scholarly journals EphA2 Interacts with Tim-4 through Association between Its FN3 Domain and the IgV Domain of Tim-4

Cells ◽  
2021 ◽  
Vol 10 (6) ◽  
pp. 1290
Author(s):  
Byeongjin Moon ◽  
Susumin Yang ◽  
Kwangwoo Kim ◽  
Juyeon Lee ◽  
Dongtak Jeong ◽  
...  
Keyword(s):  
In Silico ◽  
Transmembrane Protein ◽  
Apoptotic Cells ◽  
Interacting Protein ◽  
Polystyrene Beads ◽  
Cellular Event ◽  
Extracellular Region ◽  
Fn3 Domain

Tim-4 promotes the engulfment of apoptotic cells or exogenous particles by securing them on phagocytes. It is unable to transduce signals by itself but helps other engulfment receptors sense and internalize them. However, the identity of the engulfment receptors collaborating with Tim-4 is still incompletely understood. In this study, we searched for a candidate transmembrane protein with a FN3 domain, important for interaction with Tim-4, in silico and investigated whether it indeed interacts with Tim-4 and is involved in Tim-4-mediated phagocytosis. We found that EphA2 containing a FN3 domain in the extracellular region interacted with Tim-4, which was mediated by the IgV domain of Tim-4 and the FN3 domain of EphA2. Nevertheless, we found that EphA2 expression failed to alter Tim-4-mediated phagocytosis of apoptotic cells or polystyrene beads. Taken together, our findings suggest that EphA2, a new Tim-4 interacting protein, may intervene in a Tim-4-mediated cellular event even if it is not phagocytosis of endogenous or exogenous particles and vice versa.

scholarly journals The tetraspanin Tspan15 is an essential subunit of an ADAM10 scissor complex

2020 ◽  
Vol 295 (36) ◽  
pp. 12822-12839 ◽  
Author(s):  
Chek Ziu Koo ◽  
Neale Harrison ◽  
Peter J. Noy ◽  
Justyna Szyroka ◽  
Alexandra L. Matthews ◽  
...  
Keyword(s):  
Monoclonal Antibodies ◽  
Cell Line ◽  
Knockout Mouse ◽  
Transmembrane Protein ◽  
Mouse Cell ◽  
Primary Cells ◽  
Interacting Protein ◽  
Mouse Cell Line ◽  
Extracellular Region ◽  
Essential Subunit

A disintegrin and metalloprotease 10 (ADAM10) is a transmembrane protein essential for embryonic development, and its dysregulation underlies disorders such as cancer, Alzheimer's disease, and inflammation. ADAM10 is a “molecular scissor” that proteolytically cleaves the extracellular region from >100 substrates, including Notch, amyloid precursor protein, cadherins, growth factors, and chemokines. ADAM10 has been recently proposed to function as six distinct scissors with different substrates, depending on its association with one of six regulatory tetraspanins, termed TspanC8s. However, it remains unclear to what degree ADAM10 function critically depends on a TspanC8 partner, and a lack of monoclonal antibodies specific for most TspanC8s has hindered investigation of this question. To address this knowledge gap, here we designed an immunogen to generate the first monoclonal antibodies targeting Tspan15, a model TspanC8. The immunogen was created in an ADAM10-knockout mouse cell line stably overexpressing human Tspan15, because we hypothesized that expression in this cell line would expose epitopes that are normally blocked by ADAM10. Following immunization of mice, this immunogen strategy generated four Tspan15 antibodies. Using these antibodies, we show that endogenous Tspan15 and ADAM10 co-localize on the cell surface, that ADAM10 is the principal Tspan15-interacting protein, that endogenous Tspan15 expression requires ADAM10 in cell lines and primary cells, and that a synthetic ADAM10/Tspan15 fusion protein is a functional scissor. Furthermore, two of the four antibodies impaired ADAM10/Tspan15 activity. These findings suggest that Tspan15 directly interacts with ADAM10 in a functional scissor complex.

scholarly journals Identifying new COVID-19 receptor Neuropilin-1 in severe Alzheimer’s diseases patients group brain using genome-wide association study approach

2021 ◽  
Author(s):  
Key-Hwan Lim ◽  
Sumin Yang ◽  
Sung-Hyun Kim ◽  
Jae-Yeol Joo
Keyword(s):  
In Silico ◽  
Genome Wide Association Study ◽  
Transmembrane Protein ◽  
In Silico Analysis ◽  
Therapeutic Drug ◽  
Rna Seq ◽  
Total Rna ◽  
Neuropilin 1 ◽  
Silico Analysis ◽  
Brain Tissues

Abstract Background Numerous studies have been conducted on different aspects of the COVID-19 (coronavirus disease 2019) pandemic, which is caused by SARS-CoV-2, since its emergence in late 2019. Mutual relations among SARS-CoV-2 and neuro-pathophysiological phenomena are continuously being demonstrated, and several underlying diseases, such as those in the elderly, are positively correlated with susceptibility to SARS-CoV-2 infection. The expression of angiotensin converting enzyme 2 (ACE2), which is required for SARS-CoV-2 infection, was recently demonstrated to be increased in Alzheimer’s disease (AD) patients. Methods Recent preclinical studies have shown that Neuropilin-1 (NRP1), which is a transmembrane protein with roles in neuronal development, axonal outgrowth, and angiogenesis, also plays a role in the infectivity of SARS-CoV-2. Thus, we hypothesized that NRP1 may be upregulated in AD patients and that a correlation between AD and SARS-CoV-2 NRP1-mediated infectivity may exist. We used an AD mouse model that mimics AD and performed high throughput total RNA-seq with brain tissue and whole blood. For quantification of NPR1 in AD, brain tissues and blood were subjected to western blotting and RT-qPCR analysis. In silico analysis for NRP1 expression in AD patients has been performed on the human hippocampus data sets (GSE4226, GSE1297). Results Many cases of severe symptom of COVID-19 are concentrated in elderly group who have complications such as diabetes, degenerative disease, and brain disorders. Total RNA-seq analysis showed that Nrp1 gene was commonly overexpressed in AD model. Similar to ACE2, NRP1 protein also strongly expressed in the AD brain tissues. Interestingly, in silico analysis revealed that the level of expression for NRP1 was distinct at age and AD progression. Conclusions Given that the NRP1 is highly expressed in AD, it will be important to understand and predict that NRP1 may a risk factor for SARS-CoV-2 infection in AD patients. This will support to development of potential therapeutic drug to reduce SARS-CoV-2 transmission.

scholarly journals Multiclass Classifier for P-Glycoprotein Substrates, Inhibitors, and Non-Active Compounds

Molecules ◽  
2019 ◽  
Vol 24 (10) ◽  
pp. 2006 ◽  
Author(s):  
Liadys Mora Lagares ◽  
Nikola Minovski ◽  
Marjana Novič
Keyword(s):  
In Silico ◽  
Transmembrane Protein ◽  
External Validation ◽  
Assessment Process ◽  
Classification Model ◽  
Training Set ◽  
Test Set ◽  
Active Compounds ◽  
P Glycoprotein ◽  
Validation Set

P-glycoprotein (P-gp) is a transmembrane protein that actively transports a wide variety of chemically diverse compounds out of the cell. It is highly associated with the ADMET (absorption, distribution, metabolism, excretion and toxicity) properties of drugs/drug candidates and contributes to decreasing toxicity by eliminating compounds from cells, thereby preventing intracellular accumulation. Therefore, in the drug discovery and toxicological assessment process it is advisable to pay attention to whether a compound under development could be transported by P-gp or not. In this study, an in silico multiclass classification model capable of predicting the probability of a compound to interact with P-gp was developed using a counter-propagation artificial neural network (CP ANN) based on a set of 2D molecular descriptors, as well as an extensive dataset of 2512 compounds (1178 P-gp inhibitors, 477 P-gp substrates and 857 P-gp non-active compounds). The model provided a good classification performance, producing non error rate (NER) values of 0.93 for the training set and 0.85 for the test set, while the average precision (AvPr) was 0.93 for the training set and 0.87 for the test set. An external validation set of 385 compounds was used to challenge the model’s performance. On the external validation set the NER and AvPr values were 0.70 for both indices. We believe that this in silico classifier could be effectively used as a reliable virtual screening tool for identifying potential P-gp ligands.

scholarly journals Ethers and esters derived from apocynin avoid the interaction between p47phox and p22phox subunits of NADPH oxidase: evaluation in vitro and in silico

2013 ◽  
Vol 33 (4) ◽  
Author(s):  
Martha Edith Macías-Pérez ◽  
Federico Martínez-Ramos ◽  
Itzia Irene Padilla-Martínez ◽  
José Correa-Basurto ◽  
Lowell Kispert ◽  
...  
Keyword(s):  
Nadph Oxidase ◽  
In Silico ◽  
Transmembrane Protein ◽  
Vascular Diseases ◽  
Docking Study ◽  
Free Radical Scavengers ◽  
Phenyl Ester ◽  
Paramagnetic Resonance ◽  
In Silico Studies

NOX (NADPH oxidase) plays an important role during several pathologies because it produces the superoxide anion (O2•−), which reacts with NO (nitric oxide), diminishing its vasodilator effect. Although different isoforms of NOX are expressed in ECs (endothelial cells) of blood vessels, the NOX2 isoform has been considered the principal therapeutic target for vascular diseases because it can be up-regulated by inhibiting the interaction between its p47phox (cytosolic protein) and p22phox (transmembrane protein) subunits. In this research, two ethers, 4-(4-acetyl-2-methoxy-phenoxy)-acetic acid (1) and 4-(4-acetyl-2-methoxy-phenoxy)-butyric acid (2) and two esters, pentanedioic acid mono-(4-acetyl-2-methoxy-phenyl) ester (3) and heptanedioic acid mono-(4-acetyl-2-methoxy-phenyl) ester (4), which are apocynin derivatives were designed, synthesized and evaluated as NOX inhibitors by quantifying O2•− production using EPR (electron paramagnetic resonance) measurements. In addition, the antioxidant activity of apocynin and its derivatives were determined. A docking study was used to identify the interactions between the NOX2′s p47phox subunit and apocynin or its derivatives. The results showed that all of the compounds exhibit inhibitory activity on NOX, being 4 the best derivative. However, neither apocynin nor its derivatives were free radical scavengers. On the other hand, the in silico studies demonstrated that the apocynin and its derivatives were recognized by the polybasic SH3A and SH3B domains, which are regions of p47phox that interact with p22phox. Therefore this experimental and theoretical study suggests that compound 4 could prevent the formation of the complex between p47phox and p22phox without needing to be activated by MPO (myeloperoxidase), this being an advantage over apocynin.

scholarly journals Signal transduction by glycophorin A: role of extracellular and cytoplasmic domains in a modulatable process.

1988 ◽  
Vol 107 (4) ◽  
pp. 1351-1357 ◽  
Author(s):  
J A Chasis ◽  
M E Reid ◽  
R H Jensen ◽  
N Mohandas
Keyword(s):  
Signal Transduction ◽  
Transmembrane Protein ◽  
Fold Increase ◽  
Extracellular Domain ◽  
Cytoplasmic Domain ◽  
Glycophorin A ◽  
Domain Antibody ◽  
Extracellular Region ◽  
Cytoplasmic Side

Binding of ligands to the extracellular region of the erythrocyte transmembrane protein glycophorin A induces a decrease in membrane deformability. Since the property of membrane deformability is regulated by the skeletal proteins on the cytoplasmic side of the membrane, this suggests that ligand binding may initiate a transmembrane signal. To further study this process, we examined which domains of the extracellular region of glycophorin are involved in signal transduction and whether the cytoplasmic domain of the molecule is necessary for transmitting the signal. Using the ektacytometer, we compared the effect on deformability of four monoclonal antibodies that detect different epitopes on glycophorin A. We found that 9A3 (which recognized the amino terminus of glycophorin) caused a 5.8-fold increase in rigidity, R-10 and 10F7 (which recognized epitopes in the mid-region of the extracellular domain) caused a 10.8-fold increase in rigidity and B14 (which binds to glycophorin close to the membrane) caused a 18-fold increase in rigidity. Further, a direct relationship was observed between the degree of antibody-induced rigidity and the amount of glycophorin A that became associated with the skeletal proteins in a Triton shell assay. In Miltenberger V erythrocytes, which contain a hybrid sialoglycoprotein with no cytoplasmic domain, antibody binding did not induce an increase in rigidity. These results imply that glycophorin A is capable of a modulatable form of transmembrane signaling that is determined by the extracellular domain to which the ligand binds, and the cytoplasmic domain of glycophorin A is crucial for this process.

scholarly journals Promotion of Neurite and Filopodium Formation by CD47: Roles of Integrins, Rac, and Cdc42

2004 ◽  
Vol 15 (8) ◽  
pp. 3950-3963 ◽  
Author(s):  
Motoaki Miyashita ◽  
Hiroshi Ohnishi ◽  
Hideki Okazawa ◽  
Hiroyasu Tomonaga ◽  
Akiko Hayashi ◽  
...  
Keyword(s):  
Transmembrane Protein ◽  
Neuroblastoma Cells ◽  
Axon Extension ◽  
Entire Structure ◽  
Signaling Mechanisms ◽  
Extracellular Region ◽  
Fc Fusion Protein ◽  
Neurite Formation ◽  
Filopodium Formation

Axon extension during development is guided by many factors, but the signaling mechanisms responsible for its regulation remain largely unknown. We have now investigated the role of the transmembrane protein CD47 in this process in N1E-115 neuroblastoma cells. Forced expression of CD47 induced the formation of neurites and filopodia. Furthermore, an Fc fusion protein containing the extracellular region of the CD47 ligand SHPS-1 induced filopodium formation, and this effect was enhanced by CD47 overexpression. SHPS-1–Fc also promoted neurite and filopodium formation triggered by serum deprivation. Inhibition of Rac or Cdc42 preferentially blocked CD47-induced formation of neurites and filopodia, respectively. Overexpression of CD47 resulted in the activation of both Rac and Cdc42. The extracellular region of CD47 was sufficient for the induction of neurite formation by forced expression, but the entire structure of CD47 was required for enhancement of filopodium formation by SHPS-1–Fc. Neurite formation induced by CD47 was also inhibited by a mAb to the integrin β3 subunit. These results indicate that the interaction of SHPS-1 with CD47 promotes neurite and filopodium formation through the activation of Rac and Cdc42, and that integrins containing the β3 subunit participate in the effect of CD47 on neurite formation.

scholarly journals Identification of Most Lethal Non-synonymous SNPs in Tollip gene- An In-Silico Analysis

2021 ◽  
Author(s):  
Usman Ghani ◽  
Rahmat Ullah ◽  
Sadia Anjum ◽  
Yasir Ali ◽  
Saddam Hussain ◽  
...  
Keyword(s):  
Membrane Trafficking ◽  
In Silico ◽  
Interleukin 1 ◽  
Intracellular Localization ◽  
In Silico Analysis ◽  
Gene Interaction ◽  
Intestinal Epithelial ◽  
Interacting Protein ◽  
Synonymous Snps ◽  
Wide Range

Abstract The Toll-interacting protein (TOLLIP), first detected by hybrid screening using Interleukin-1 receptor accessory protein in 2000, is ubiquitous and its TLR signaling cascade gets negatively regulated by TOLLIP in particular by impeding the TLR4 and TLR2 pathways. Toll-interacting protein facilitates TLR and TGF-β type 1 receptor intracellular localization and lysosomal degradation and exerts its anti-apoptosis and pro-autophagy effects through interaction with a target of Myb1 membrane trafficking protein 1 (TOM1) in autoimmunity. It also protects intestinal epithelial cells from apoptosis induced by (TNF-α) (IFN-γ) signaling and acts as a cargo adaptor linking (ATG8) autophagy gene 8 and microtubule-associated protein 1 light chain 3. Ubiquitin-modified cell debris along with coated autophagosomes removes harmful protein aggregates and maintains cellular homeostasis. It is therefore structurally and functionally important to detect TOLLIP polymorphisms to indicate the possible malfunctions and therapeutics. We have identified the gap of available data on nsSNPs in the TOLLIP gene in previous studies. Hence, We have used a wide range of bioinformatic techniques in this study to identify the most destructive nsSNPs in the TOLLIP gene. The in-silico tools such as PROVEAN, SIFT, SNP&GO, PhD SNP, and PolyPhen2 have been used followed by I Mutant, MutPred, and ConSurf. The 3-D mapping was carried out with I-TASSER and Phyre2. Though, STRING and GeneMANIA proposed the gene to gene interaction of TOLLIP. Our study identified G19D (rs866744102), G32R (rs1308704061), D71N (rs777772934), and E72G (rs1202660177) as the four most lethal non-synonymous SNPs in TOLLIP genes, which may play an essential part in defects of TOLLIP Protein and probably cause a different type of diseases. This is the first study of its kind, and it could pave the way in the field of medical drugs in the future. The influence of these nsSNPs in the normal functioning of TOLLIP can also be evaluated in animal models to identify the underlying biochemical pathway involved in diseases.

scholarly journals Mapping transmembrane binding partners for E-cadherin ectodomains

2020 ◽  
Author(s):  
Omer Shafraz ◽  
Bin Xie ◽  
Soichiro Yamada ◽  
Sanjeevi Sivasankar
Keyword(s):  
Single Molecule ◽  
Protein Interactions ◽  
Transmembrane Protein ◽  
Binding Assays ◽  
Binding Interactions ◽  
Adhesion Protein ◽  
Bait Protein ◽  
Binding Partners ◽  
Extracellular Region ◽  
E Cadherin

ABSTRACTWe combine proximity labeling and single molecule binding assays, to discover novel transmembrane protein interactions in cells. We first screen for candidate binding partners by tagging the extracellular and cytoplasmic regions of a bait protein with TurboID biotin ligase, and identify proximal proteins that are biotin-tagged on both their extracellular and intracellular regions. We then test direct binding interactions between the proximal proteins and the bait, using single molecule Atomic Force Microscope binding assays. Using this approach, we identify novel binding partners for the extracellular region of E-cadherin, an essential cell-cell adhesion protein. We show that the desmosomal proteins desmoglein-2 and desmocollin-3, the focal adhesion protein integrin-α2β1, and the receptor tyrosine kinase ligand ephrin-B1, all directly interact with E-cadherin ectodomains. Our discovery of previously unknown heterophilic E-cadherin binding interactions, suggest the existence of novel cadherin cross-talk in epithelial cells.

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