Biophysical characterization of melanoma cell phenotype markers during metastatic progression

AbstractMelanoma is the most fatal form of skin cancer, with increasing prevalence worldwide. The most common melanoma genetic driver is mutation of the proto-oncogene serine/threonine kinase BRAF; thus, the inhibition of its MAP kinase pathway by specific inhibitors is a commonly applied therapy. However, many patients are resistant, or develop resistance to this type of monotherapy, and therefore combined therapies which target other signaling pathways through various molecular mechanisms are required. A possible strategy may involve targeting cellular energy metabolism, which has been recognized as crucial for cancer development and progression and which connects through glycolysis to cell surface glycan biosynthetic pathways. Protein glycosylation is a hallmark of more than 50% of the human proteome and it has been recognized that altered glycosylation occurs during the metastatic progression of melanoma cells which, in turn facilitates their migration. This review provides a description of recent advances in the search for factors able to remodel cell metabolism between glycolysis and oxidative phosphorylation, and of changes in specific markers and in the biophysical properties of cells during melanoma development from a nevus to metastasis. This development is accompanied by changes in the expression of surface glycans, with corresponding changes in ligand-receptor affinity, giving rise to structural features and viscoelastic parameters particularly well suited to study by label-free biophysical methods.

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EGFL9 promotes breast cancer metastasis by inducing cMET activation and metabolic reprogramming

Nature Communications ◽

10.1038/s41467-019-13034-3 ◽

2019 ◽

Vol 10 (1) ◽

Cited By ~ 7

Author(s):

Fanyan Meng ◽

Ling Wu ◽

Lun Dong ◽

Allison V. Mitchell ◽

C. James Block ◽

...

Keyword(s):

Breast Cancer ◽

Molecular Mechanisms ◽

Cancer Metastasis ◽

Cell Metabolism ◽

Breast Cancer Metastasis ◽

Metabolic Reprogramming ◽

Metabolic Phenotype ◽

Migration And Invasion ◽

Metastatic Progression ◽

Basal Like Breast Cancer

Abstract The molecular mechanisms driving metastatic progression in triple-negative breast cancer (TNBC) patients are poorly understood. In this study, we demonstrate that epidermal growth factor-like 9 (EGFL9) is significantly upregulated in basal-like breast cancer cells and associated with metastatic progression in breast tumor samples. Functionally, EGFL9 is both necessary and sufficient to enhance cancer cell migration and invasion, as well as distant metastasis. Mechanistically, we demonstrate that EGFL9 binds cMET, activating cMET-mediated downstream signaling. EGFL9 and cMET co-localize at both the cell membrane and within the mitochondria. We further identify an interaction between EGFL9 and the cytochrome c oxidase (COX) assembly factor COA3. Consequently, EGFL9 regulates COX activity and modulates cell metabolism, promoting a Warburg-like metabolic phenotype. Finally, we show that combined pharmacological inhibition of cMET and glycolysis reverses EGFL9-driven stemness. Our results identify EGFL9 as a therapeutic target for combating metastatic progression in TNBC.

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The Impact of Exercise Serum on Selected Parameters of CD4+ T Cell Metabolism

Immuno ◽

10.3390/immuno1030008 ◽

2021 ◽

Vol 1 (3) ◽

pp. 119-131

Author(s):

Jana Palmowski ◽

Kristina Gebhardt ◽

Thomas Reichel ◽

Torsten Frech ◽

Robert Ringseis ◽

...

Keyword(s):

T Cells ◽

Oxygen Consumption ◽

T Cell ◽

Real Time ◽

Cd4 T Cells ◽

Cell Metabolism ◽

Cd4 T Cell ◽

Autologous Serum ◽

Cell Phenotype ◽

The Impact

CD4+ T cells are sensitive to peripheral changes of cytokine levels and metabolic substrates such as glucose and lactate. This study aimed to analyze whether factors released after exercise alter parameters of human T cell metabolism, specifically glycolysis and oxidative phosphorylation. We used primary human CD4+ T cells activated in the presence of autologous serum, which was collected before (CO) and after a 30-min exercise intervention (EX). In the course of activation, cells and supernatants were analyzed for cell viability and diameter, real-time oxygen consumption by using PreSens Technology, mRNA expression of glycolytic enzymes and complexes of the electron transport chain by real-time PCR, glucose, and lactate levels in supernatants, and in vitro differentiation by flow cytometry. EX did not alter T cell phenotype, viability, or on-blast formation. Similarly, no difference between CO and EX were found for CD4+ T cell activation and cellular oxygen consumption. In contrast, higher levels of glucose were found after 48 h activation in EX conditions. T cells activated in autologous exercise serum expressed lower HK1 mRNA and higher IFN-γ receptor 1. We suggest that the exercise protocol used was not sufficient to destabilize the immune metabolism of T cells. Therefore, more intense and prolonged exercise should be used in future studies.

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Impairing flow-mediated endothelial remodeling reduces extravasation of tumor cells

Scientific Reports ◽

10.1038/s41598-021-92515-2 ◽

2021 ◽

Vol 11 (1) ◽

Author(s):

Gautier Follain ◽

Naël Osmani ◽

Valentin Gensbittel ◽

Nandini Asokan ◽

Annabel Larnicol ◽

...

Keyword(s):

Blood Flow ◽

Tumor Cells ◽

Molecular Mechanisms ◽

Metastatic Progression ◽

Metastatic Dissemination ◽

Secondary Tumors ◽

Flow Profiles ◽

Endothelial Response ◽

Life Threatening

AbstractTumor progression and metastatic dissemination are driven by cell-intrinsic and biomechanical cues that favor the growth of life-threatening secondary tumors. We recently identified pro-metastatic vascular regions with blood flow profiles that are permissive for the arrest of circulating tumor cells. We have further established that such flow profiles also control endothelial remodeling, which favors extravasation of arrested CTCs. Yet, how shear forces control endothelial remodeling is unknown. In the present work, we aimed at dissecting the cellular and molecular mechanisms driving blood flow-dependent endothelial remodeling. Transcriptomic analysis of endothelial cells revealed that blood flow enhanced VEGFR signaling, among others. Using a combination of in vitro microfluidics and intravital imaging in zebrafish embryos, we now demonstrate that the early flow-driven endothelial response can be prevented upon specific inhibition of VEGFR tyrosine kinase and subsequent signaling. Inhibitory targeting of VEGFRs reduced endothelial remodeling and subsequent metastatic extravasation. These results confirm the importance of VEGFR-dependent endothelial remodeling as a driving force of CTC extravasation and metastatic dissemination. Furthermore, the present work suggests that therapies targeting endothelial remodeling might be a relevant clinical strategy in order to impede metastatic progression.

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Membrane traffic related to endosome dynamics and protein secretion in filamentous fungi

Bioscience Biotechnology and Biochemistry ◽

10.1093/bbb/zbab004 ◽

2021 ◽

Author(s):

Yujiro Higuchi

Keyword(s):

Filamentous Fungi ◽

Protein Secretion ◽

Secretory Pathway ◽

Molecular Mechanisms ◽

Membrane Traffic ◽

Early Endosome ◽

Protein Glycosylation ◽

Hyphal Cell ◽

Cell Factories ◽

Membrane Contacts

ABSTRACT In eukaryotic cells, membrane-surrounded organelles are orchestrally organized spatiotemporally under environmental situations. Among such organelles, vesicular transports and membrane contacts occur to communicate each other, so-called membrane traffic. Filamentous fungal cells are highly polarized and thus membrane traffic is developed to have versatile functions. Early endosome (EE) is an endocytic organelle that dynamically exhibits constant long-range motility through the hyphal cell, which is proven to have physiological roles, such as other organelle distribution and signal transduction. Since filamentous fungal cells are also considered as cell factories, to produce valuable proteins extracellularly, molecular mechanisms of secretory pathway including protein glycosylation have been well investigated. In this review, molecular and physiological aspects of membrane traffic especially related to EE dynamics and protein secretion in filamentous fungi are summarized, and perspectives for application are also described.

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The dimer-monomer equilibrium of SARS-CoV-2 main protease is affected by small molecule inhibitors

Scientific Reports ◽

10.1038/s41598-021-88630-9 ◽

2021 ◽

Vol 11 (1) ◽

Author(s):

Lucia Silvestrini ◽

Norhan Belhaj ◽

Lucia Comez ◽

Yuri Gerelli ◽

Antonino Lauria ◽

...

Keyword(s):

Enzymatic Activity ◽

Dissociation Constant ◽

Molecular Mechanisms ◽

Antiviral Drug ◽

Structural Features ◽

Experimental Information ◽

Detailed Knowledge ◽

Main Protease ◽

Equilibrium Dissociation ◽

Drug Leads

AbstractThe maturation of coronavirus SARS-CoV-2, which is the etiological agent at the origin of the COVID-19 pandemic, requires a main protease Mpro to cleave the virus-encoded polyproteins. Despite a wealth of experimental information already available, there is wide disagreement about the Mpro monomer-dimer equilibrium dissociation constant. Since the functional unit of Mpro is a homodimer, the detailed knowledge of the thermodynamics of this equilibrium is a key piece of information for possible therapeutic intervention, with small molecules interfering with dimerization being potential broad-spectrum antiviral drug leads. In the present study, we exploit Small Angle X-ray Scattering (SAXS) to investigate the structural features of SARS-CoV-2 Mpro in solution as a function of protein concentration and temperature. A detailed thermodynamic picture of the monomer-dimer equilibrium is derived, together with the temperature-dependent value of the dissociation constant. SAXS is also used to study how the Mpro dissociation process is affected by small inhibitors selected by virtual screening. We find that these inhibitors affect dimerization and enzymatic activity to a different extent and sometimes in an opposite way, likely due to the different molecular mechanisms underlying the two processes. The Mpro residues that emerge as key to optimize both dissociation and enzymatic activity inhibition are discussed.

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G-protein-coupled receptors signalling at the cell nucleus: an emerging paradigmThis paper is one of a selection of papers published in this Special Issue, entitled The Nucleus: A Cell Within A Cell.

Canadian Journal of Physiology and Pharmacology ◽

10.1139/y05-127 ◽

2006 ◽

Vol 84 (3-4) ◽

pp. 287-297 ◽

Cited By ~ 124

Author(s):

Fernand Gobeil ◽

Audrey Fortier ◽

Tang Zhu ◽

Michela Bossolasco ◽

Martin Leduc ◽

...

Keyword(s):

G Protein ◽

Cell Nucleus ◽

Intracellular Signaling ◽

Molecular Mechanisms ◽

Nuclear Translocation ◽

Cell Metabolism ◽

Hormone Secretion ◽

G Protein Coupled Receptors ◽

A Cell ◽

G Protein Coupled

G-protein-coupled receptors (GPCRs) comprise a wide family of monomeric heptahelical glycoproteins that recognize a broad array of extracellular mediators including cationic amines, lipids, peptides, proteins, and sensory agents. Thus far, much attention has been given towards the comprehension of intracellular signaling mechanisms activated by cell membrane GPCRs, which convert extracellular hormonal stimuli into acute, non-genomic (e.g., hormone secretion, muscle contraction, and cell metabolism) and delayed, genomic biological responses (e.g., cell division, proliferation, and apoptosis). However, with respect to the latter response, there is compelling evidence for a novel intracrine mode of genomic regulation by GPCRs that implies either the endocytosis and nuclear translocation of peripheral-liganded GPCR and (or) the activation of nuclearly located GPCR by endogenously produced, nonsecreted ligands. A noteworthy example of the last scenario is given by heptahelical receptors that are activated by bioactive lipoids (e.g., PGE2 and PAF), many of which may be formed from bilayer membranes including those of the nucleus. The experimental evidence for the nuclear localization and signalling of GPCRs will be reviewed. We will also discuss possible molecular mechanisms responsible for the atypical compartmentalization of GPCRs at the cell nucleus, along with their role in gene expression.

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Temporal Quantitative Proteomics Analysis of Neuroblastoma Cells Treated with Bovine Milk-Derived Extracellular Vesicles Highlights the Anti-Proliferative Properties of Milk-Derived Extracellular Vesicles

Cells ◽

10.3390/cells10040750 ◽

2021 ◽

Vol 10 (4) ◽

pp. 750

Author(s):

Pamali Fonseka ◽

Taeyoung Kang ◽

Sing Chee ◽

Sai V. Chitti ◽

Rahul Sanwlani ◽

...

Keyword(s):

High Risk ◽

Extracellular Vesicles ◽

Quantitative Proteomics ◽

Cell Metabolism ◽

Bovine Milk ◽

Neuroblastoma Cells ◽

Treatment Strategies ◽

Treatment Resistance ◽

Label Free ◽

The Impact

Neuroblastoma (NBL) is a pediatric cancer that accounts for 15% of childhood cancer mortality. Amplification of the oncogene N-Myc occurs in 20% of NBL patients and is considered high risk as it correlates with aggressiveness, treatment resistance and poor prognosis. Even though the treatment strategies have improved in the recent years, the survival rate of high-risk NBL patients remain poor. Hence, it is crucial to explore new therapeutic avenues to sensitise NBL. Recently, bovine milk-derived extracellular vesicles (MEVs) have been proposed to contain anti-cancer properties. However, the impact of MEVs on NBL cells is not understood. In this study, we characterised MEVs using Western blotting, NTA and TEM. Importantly, treatment of NBL cells with MEVs decreased the proliferation and increased the sensitivity of NBL cells to doxorubicin. Temporal label-free quantitative proteomics of NBL cells highlighted the depletion of proteins involved in cell metabolism, cell growth and Wnt signalling upon treatment with MEVs. Furthermore, proteins implicated in cellular senescence and apoptosis were enriched in NBL cells treated with MEVs. For the first time, this study highlights the temporal proteomic profile that occurs in cancer cells upon MEVs treatment.

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When nutrition interacts with osteoblast function: molecular mechanisms of polyphenols

Nutrition Research Reviews ◽

10.1017/s095442240926402x ◽

2009 ◽

Vol 22 (1) ◽

pp. 68-81 ◽

Cited By ~ 42

Author(s):

Anna Trzeciakiewicz ◽

Véronique Habauzit ◽

Marie-Noëlle Horcajada

Keyword(s):

Bone Health ◽

Molecular Mechanisms ◽

Vegetable Intake ◽

Cell Metabolism ◽

Bone Cell ◽

Mitogen Activated Protein Kinase ◽

Present Review ◽

Activator Protein 1 ◽

Cellular Signalling ◽

Osteoblast Function

Recent research has provided insights into dietary components that may optimise bone health and stimulate bone formation. Fruit and vegetable intake, as well as grains and other plant-derived food, have been linked to decreased risk of major chronic diseases including osteoporosis. This effect has been partially attributed to the polyphenols found in these foods. Thus, it has been suggested that these compounds may provide desirable bone health benefits through an action on bone cell metabolism. The present review will focus on how some polyphenols can modulate osteoblast function and reports which cellular signalling pathways are potentially implicated. However, to date, despite numerous investigations, few studies have provided clear evidence that phenolic compounds can act on osteoblasts. Polyphenols cited in the present review seem to be able to modulate the expression of transcription factors such as runt-related transcription factor-2 (Runx2) and Osterix, NF-κB and activator protein-1 (AP-1). It appears that polyphenols may act on cellular signalling such as mitogen-activated protein kinase (MAPK), bone morphogenetic protein (BMP), oestrogen receptor and osteoprotegerin/receptor activator of NF-κB ligand (OPG/RANKL) and thus may affect osteoblast functions. However, it is also important to take in account the possible interaction of these compounds on osteoclast metabolism to better understand the positive correlation reported between the consumption of fruit and vegetables and bone mass.

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Unique structural features of a BCL-2 family protein CED-9 and biophysical characterization of CED-9/EGL-1 interactions

Cell Death and Differentiation ◽

10.1038/sj.cdd.4401303 ◽

2003 ◽

Vol 10 (12) ◽

pp. 1310-1319 ◽

Cited By ~ 30

Author(s):

J-S Woo ◽

J-S Jung ◽

N-C Ha ◽

J Shin ◽

K-H Kim ◽

...

Keyword(s):

Structural Features ◽

Biophysical Characterization ◽

Family Protein

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Quantitative Proteomics Reveals that GmENO2 Proteins Are Involved in Response to Phosphate Starvation in the Leaves of Glycine max L.

International Journal of Molecular Sciences ◽

10.3390/ijms22020920 ◽

2021 ◽

Vol 22 (2) ◽

pp. 920

Author(s):

Ling Cheng ◽

Wanling Min ◽

Man Li ◽

Lili Zhou ◽

Chuan-Chih Hsu ◽

...

Keyword(s):

Glycine Max ◽

Molecular Mechanisms ◽

Human Life ◽

Phosphate Starvation ◽

Limiting Factor ◽

Transcriptional Level ◽

Label Free ◽

Promoter Regions ◽

Glycine Max L ◽

Soybean Plants

Soybean (Glycine max L.) is a major crop providing important source for protein and oil for human life. Low phosphate (LP) availability is a critical limiting factor affecting soybean production. Soybean plants develop a series of strategies to adapt to phosphate (Pi) limitation condition. However, the underlying molecular mechanisms responsible for LP stress response remain largely unknown. Here, we performed a label-free quantification (LFQ) analysis of soybean leaves grown under low and high phosphate conditions. We identified 267 induced and 440 reduced differential proteins from phosphate-starved leaves. Almost a quarter of the LP decreased proteins are involved in translation processes, while the LP increased proteins are accumulated in chlorophyll biosynthetic and carbon metabolic processes. Among these induced proteins, an enolase protein, GmENO2a was found to be mostly induced protein. On the transcriptional level, GmENO2a and GmENO2b, but not GmENO2c or GmENO2d, were dramatically induced by phosphate starvation. Among 14 enolase genes, only GmENO2a and GmENO2b genes contain the P1BS motif in their promoter regions. Furthermore, GmENO2b was specifically induced in the GmPHR31 overexpressing soybean plants. Our findings provide molecular insights into how soybean plants tune basic carbon metabolic pathway to adapt to Pi deprivation through the ENO2 enzymes.

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