Molecular mechanism of α-tocopheryl-phosphate transport across the cell membrane

Adenylate cyclase toxin (ACT or CyaA) plays a crucial role in respiratory tract colonization and virulence of the whooping cough causative bacteriumBordetella pertussis. Secreted as soluble protein, it targets myeloid cells expressing the CD11b/CD18 integrin and on delivery of its N-terminal adenylate cyclase catalytic domain (AC domain) into the cytosol, generates uncontrolled toxic levels of cAMP that ablates bactericidal capacities of phagocytes. Our study deciphers the fundamentals of the heretofore poorly understood molecular mechanism by which the ACT enzyme domain directly crosses the host cell membrane. By combining molecular biology, biochemistry, and biophysics techniques, we discover that ACT has intrinsic phospholipase A (PLA) activity, and that such activity determines AC translocation. Moreover, we show that elimination of the ACT–PLA activity abrogates ACT toxicity in macrophages, particularly at toxin concentrations close to biological reality of bacterial infection. Our data support a molecular mechanism in which in situ generation of nonlamellar lysophospholipids by ACT–PLA activity into the cell membrane would form, likely in combination with membrane-interacting ACT segments, a proteolipidic toroidal pore through which AC domain transfer could directly take place. Regulation of ACT–PLA activity thus emerges as novel target for therapeutic control of the disease.

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Sulphate and phosphate transport in the renal proximal tubule

Philosophical Transactions of the Royal Society of London Series B Biological Sciences ◽

10.1098/rstb.1982.0151 ◽

1982 ◽

Vol 299 (1097) ◽

pp. 549-558 ◽

Cited By ~ 40

Keyword(s):

Cell Membrane ◽

Proximal Tubule ◽

Brush Border ◽

Inorganic Phosphate ◽

Brush Border Membrane ◽

Ph Dependence ◽

Membrane Vesicles ◽

Dicarboxylic Acids ◽

Phosphate Transport ◽

Brush Border Membrane Vesicles

Experiments performed on microperfused proximal tubules and brush-border membrane vesicles revealed that inorganic phosphate is actively reabsorbed in the proximal tubule involving a 2 Na + -HPO 2- 4 or H 2 PO 4 - co-transport step in the brush-border membrane and a sodium-independent exit step in the basolateral cell membrane. Na + - phosphate co-transport is competitively inhibited by arsenate. The transtubular transport regulation is mirrored by the brush-border transport step: it is inhibited by parathyroid hormone intracellularly mediated by cyclic AMP. Transepithelial inorganic phosphate (P i ) transport and Na + -dependent P i transport across the brush-border membrane correlates inversely with the P i content of the diet. Intraluminal acidification as well as intracellular alkalinization led to a reduction of transepithelial P i transport. Data from brush-border membrane vesicles indicate that high luminal H + concentrations reduce the affinity for Na + of the Na + -phosphate co-transport system, and that this mechanism might be responsible for the pH dependence of phosphate reabsorption. Contraluminal influx of P i from the interstitium into the cell could be partly inhibited by 4,4'-diisothiocyanostilbene-2,2'-disulphonic acid (DIDS). It is not, however, changed when dicarboxylic acids are present or when the pH of the perfusate is reduced to pH 6. Sulphate is actively reabsorbed, involving electroneutral 2 Na + -SO 2 - 4 co-transport through the brush-border membrane. This transport step is inhibited by thiosulphate and molybdate, but not by phosphate or tungstate. The transtubular active sulphate reabsorption is not pH dependent, but is diminished by the absence of bicarbonate. The transport of sulphate through the contraluminal cell side is inhibited by DIDS and diminished when the capillary perfusate contains no bicarbonate or chloride. The latter data indicate the presence of an anion exchange system in the contraluminal cell membrane like that in the erythrocyte membrane.

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Molecular Mechanism of the Cell Membrane Pore Formation Induced by Bubble Stable Cavitation

The Journal of Physical Chemistry B ◽

10.1021/acs.jpcb.8b09391 ◽

2018 ◽

Vol 123 (1) ◽

pp. 71-78 ◽

Cited By ~ 8

Author(s):

Viet Hoang Man ◽

Phan Minh Truong ◽

Mai Suan Li ◽

Junmei Wang ◽

Nguyen-Thi Van-Oanh ◽

...

Keyword(s):

Cell Membrane ◽

Molecular Mechanism ◽

Pore Formation ◽

Membrane Pore

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Molecular mechanism of pyroptosis of mononuclear macrophages induced by pathogenic E. coli high pathogenicity island (HPI) in Yunnan Saba pigs

10.21203/rs.2.24631/v1 ◽

2020 ◽

Author(s):

Chunlan Shan ◽

Shushu Miao ◽

Chaoying Liu ◽

Weiwei Zhao ◽

Bo Zhang ◽

...

Keyword(s):

Cell Membrane ◽

Molecular Mechanism ◽

Pathogenicity Island ◽

Control Group ◽

Tunel Staining ◽

Membrane Pore ◽

Expression Levels ◽

E Coli ◽

Caspase 1 ◽

High Pathogenicity

Abstract Background In this study we evaluated the molecular mechanism by which pyroptosis is induced in mononuclear macrophages isolated from Saba pigs following infection with pathogenic E. coli high pathogenicity island (HPI). Mononuclear macrophages were divided into four treatment groups: control, Lipopolysaccharide (LPS) + adenosine triphosphate (ATP), HPI positive (+) strain and HPI negative (-) strain. The mononuclear macrophages and their culture supernatants were collected at 0.5, 3, 6, 9, 12 and 24 h after infection. DNA changes were detected by TUNEL staining and the integrity of the cell membrane was evaluated by propidium iodide (PI) staining. Changes in mRNA expression levels of NLRP3, caspase-1, IL-1β, and IL-18 gene in mononuclear macrophages were analyzed by quantitative real-time polymerase chain reaction (RT-PCR) and caspase-1 protein expression was detected by indirect immunofluorescence. IL-1β and IL-18 concentration in the mononuclear macrophage culture supernatant were measured by ELISA. Results Compared with the control group, TUNEL and PI staining of mononuclear macrophages was significantly increased following infection with the HPI + /HPI - strains ( P < 0.01 or P < 0.05), with significantly higher levels detected in the HPI + group compared with those in the HPI - group ( P < 0.01 and P < 0.05). Compared with the control group, the expression levels of NLRP3, caspase-1, IL-1β, and IL-18 in the HPI groups were upregulated after pathogenic E. coli infection, with significantly higher levels detected in the HPI + group compared with those in the HPI - group ( P < 0.01 or P < 0.05). Conclusions These findings showed that pathogenic E. coli HPI infection of Saba pigs results induced pyroptosis of mononuclear macrophages characterized by increased expression of NLRP3, caspase-1, IL-1β and IL-18 mRNA in mononuclear macrophages, the induction of cell membrane pore formation, nuclear DNA damage, and the secretion of IL-1β and IL-18 to enhance the inflammatory response.

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Inorganic phosphate transport across the red blood cell membrane: the effect of exposure to hyperoxia

Clinica Chimica Acta ◽

10.1016/0009-8981(87)90345-7 ◽

1987 ◽

Vol 167 (3) ◽

pp. 259-265 ◽

Cited By ~ 6

Author(s):

Nigel R. Webster ◽

Colin Toothill

Keyword(s):

Cell Membrane ◽

Blood Cell ◽

Red Blood Cell ◽

Inorganic Phosphate ◽

Phosphate Transport ◽

Red Blood Cell Membrane

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Sarcolemmal permeability changes during early myocardial anoxia

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100084089 ◽

1978 ◽

Vol 36 (2) ◽

pp. 642-643

Author(s):

M. Ashraf ◽

L. Landa ◽

L. Nimmo ◽

C. M. Bloor

Keyword(s):

Cell Membrane ◽

Membrane Permeability ◽

Coronary Artery Occlusion ◽

Artery Occlusion ◽

Cell Membrane Permeability ◽

Enzyme Release ◽

Sprague Dawley ◽

Sprague Dawley Rats ◽

Sarcolemmal Permeability ◽

Membrane Changes

Following coronary artery occlusion, the myocardial cells lose intracellular enzymes that appear in the serum 3 hrs later. By this time the cells in the ischemic zone have already undergone irreversible changes, and the cell membrane permeability is variably altered in the ischemic cells. At certain stages or intervals the cell membrane changes, allowing release of cytoplasmic enzymes. To correlate the changes in cell membrane permeability with the enzyme release, we used colloidal lanthanum (La+++) as a histological permeability marker in the isolated perfused hearts. The hearts removed from sprague-Dawley rats were perfused with standard Krebs-Henseleit medium gassed with 95% O2 + 5% CO2. The hypoxic medium contained mannitol instead of dextrose and was bubbled with 95% N2 + 5% CO2. The final osmolarity of the medium was 295 M osmol, pH 7. 4.

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