  
Jeffrey H. Teckman ◽  
Reid Gilmore ◽  
David H. Perlmutter
Keyword(s):  
Endoplasmic Reticulum ◽  
Cell Lines ◽  
Human Fibroblast ◽  
Fibroblast Cell ◽  
Proteasomal Degradation ◽  
Intact Cells ◽  
Free System ◽  
Ubiquitin System ◽  
Human Fibroblast Cell ◽  
Fibroblast Cell Lines

A delay in intracellular degradation of the mutant α1-antitrypsin (α1AT)Z molecule is associated with greater retention within the endoplasmic reticulum (ER) and susceptibility to liver disease in a subgroup of patients with α1AT deficiency. Recent studies have shown that α1ATZ is ordinarily degraded in the ER by a mechanism that involves the proteasome, as demonstrated in intact cells using human fibroblast cell lines engineered for expression of α1ATZ and in a cell-free microsomal translocation assay system programmed with purified α1ATZ mRNA. To determine whether the ubiquitin system is required for proteasomal degradation of α1ATZ and whether specific components of the ubiquitin system can be implicated, we have now used two approaches. First, we overexpressed a dominant-negative ubiquitin mutant (UbK48R-G76A) by transient transfection in the human fibroblast cell lines expressing α1ATZ. The results showed that there was marked, specific, and selective inhibition of α1ATZ degradation mediated by UbK48R-G76A, indicating that the ubiquitin system is at least in part involved in ER degradation of α1ATZ. Second, we subjected reticulocyte lysate to DE52 chromatography and tested the resulting well-characterized fractions in the cell-free system. The results showed that there were both ubiquitin-dependent and -independent proteasomal mechanisms for degradation of α1ATZ and that the ubiquitin-conjugating enzyme E2-F1 may play a role in the ubiquitin-dependent proteasomal mechanism.

scholarly journals Histone Deacetylase Inhibitors Induce VHL and Ubiquitin-Independent Proteasomal Degradation of Hypoxia-Inducible Factor 1α

2006 ◽  
Vol 26 (6) ◽  
pp. 2019-2028 ◽  
Author(s):  
Xianguo Kong ◽  
Zhao Lin ◽  
Dongming Liang ◽  
Donna Fath ◽  
Nianli Sang ◽  
...  
Keyword(s):  
Histone Deacetylase ◽  
Histone Deacetylase Inhibitors ◽  
Anaerobic Metabolism ◽  
Hypoxia Inducible Factor ◽  
Proteasomal Degradation ◽  
Degradation Pathway ◽  
Von Hippel Lindau ◽  
Hypoxia Inducible Factor 1Α ◽  
Ubiquitin System ◽  
Tumor Survival

ABSTRACT Adaptation to hypoxic microenvironment is critical for tumor survival and metastatic spread. Hypoxia-inducible factor 1α (HIF-1α) plays a key role in this adaptation by stimulating the production of proangiogenic factors and inducing enzymes necessary for anaerobic metabolism. Histone deacetylase inhibitors (HDACIs) produce a marked inhibition of HIF-1α expression and are currently in clinical trials partly based on their potent antiangiogenic effects. Although it has been postulated that HDACIs affect HIF-1α expression by enhancing its interactions with VHL (von Hippel Lindau), thus promoting its ubiquitination and degradation, the actual mechanisms by which HDACIs decrease HIF-1α levels are not clear. Here, we present data indicating that HDACIs induce the proteasomal degradation of HIF-1α by a mechanism that is independent of VHL and p53 and does not require the ubiquitin system. This degradation pathway involves the enhanced interaction of HIF-1α with HSP70 and is secondary to a disruption of the HSP70/HSP90 axis function that appears mediated by the activity of HDAC-6.

scholarly journals A general in vitro assay to study enzymatic activities of the ubiquitin system

2019 ◽  
Author(s):  
Yukun Zuo ◽  
Boon Keat Chong ◽  
Kun Jiang ◽  
Daniel Finley ◽  
David Klenerman ◽  
...  
Keyword(s):  
Reaction Rates ◽  
Vitro Assay ◽  
Protein Ubiquitination ◽  
Ubiquitination Assay ◽  
Ubiquitin System ◽  
Wide Range ◽  
Machine Learning Approach ◽  
Fret Efficiency ◽  
Kinetics Of

AbstractThe ubiquitin (Ub) system regulates a wide range of cellular signaling pathways. Several hundred E1, E2 and E3 enzymes are together responsible for protein ubiquitination, thereby controlling cellular activities. Due to the numerous enzymes and processes involved, studies on ubiquitination activities have been challenging. We here report a novel FRET-based assay to study the in vitro kinetics of ubiquitination. FRET is established between binding of fluorophore-labeled Ub to eGFP-tagged ZnUBP, a domain that exclusively binds unconjugated Ub. We name this assay the Free Ub Sensor System (FUSS). Using Uba1, UbcH5 and CHIP as model E1, E2 and E3 enzymes, respectively, we demonstrate that ubiquitination results in decreasing FRET efficiency, from which reaction rates can be determined. Further treatment with USP21, a deubiquitinase, leads to increased FRET efficiency, confirming the reversibility of the assay. We subsequently use this assay to show that increasing the concentration of CHIP or UbcH5 but not Uba1 enhances ubiquitination rates, and develop a novel machine learning approach to model ubiquitination. The overall ubiquitination activity is also increased upon incubation with tau, a substrate of CHIP. Our data together demonstrate the versatile applications of a novel ubiquitination assay that does not require labeling of E1, E2, E3 or substrates, and is thus likely compatible with any E1-E2-E3 combinations.

3-D imaging of cells using a confocal laser scanning microscope and digital image processing

1988 ◽  
Vol 46 ◽  
pp. 96-97
Author(s):  
Thomas M. Jovin ◽  
Michel Robert-Nicoud ◽  
Donna J. Arndt-Jovin ◽  
Thorsten Schormann
Keyword(s):  
Laser Scanning ◽  
Confocal Laser Scanning Microscope ◽  
Laser Scanning Microscopy ◽  
Confocal Laser Scanning ◽  
Confocal Laser ◽  
Scanning Microscope ◽  
Laser Scanning Microscope ◽  
Biochemical Processes ◽  
Adjacent Structures

Light microscopic techniques for visualizing biomolecules and biochemical processes in situ have become indispensable in studies concerning the structural organization of supramolecular assemblies in cells and of processes during the cell cycle, transformation, differentiation, and development. Confocal laser scanning microscopy offers a number of advantages for the in situ localization and quantitation of fluorescence labeled targets and probes: (i) rejection of interfering signals emanating from out-of-focus and adjacent structures, allowing the “optical sectioning” of the specimen and 3-D reconstruction without time consuming deconvolution; (ii) increased spatial resolution; (iii) electronic control of contrast and magnification; (iv) simultanous imaging of the specimen by optical phenomena based on incident, scattered, emitted, and transmitted light; and (v) simultanous use of different fluorescent probes and types of detectors.We currently use a confocal laser scanning microscope CLSM (Zeiss, Oberkochen) equipped with 3-laser excitation (u.v - visible) and confocal optics in the fluorescence mode, as well as a computer-controlled X-Y-Z scanning stage with 0.1 μ resolution.

Analytical Electron Microscopy of Surface-Modified Ceramics

1985 ◽  
Vol 43 ◽  
pp. 276-279
Author(s):  
P. S. Sklad
Keyword(s):  
Electron Microscopy ◽  
Analytical Electron Microscopy ◽  
Theoretical Models ◽  
Ceramic Materials ◽  
Solute Concentration ◽  
Second Phase ◽  
Analytical Electron ◽  
Implantation Techniques ◽  
Lattice Damage ◽  
Surface Modified

Over the past several years, it has become increasingly evident that materials for proposed advanced energy systems will be required to operate at high temperatures and in aggressive environments. These constraints make structural ceramics attractive materials for these systems. However it is well known that the condition of the specimen surface of ceramic materials is often critical in controlling properties such as fracture toughness, oxidation resistance, and wear resistance. Ion implantation techniques offer the potential of overcoming some of the surface related limitations.While the effects of implantation on surface sensitive properties may be measured indpendently, it is important to understand the microstructural evolution leading to these changes. Analytical electron microscopy provides a useful tool for characterizing the microstructures produced in terms of solute concentration profiles, second phase formation, lattice damage, crystallinity of the implanted layer, and annealing behavior. Such analyses allow correlations to be made with theoretical models, property measurements, and results of complimentary techniques.

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