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1.
Pseudomonas aeruginosa Triggered Exosomal Release of ADAM10 Mediates Proteolytic Cleavage in Trans.
Aljohmani, A, Opitz, B, Bischoff, M, Yildiz, D
International journal of molecular sciences. 2022;(3)
Abstract
Pneumonia is a life-threatening disease often caused by infection with Streptococcus pneumoniae and Pseudomonas aeruginosa. Many of the mediators (e.g., TNF, IL-6R) and junction molecules (e.g., E-cadherin) orchestrating inflammatory cell recruitment and loss of barrier integrity are proteolytically cleaved through a disintegrin and metalloproteinases (ADAMs). We could show by Western blot, surface expression analysis and measurement of proteolytic activity in cell-based assays, that ADAM10 in epithelial cells is upregulated and activated upon infection with Pseudomonas aeruginosa and Exotoxin A (ExoA), but not upon infection with Streptococcus pneumoniae. Targeting ADAM10 by pharmacological inhibition or gene silencing, we demonstrated that this activation was critical for cleavage of E-cadherin and modulated permeability and epithelial integrity. Stimulation with heat-inactivated bacteria revealed that the activation was based on the toxin repertoire rather than the interaction with the bacterial particle itself. Furthermore, calcium imaging experiments showed that the ExoA action was based on the induction of calcium influx. Investigating the extracellular vesicles and their proteolytic activity, we could show that Pseudomonas aeruginosa triggered exosomal release of ADAM10 and proteolytic cleavage in trans. This newly described mechanism could constitute an essential mechanism causing systemic inflammation in patients suffering from Pseudomonas aeruginosa-induced pneumonia stimulating future translational studies.
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2.
Necrotic debris and STING exert therapeutically relevant effects on tumor cholesterol homeostasis.
Katakam, S, Anand, S, Martin, P, Riggi, N, Stamenkovic, I
Life science alliance. 2022;(3)
Abstract
Malignant tumors commonly display necrosis, which invariably triggers an inflammatory response that supports tumor growth. However, the effect on tumor cells of necrotic debris, or damage-associated molecular patterns (DAMPs) released by dying cells is unknown. Here, we addressed the effect of DAMPs on primary Ewing sarcoma (EwS) cells and cell lines grown in 3D (spheroids) and 2D culture. We show that DAMPs promote the growth of EwS spheroids but not 2D cultures and that the underlying mechanism implicates an increase in cholesterol load in spheroids. In contrast, stimulation of the nucleic acid sensor signaling platform STING by its ligand cyclic GMP-AMP decreases the tumor cell cholesterol load and reduces their tumor initiating ability. Overexpression of STING or stimulation with cyclic GMP-AMP opposes the growth stimulatory effect of DAMPs and synergizes with the cholesterol synthesis inhibitor simvastatin to inhibit tumor growth. Our observations show that modulation of cholesterol homeostasis is a major effect of necrotic cell debris and STING and suggest that combining STING agonists with statins may help control tumor growth.
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3.
Traptamer screening: a new functional genomics approach to study virus entry and other cellular processes.
Xie, J, DiMaio, D
The FEBS journal. 2022;(2):355-362
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Abstract
Historically, the genetic analysis of mammalian cells entailed the isolation of randomly arising mutant cell lines with altered properties, followed by laborious genetic mapping experiments to identify the mutant gene responsible for the phenotype. In recent years, somatic cell genetics has been revolutionized by functional genomics screens, in which expression of every protein-coding gene is systematically perturbed, and the phenotype of the perturbed cells is determined. We outline here a novel functional genomics screening strategy that differs fundamentally from commonly used approaches. In this strategy, we express libraries of artificial transmembrane proteins named traptamers and select rare cells with the desired phenotype because, by chance, a traptamer specifically perturbs the expression or activity of a target protein. Active traptamers are then recovered from the selected cells and can be used as tools to dissect the biological process under study. We also briefly describe how we have used this new strategy to provide insights into the complex process by which human papillomaviruses enter cells.
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4.
Non-metabolic functions of pyruvate kinase M2: PKM2 in tumorigenesis and therapy resistance.
İlhan, M
Neoplasma. 2022;(4):747-754
Abstract
Cancer is the disease of uncontrollably dividing cells in the body. As cancer cells proliferate at higher rates, they need more energy in a short time necessitating deregulation of energy-generating pathways for their benefit. Although oxidative phosphorylation generates more energy from a glucose molecule, cancer cells have a tendency to enhance aerobic glycolysis by consuming more glucose and producing lactate as a by-product even if oxygen is present. In addition to the generation of rapid energy to fulfill their increasing demands, this strategy also provides the use of glucose metabolites such as lactate as a source for the synthesis of anabolic molecules, such as nucleotides, amino acids, and lipids during the rapid phase of the proliferation. Pyruvate kinase M2 (PKM2) is an isoform of pyruvate kinase, which mediates the balancing of energy generation mechanisms during the anabolic and catabolic events. Due to its vital role in glycolysis, PKM2 has been investigated to target cancer cell metabolism for several years. However, recent studies demonstrate that PKM2 may also promote cancer progression by regulating core steps in metastasis such as migration, angiogenesis, and stemness. Of note, it is estimated that 90% of cancer-related deaths are due to metastasis. This review is intended to summarize the recent advances in the non-metabolic roles of PKM2 in cancer progression and to indicate its potential uses for the development of new treatment strategies.
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5.
The roles of transmembrane family proteins in the regulation of store-operated Ca2+ entry.
Zhang, N, Pan, H, Liang, X, Xie, J, Han, W
Cellular and molecular life sciences : CMLS. 2022;(2):118
Abstract
Store-operated Ca2+ entry (SOCE) is a major pathway for calcium signaling, which regulates almost every biological process, involving cell proliferation, differentiation, movement and death. Stromal interaction molecule (STIM) and ORAI calcium release-activated calcium modulator (ORAI) are the two major proteins involved in SOCE. With the deepening of studies, more and more proteins are found to be able to regulate SOCE, among which the transmembrane (TMEM) family proteins are worth paying more attention. In addition, the ORAI proteins belong to the TMEM family themselves. As the name suggests, TMEM family is a type of proteins that spans biological membranes including plasma membrane and membrane of organelles. TMEM proteins are in a large family with more than 300 proteins that have been already identified, while the functional knowledge about the proteins is preliminary. In this review, we mainly summarized the TMEM proteins that are involved in SOCE, to better describe a picture of the interaction between STIM and ORAI proteins during SOCE and its downstream signaling pathways, as well as to provide an idea for the study of the TMEM family proteins.
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6.
PIP aquaporin pH-sensing is regulated by the length and charge of the C-terminal region.
Scochera, F, Zerbetto De Palma, G, Canessa Fortuna, A, Chevriau, J, Toriano, R, Soto, G, Zeida, A, Alleva, K
The FEBS journal. 2022;(1):246-261
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Abstract
Plant PIP aquaporins play a central role in controlling plant water status. The current structural model for PIP pH-gating states that the main pH sensor is located in loopD and that all the mobile cytosolic elements participate in a complex interaction network that ensures the closed structure. However, the precise participation of the last part of the C-terminal domain (CT) in PIP pH gating remains unknown. This last part has not been resolved in PIP crystal structures and is a key difference between PIP1 and PIP2 paralogues. Here, by a combined experimental and computational approach, we provide data about the role of CT in pH gating of Beta vulgaris PIP. We demonstrate that the length of CT and the positive charge located among its last residues modulate the pH at which the open/closed transition occurs. We also postulate a molecular-based mechanism for the differential pH sensing in PIP homo- or heterotetramers by performing atomistic molecular dynamics simulations (MDS) on complete models of PIP tetramers. Our findings show that the last part of CT can affect the environment of loopD pH sensors in the closed state. Results presented herein contribute to the understanding of how the characteristics of CT in PIP channels play a crucial role in determining the pH at which water transport through these channels is blocked, highlighting the relevance of the differentially conserved very last residues in PIP1 and PIP2 paralogues.
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7.
Advanced Glycation End Products: A Sweet Flavor That Embitters Cardiovascular Disease.
Pinto, RS, Minanni, CA, de Araújo Lira, AL, Passarelli, M
International journal of molecular sciences. 2022;(5)
Abstract
Epidemiological studies demonstrate the role of early and intensive glycemic control in the prevention of micro and macrovascular disease in both type 1 and type 2 diabetes mellitus (DM). Hyperglycemia elicits several pathways related to the etiopathogenesis of cardiovascular disease (CVD), including the generation of advanced glycation end products (AGEs). In this review, we revisit the role played by AGEs in CVD based in clinical trials and experimental evidence. Mechanistic aspects concerning the recognition of AGEs by the advanced glycosylation end product-specific receptor (AGER) and its counterpart, the dolichyl-diphosphooligosaccharide-protein glycosyltransferase (DDOST) and soluble AGER are discussed. A special focus is offered to the AGE-elicited pathways that promote cholesterol accumulation in the arterial wall by enhanced oxidative stress, inflammation, endoplasmic reticulum stress and impairment in the reverse cholesterol transport (RCT).
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8.
Bone marrow stromal cells from MDS and AML patients show increased adipogenic potential with reduced Delta-like-1 expression.
Weickert, MT, Hecker, JS, Buck, MC, Schreck, C, Rivière, J, Schiemann, M, Schallmoser, K, Bassermann, F, Strunk, D, Oostendorp, RAJ, et al
Scientific reports. 2021;(1):5944
Abstract
Myelodysplastic syndromes (MDS) and acute myeloid leukemia (AML) are clonal hematopoietic stem cell disorders with a poor prognosis, especially for elderly patients. Increasing evidence suggests that alterations in the non-hematopoietic microenvironment (bone marrow niche) can contribute to or initiate malignant transformation and promote disease progression. One of the key components of the bone marrow (BM) niche are BM stromal cells (BMSC) that give rise to osteoblasts and adipocytes. It has been shown that the balance between these two cell types plays an important role in the regulation of hematopoiesis. However, data on the number of BMSC and the regulation of their differentiation balance in the context of hematopoietic malignancies is scarce. We established a stringent flow cytometric protocol for the prospective isolation of a CD73+ CD105+ CD271+ BMSC subpopulation from uncultivated cryopreserved BM of MDS and AML patients as well as age-matched healthy donors. BMSC from MDS and AML patients showed a strongly reduced frequency of CFU-F (colony forming unit-fibroblast). Moreover, we found an altered phenotype and reduced replating efficiency upon passaging of BMSC from MDS and AML samples. Expression analysis of genes involved in adipo- and osteogenic differentiation as well as Wnt- and Notch-signalling pathways showed significantly reduced levels of DLK1, an early adipogenic cell fate inhibitor in MDS and AML BMSC. Matching this observation, functional analysis showed significantly increased in vitro adipogenic differentiation potential in BMSC from MDS and AML patients. Overall, our data show BMSC with a reduced CFU-F capacity, and an altered molecular and functional profile from MDS and AML patients in culture, indicating an increased adipogenic lineage potential that is likely to provide a disease-promoting microenvironment.
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9.
The Interactome of the VAP Family of Proteins: An Overview.
James, C, Kehlenbach, RH
Cells. 2021;(7)
Abstract
Membrane contact sites (MCS) are sites of close apposition of two organelles that help in lipid transport and synthesis, calcium homeostasis and several other biological processes. The VAMP-associated proteins (VAPs) VAPA, VAPB, MOSPD2 and the recently described MOSPD1 and MOSPD3 are tether proteins of MCSs that are mainly found at the endoplasmic reticulum (ER). VAPs interact with various proteins with a motif called FFAT (two phenylalanines in an acidic tract), recruiting the associated organelle to the ER. In addition to the conventional FFAT motif, the recently described FFNT (two phenylalanines in a neutral tract) and phospho-FFAT motifs contribute to the interaction with VAPs. In this review, we summarize and compare the recent interactome studies described for VAPs, including in silico and proximity labeling methods. Collectively, the interaction repertoire of VAPs is very diverse and highlights the complexity of interactions mediated by the different FFAT motifs to the VAPs.
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10.
Role of candidate gene variants in modulating the risk and severity of alcoholic hepatitis.
Beaudoin, JJ, Liang, T, Tang, Q, Banini, BA, Shah, VH, Sanyal, AJ, Chalasani, NP, Gawrieh, S
Alcoholism, clinical and experimental research. 2021;(4):709-719
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Abstract
BACKGROUND Alcoholic hepatitis (AH) is a severe and life-threatening alcohol-associated liver disease. Only a minority of heavy drinkers acquires AH and severity varies among affected individuals, suggesting a genetic basis for the susceptibility to and severity of AH. METHODS A cohort consisting of 211 patients with AH and 176 heavy drinking controls was genotyped for five variants in five candidate genes that have been associated with chronic liver diseases: rs738409 in patatin-like phospholipase domain-containing protein 3 (PNPLA3), rs72613567 in hydroxysteroid 17-beta dehydrogenase 13 (HSD17B13), rs58542926 in transmembrane 6 superfamily member 2 (TM6SF2), rs641738 in membrane bound O-acyltransferase domain containing 7 (MBOAT7), and a copy number variant in the haptoglobin (HP) gene. We tested the effects of individual variants and the combined/interacting effects of variants on AH risk and severity. RESULTS We found significant associations between AH risk and the risk alleles of rs738409 (p = 0.0081) and HP (p = 0.0371), but not rs72613567 (p = 0.3132), rs58542926 (p = 0.2180), or rs641738 (p = 0.7630), after adjusting for patient's age and sex. A multiple regression model indicated that PNPLA3 rs738409:G [OR = 1.59 (95% CI: 1.15-2.22), p = 0.0055] and HP*2 [OR = 1.38 (95% CI: 1.04-1.82), p = 0.0245], when combined and adjusted for age and sex also had a large influence on AH risk among heavy drinkers. In the entire cohort, variants in PNPLA3 and HP were associated with increased total bilirubin and Model for End-stage Liver Disease (MELD) score, both measures of AH severity. The HSD17B13 rs72613567:AA allele was not found to reduce risk of AH in patients carrying the G allele of PNPLA3 rs738409 (p = 0.0921). CONCLUSION PNPLA3 and HP genetic variants increase AH risk and are associated with total bilirubin and MELD score, surrogates of AH severity.