scholarly journals Super Spy variants implicate flexibility in chaperone action

eLife ◽  
2014 ◽  
Vol 3 ◽  
Author(s):  
Shu Quan ◽  
Lili Wang ◽  
Evgeniy V Petrotchenko ◽  
Karl AT Makepeace ◽  
Scott Horowitz ◽  
...  
Keyword(s):  
Protein Function ◽  
Genetic Selection ◽  
Chaperone Activity ◽  
Wild Type ◽  
Good Relationship ◽  
Chaperone Function ◽  
Binding Partners ◽  
Client Proteins ◽  
Adaptive Recognition

Experimental study of the role of disorder in protein function is challenging. It has been proposed that proteins utilize disordered regions in the adaptive recognition of their various binding partners. However apart from a few exceptions, defining the importance of disorder in promiscuous binding interactions has proven to be difficult. In this paper, we have utilized a genetic selection that links protein stability to antibiotic resistance to isolate variants of the newly discovered chaperone Spy that show an up to 7 fold improved chaperone activity against a variety of substrates. These “Super Spy” variants show tighter binding to client proteins and are generally more unstable than is wild type Spy and show increases in apparent flexibility. We establish a good relationship between the degree of their instability and the improvement they show in their chaperone activity. Our results provide evidence for the importance of disorder and flexibility in chaperone function.

scholarly journals Genetic Analysis of the Hsm3 Protein Function in Yeast Saccharomyces cerevisiae NuB4 Complex

Genes ◽  
2021 ◽  
Vol 12 (7) ◽  
pp. 1083
Author(s):  
Tatiyana A. Evstyukhina ◽  
Elena A. Alekseeva ◽  
Dmitriy V. Fedorov ◽  
Vyacheslav T. Peshekhonov ◽  
Vladimir G. Korolev
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Protein Function ◽  
Wild Type ◽  
Core Complex ◽  
Yeast Saccharomyces Cerevisiae ◽  
Nuclear Compartment ◽  
Spontaneous Mutagenesis ◽  
Chaperone Function ◽  
Induced Mutagenesis

In the nuclear compartment of yeast, NuB4 core complex consists of three proteins, Hat1, Hat2, and Hif1, and interacts with a number of other factors. In particular, it was shown that NuB4 complex physically interacts with Hsm3p. Early we demonstrated that the gene HSM3 participates in the control of replicative and reparative spontaneous mutagenesis, and that hsm3Δ mutants increase the frequency of mutations induced by different mutagens. It was previously believed that the HSM3 gene controlled only some minor repair processes in the cell, but later it was suggested that it had a chaperone function with its participation in proteasome assembly. In this work, we analyzed the properties of three hsm3Δ, hif1Δ, and hat1Δ mutants. The results obtained showed that the Hsm3 protein may be a functional subunit of NuB4 complex. It has been shown that hsm3- and hif1-dependent UV-induced mutagenesis is completely suppressed by inactivation of the Polη polymerase. We showed a significant role of Polη for hsm3-dependent mutagenesis at non-bipyrimidine sites (NBP sites). The efficiency of expression of RNR (RiboNucleotid Reducase) genes after UV irradiation in hsm3Δ and hif1Δ mutants was several times lower than in wild-type cells. Thus, we have presented evidence that significant increase in the dNTP levels suppress hsm3- and hif1-dependent mutagenesis and Polη is responsible for hsm3- and hif1-dependent mutagenesis.

ANGPTL4 modulates vascular junction integrity by integrin signaling and disruption of intercellular VE-cadherin and claudin-5 clusters

Blood ◽  
2011 ◽  
Vol 118 (14) ◽  
pp. 3990-4002 ◽  
Author(s):  
Royston-Luke Huang ◽  
Ziqiang Teo ◽  
Han Chung Chong ◽  
Pengcheng Zhu ◽  
Ming Jie Tan ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Integrin Signaling ◽  
Wild Type ◽  
Integrin Α5β1 ◽  
Vascular Integrity ◽  
Vascular Disruption ◽  
Binding Partners ◽  
Adhesive Proteins

Abstract Vascular disruption induced by interactions between tumor-secreted permeability factors and adhesive proteins on endothelial cells facilitates metastasis. The role of tumor-secreted C-terminal fibrinogen-like domain of angiopoietin-like 4 (cANGPTL4) in vascular leakiness and metastasis is controversial because of the lack of understanding of how cANGPTL4 modulates vascular integrity. Here, we show that cANGPTL4 instigated the disruption of endothelial continuity by directly interacting with 3 novel binding partners, integrin α5β1, VE-cadherin, and claudin-5, in a temporally sequential manner, thus facilitating metastasis. We showed that cANGPTL4 binds and activates integrin α5β1-mediated Rac1/PAK signaling to weaken cell–cell contacts. cANGPTL4 subsequently associated with and declustered VE-cadherin and claudin-5, leading to endothelial disruption. Interfering with the formation of these cANGPTL4 complexes delayed vascular disruption. In vivo vascular permeability and metastatic assays performed using ANGPTL4-knockout and wild-type mice injected with either control or ANGPTL4-knockdown tumors confirmed that cANGPTL4 induced vascular leakiness and facilitated lung metastasis in mice. Thus, our findings elucidate how cANGPTL4 induces endothelial disruption. Our findings have direct implications for targeting cANGPTL4 to treat cancer and other vascular pathologies.

scholarly journals Role of N-Linked Glycans in a Human Immunodeficiency Virus Envelope Glycoprotein: Effects on Protein Function and the Neutralizing Antibody Response

2002 ◽  
Vol 76 (9) ◽  
pp. 4199-4211 ◽  
Author(s):  
Miriam I. Quiñones-Kochs ◽  
Linda Buonocore ◽  
John K. Rose
Keyword(s):  
Human Immunodeficiency Virus ◽  
Envelope Glycoprotein ◽  
Protein Function ◽  
Neutralizing Antibodies ◽  
Immune Recognition ◽  
Wild Type ◽  
Virus Envelope ◽  
Mutant Proteins ◽  
Immunodeficiency Virus

ABSTRACT The envelope (Env) glycoprotein of human immunodeficiency virus (HIV) contains 24 N-glycosylation sites covering much of the protein surface. It has been proposed that one role of these carbohydrates is to form a shield that protects the virus from immune recognition. Strong evidence for such a role for glycosylation has been reported for simian immunodeficiency virus (SIV) mutants lacking glycans in the V1 region of Env (J. N. Reitter, R. E. Means, and R. C. Desrosiers, Nat. Med. 4:679-684, 1998). Here we used recombinant vesicular stomatitis viruses (VSVs) expressing HIV Env glycosylation mutants to determine if removal of carbohydrates in the V1 and V2 domains affected protein function and the generation of neutralizing antibodies in mice. Mutations that eliminated one to six of the sites for N-linked glycosylation in the V1 and V2 loops were introduced into a gene encoding the HIV type 1 primary isolate 89.6 envelope glycoprotein with its cytoplasmic domain replaced by that of the VSV G glycoprotein. The membrane fusion activities of the mutant proteins were studied in a syncytium induction assay. The transport and processing of the mutant proteins were studied with recombinant VSVs expressing mutant Env G proteins. We found that HIV Env V1 and V2 glycosylation mutants were no better than wild-type envelope at inducing antibodies neutralizing wild-type Env, although an Env mutant lacking glycans appeared somewhat more sensitive to neutralization by antibodies raised to mutant or wild-type Env. These results indicate significant differences between SIV and HIV with regard to the roles of glycans in the V1 and V2 domains.

scholarly journals Mitochondrial peroxiredoxin functions as crucial chaperone reservoir in Leishmania infantum

2015 ◽  
Vol 112 (7) ◽  
pp. E616-E624 ◽  
Author(s):  
Filipa Teixeira ◽  
Helena Castro ◽  
Tânia Cruz ◽  
Eric Tse ◽  
Philipp Koldewey ◽  
...  
Keyword(s):  
Leishmania Infantum ◽  
Protein Unfolding ◽  
Protozoan Parasite ◽  
Chaperone Activity ◽  
Dual Function ◽  
Active Site Cysteine ◽  
Chaperone Function ◽  
Client Proteins

Cytosolic eukaryotic 2-Cys-peroxiredoxins have been widely reported to act as dual-function proteins, either detoxifying reactive oxygen species or acting as chaperones to prevent protein aggregation. Several stimuli, including peroxide-mediated sulfinic acid formation at the active site cysteine, have been proposed to trigger the chaperone activity. However, the mechanism underlying this activation and the extent to which the chaperone function is crucial under physiological conditions in vivo remained unknown. Here we demonstrate that in the vector-borne protozoan parasite Leishmania infantum, mitochondrial peroxiredoxin (Prx) exerts intrinsic ATP-independent chaperone activity, protecting a wide variety of different proteins against heat stress-mediated unfolding in vitro and in vivo. Activation of the chaperone function appears to be induced by temperature-mediated restructuring of the reduced decamers, promoting binding of unfolding client proteins in the center of Prx’s ringlike structure. Client proteins are maintained in a folding-competent conformation until restoration of nonstress conditions, upon which they are released and transferred to ATP-dependent chaperones for refolding. Interference with client binding impairs parasite infectivity, providing compelling evidence for the in vivo importance of Prx’s chaperone function. Our results suggest that reduced Prx provides a mitochondrial chaperone reservoir, which allows L. infantum to deal successfully with protein unfolding conditions during the transition from insect to the mammalian hosts and to generate viable parasites capable of perpetuating infection.

scholarly journals A novel multifunctional role for Hsp70 in binding post-translational modifications on clients

2021 ◽  
Author(s):  
Nitika ◽  
Bo Zheng ◽  
Linhao Ruan ◽  
Jake Kline ◽  
Jacek Sikora ◽  
...  
Keyword(s):  
Protein Function ◽  
Point Interaction ◽  
Post Translational Modifications ◽  
Definitive Evidence ◽  
Chaperone Function ◽  
Transient Nature ◽  
Hsp70 Protein ◽  
Response To Stress ◽  
Client Proteins ◽  
Self Association

Hsp70 interactions are critical for cellular viability and the response to stress. Previous attempts to characterize Hsp70 interactions have been limited by their transient nature and inability of current technologies to distinguish direct vs bridged interactions. We report the novel use of cross-linking mass spectrometry (XL-MS) to comprehensively characterize the budding yeast Hsp70 protein interactome. Using this approach, we have gained fundamental new insights into Hsp70 function, including definitive evidence of Hsp70 self-association as well as multi-point interaction with its client proteins. In addition to identifying a novel set of direct Hsp70 interactors which can be used to probe chaperone function in cells, we have also identified a suite of PTM-associated Hsp70 interactions. The majority of these PTMs have not been previously reported and appear to be critical in the regulation of client protein function. These data indicate that one of the mechanisms by which PTMs contribute to protein function is by facilitating interaction with chaperones. Taken together, we propose that XL-MS analysis of chaperone complexes may be used as a unique way to identify biologically-important PTMs on client proteins.

scholarly journals Molecular analysis of scabrous mutant alleles from Drosophila melanogaster indicates a secreted protein with two functional domains.

Genetics ◽  
1995 ◽  
Vol 141 (2) ◽  
pp. 607-617
Author(s):  
X Hu ◽  
E C Lee ◽  
N E Baker
Keyword(s):  
Protein Interactions ◽  
Neural Development ◽  
Protein Function ◽  
Blood Clotting ◽  
Dominant Negative ◽  
Wild Type ◽  
Amino Terminal ◽  
Amino Terminal Domain ◽  
Amino Acid Segment

Abstract Mutations at the scabrous locus (sca) affect cell-cell signaling during neural development. Twenty-one mutant alleles of scabrous have been analyzed. Many synthesize no sca protein. In others, a defective protein is arrested intracellularly. Two mutants in which protein is not arrested must affect sca protein function outside the cell. Both affect the fibrinogen related domain (FReD), a 200-amino acid segment conserved in fibrinogen, tenascins, and other proteins. In fibrinogen, this region is involved in protein interactions and is altered in human mutations affecting blood clotting. In sca(UM2), an invariant Asp residue is replaced by Asn. In sca(MSKF) allele has dominant negative properties, indicating that the truncated amino-terminal portion interferes with the function of so me other gene product. These mutations show that the conserved FReD is essential for wild-type sca function, but suggest that the amino-terminal domain also interacts with other proteins, but other neural mutations were without effect. Models for the role of a two-domain protein in neural development are discussed.

scholarly journals RICE CENTRORADIALIS 1, a TFL1-like gene, Responses to Drought Stress and Regulates Rice Flowering Transition

2020 ◽  
Author(s):  
Yan Wang ◽  
Yuyang Lu ◽  
Ziyu Guo ◽  
Yanfeng Ding ◽  
Chengqiang Ding
Keyword(s):  
Drought Stress ◽  
Protein Function ◽  
Heading Date ◽  
Wild Type ◽  
Terminal Flower ◽  
Complex Process ◽  
Genes And Environment ◽  
Repression Complex ◽  
Normal Water

Abstract Background: The initiation of flowering transition in rice (Oryza sativa) is a complex process regulated by genes and environment. In particular, drought can interfere with flowering; therefore, many plants hasten this process to shorten their life cycle under water scarcity, and this is known as drought-escape response. However, rice has other strategies; for example, drought stress can delay flowering instead of accelerating it. RICE CENTRORADIALIS 1 (RCN1) is a TERMINAL FLOWER-like gene that influences rice flowering transition and spike differentiation. It interacts with 14-3-3 proteins and transcription factor OsFD1 to form a florigen repression complex that suppresses flowering transition in rice.Results: In this study, we explored the role of RCN1 in the molecular pathway of drought-regulated flowering transition. The rcn1 mutant plants displayed early heading under both normal water and drought stress conditions, and they were more insensitive to drought stress than the wild-type plants. Abscisic acid (ABA) signaling-mediated drought-induced RCN1 is involved in this process.Conclusions: Thus, RCN1 plays an important role in the process of drought stress inhibiting flowering transition. It may worked by suppressing the protein function rather than transcription of HEADING DATE 3a.

Effect of methylglyoxal modification on stress-induced aggregation of client proteins and their chaperoning by human αA-crystallin

2008 ◽  
Vol 409 (3) ◽  
pp. 771-777 ◽  
Author(s):  
Ashis Biswas ◽  
Benlian Wang ◽  
Masaru Miyagi ◽  
Ram H. Nagaraj
Keyword(s):  
Alcohol Dehydrogenase ◽  
Protein Aggregation ◽  
Surface Hydrophobicity ◽  
Stress Conditions ◽  
Thermal Aggregation ◽  
Chaperone Function ◽  
Client Proteins ◽  
Modified Proteins ◽  
Induced Aggregation

α-Crystallin prevents protein aggregation under various stress conditions through its chaperone-like properties. Previously, we demonstrated that MGO (methylglyoxal) modification of αA-crystallin enhances its chaperone function and thus may affect transparency of the lens. During aging of the lens, not only αA-crystallin, but its client proteins are also likely to be modified by MGO. We have investigated the role of MGO modification of four model client proteins (insulin, α-lactalbumin, alcohol dehydrogenase and γ-crystallin) in their aggregation and structure and the ability of human αA-crystallin to chaperone them. We found that MGO modification (10–1000 μM) decreased the chemical aggregation of insulin and α-lactalbumin and thermal aggregation of alcohol dehydrogenase and γ-crystallin. Surface hydrophobicity in MGO-modified proteins decreased slightly relative to unmodified proteins. HPLC and MS analyses revealed argpyrimidine and hydroimidazolone in MGO-modified client proteins. The degree of chaperoning by αA-crystallin towards MGO-modified and unmodified client proteins was similar. Co-modification of client proteins and αA-crystallin by MGO completely inhibited stress-induced aggregation of client proteins. Our results indicate that minor modifications of client proteins and αA-crystallin by MGO might prevent protein aggregation and thus help maintain transparency of the aging lens.

scholarly journals RICE CENTRORADIALIS 1, a TFL1-like Gene, Responses to Drought Stress and Regulates Rice Flowering Transition

Rice ◽  
2020 ◽  
Vol 13 (1) ◽  
Author(s):  
Yan Wang ◽  
Yuyang Lu ◽  
Ziyu Guo ◽  
Yanfeng Ding ◽  
Chengqiang Ding
Keyword(s):  
Drought Stress ◽  
Protein Function ◽  
Heading Date ◽  
Wild Type ◽  
Terminal Flower ◽  
Complex Process ◽  
Genes And Environment ◽  
Repression Complex ◽  
Normal Water

Abstract Background The initiation of flowering transition in rice (Oryza sativa) is a complex process regulated by genes and environment. In particular, drought can interfere with flowering; therefore, many plants hasten this process to shorten their life cycle under water scarcity, and this is known as drought-escape response. However, rice has other strategies; for example, drought stress can delay flowering instead of accelerating it. RICE CENTRORADIALIS 1 (RCN1) is a TERMINAL FLOWER-like gene that influences rice flowering transition and spike differentiation. It interacts with 14–3-3 proteins and transcription factor OsFD1 to form a florigen repression complex that suppresses flowering transition in rice. Results In this study, we explored the role of RCN1 in the molecular pathway of drought-regulated flowering transition. The rcn1 mutant plants displayed early heading under both normal water and drought stress conditions, and they were more insensitive to drought stress than the wild-type plants. Abscisic acid (ABA) signaling-mediated drought-induced RCN1 is involved in this process. Conclusions Thus, RCN1 plays an important role in the process of drought stress inhibiting flowering transition. It may worked by suppressing the protein function rather than transcription of HEADING DATE 3a.

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