Identification of Band 3-like Proteins and Cl-/HCO3- Exchange in Isolated Cardiomyocytes

Pucéat, Michel; Korichneva, Irina; Cassoly, Robert; Vassort, Guy

doi:10.1074/jbc.270.3.1315

Cited by 51 publications

(63 citation statements)

References 39 publications

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“…In the present work, we demonstrate that GABA B R1 and GABA B R2 are actually colocalized, functional, and identically distributed along the sarcolemma and the t-system of cardiomyocytes (Fig. 4), following a pattern very similar to that found for other cardiac sarcolemmal proteins (32,45).…”

Section: Discussionsupporting

confidence: 77%

“…Ventricular cardiomyocytes from Wistar rats (250 g) were isolated as described (32). Whole-cell patch-clamp recording was performed with soft glass capillaries pulled to a tip resistance of 1.5-2 M⍀ (Sutter Instruments, Novato, CA).…”

Section: Methodsmentioning

confidence: 99%

“…Ab174.1 was directed against carboxylterminal epitopes of GABA B R1a͞b (33), whereas AbC22 was raised against amino acids 806-907 of the carboxyl-terminal sequence of GABA B R2 (17). Immunoblots with or without coimmunoprecipitation were subsequently probed with either antibody according to previously described techniques (32).…”

Section: Methodsmentioning

confidence: 99%

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γ-Aminobutyric acid type B receptors are expressed and functional in mammalian cardiomyocytes

Lorente

Lacampagne

Pouzeratte

et al. 2000

Proc. Natl. Acad. Sci. U.S.A.

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␥-Hydroxybutyrate (GHB), an anesthetic adjuvant analog of ␥-aminobutyrate (GABA), depresses cell excitability in hippocampal neurons by inducing hyperpolarization through the activation of a prominent inwardly rectifying K ؉ (Kir3) conductance. These GABA type B (GABAB)-like effects are clearly shown at high concentrations of GHB corresponding to blood levels usually reached during anesthesia and are mimicked by the GABAB agonist baclofen. Recent studies of native GABAB receptors (GABABRs) have favored the concept that GHB is also a selective agonist. Furthermore, cloning has demonstrated that GABABRs assemble heteromeric complexes from the GABABR1 and GABABR2 subtypes and that these assemblies are activated by GHB. The surprisingly high tissue content, together with anti-ischemic and protective effects of GHB in the heart, raises the question of a possible influence of GABAB agonists on excitable cardiac cells. In the present study, we provide electrophysiological evidence that GHB activates an inwardly rectifying K ؉ current in rat ventricular myocytes. This effect is mimicked by baclofen, reversibly inhibited by GABAB antagonists, and prevented by pertussis toxin pretreatment. Both GABA BR1 and GABABR2 are detected in cardiomyocytes by Western blotting and are shown to coimmunoprecipitate. Laser scanning confocal microscopy discloses an even distribution of the two receptors in the sarcolemma and along the transverse tubular system. Hence, we conclude that GABABRs are distributed not only in neuronal tissues but also in the heart, where they can be activated and induce electrophysiological alterations through G-protein-coupled inward rectifier potassium channels.

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Section: Discussionsupporting

confidence: 77%

Section: Methodsmentioning

confidence: 99%

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γ-Aminobutyric acid type B receptors are expressed and functional in mammalian cardiomyocytes

Lorente

Lacampagne

Pouzeratte

et al. 2000

Proc. Natl. Acad. Sci. U.S.A.

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“…4 (11). After a 15-min incubation, the reaction was quenched with buffer (192 mM glycine, 25 mM Tris-HCl, pH 7.5, 0.1 mM CaCl 2 , 1 mM MgSO 4 ), and oocytes were washed in MBS, pH 8.…”

Section: Methodsmentioning

confidence: 99%

“…These two entities seem to function independently: the large cytoplasmic domain is involved in interactions with enzymes, hemoglobin, and structural proteins, whereas the membrane-spanning domain is responsible for the transport activity of the protein (2,3). In addition to red cells, AE1 is also expressed in kidney ␣-intercalated cells and cardiac myocytes (4). In physiological conditions, AE1 exchanges one chloride for one bicarbonate by an electroneutral transport mechanism with the driving force for ion movement provided by their electrochemical gradient.…”

mentioning

confidence: 99%

Structural Model of the Anion Exchanger 1 (SLC4A1) and Identification of Transmembrane Segments Forming the Transport Site

Barneaud-Rocca

Etchebest

Guizouarn

2013

Journal of Biological Chemistry

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Background: There is still no high resolution three-dimensional structure available for the membrane-spanning domain of anion exchanger 1 (AE1). Results: A three-dimensional model of AE1 membrane-spanning domain has been generated in silico and experimentally assessed. Conclusion: Transmembrane segments forming AE1 transport site have been identified. Significance: This is the first three-dimensional model of AE1 membrane-spanning domain based on a cation symporter.The anion exchanger 1 (AE1), a member of bicarbonate transporter family SLC4, mediates an electroneutral chloride/bicarbonate exchange in physiological conditions. However, some point mutations in AE1 membrane-spanning domain convert the electroneutral anion exchanger into a Na ؉ and K ؉ conductance or induce a cation leak in a still functional anion exchanger. The molecular determinants that govern ion movement through this transporter are still unknown. The present study was intended to identify the ion translocation pathway within AE1. In the absence of a resolutive three-dimensional structure of AE1 membrane-spanning domain, in silico modeling combined with site-directed mutagenesis experiments was done. A structural model of AE1 membrane-spanning domain is proposed, and this model is based on the structure of a uracilproton symporter. This model was used to design cysteine-scanning mutagenesis on transmembrane (TM) segments 3 and 5. By measuring AE1 anion exchange activity or cation leak, it is proposed that there is a unique transport site comprising TM3-5 and TM8 that should function as an anion exchanger and a cation leak.The anion exchanger 1 (AE1, 2 SLC4A1, or band 3) is the main membrane protein in vertebrate red cells. It fulfills different tasks in these cells: a structural role by linking plasma membrane to cytoskeleton, a respiratory role by improving CO 2 transport capacity of red cells, and an antigenic function (Diego blood group and senescence), and it is also involved in cytokinesis and red cell volume regulation (1). At the molecular level, this protein is divided into two main structural domains: a cytoplasmic N-terminal domain (about 400 amino acids) and a membrane-spanning domain (about 450 amino acids) with a short C-terminal tail in the cytoplasm. These two entities seem to function independently: the large cytoplasmic domain is involved in interactions with enzymes, hemoglobin, and structural proteins, whereas the membrane-spanning domain is responsible for the transport activity of the protein (2, 3). In addition to red cells, AE1 is also expressed in kidney ␣-intercalated cells and cardiac myocytes (4). In physiological conditions, AE1 exchanges one chloride for one bicarbonate by an electroneutral transport mechanism with the driving force for ion movement provided by their electrochemical gradient. A few years ago, some specific point mutations were characterized and proposed to convert the electroneutral anion exchanger into a cation-conductive pathway (5, 6). These mutations are associated with human pathologies (hered...

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Multiple G-protein involvement in parathyroid hormone regulation of acid production by osteoclasts

May

1997

J. Cell. Biochem.

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The involvement of multiple G-proteins in parathyroid hormone regulation of acid production was demonstrated in a highly enriched osteoclast population. Osteoclasts were isolated from the endosteum of 2.5 to 3-week-old chicken tibia using sequential enzymatic digestion. Single cell analysis of acid production was accomplished using microscope photometry and vital staining with acridine orange, a hydrogen ion concentration sensitive fluorescent dye. Lithium chloride, an uncoupler of G-proteins from their respective receptors, blocked parathyroid hormone stimulated production of acid. Cholera toxin, which permanently activates Gs-proteins, mimicked PTH stimulation. Pertussis toxin, which prevents receptor interaction with Gi- and Go-proteins, blocked both 10(-8) M and 10(-11) M PTH stimulated acid production, suggesting that the pertussis toxin-sensitive G-protein is utilized at both PTH concentrations. Immunoblots of osteoclast plasma membrane proteins, using a panel of antibodies generated against specific G-protein alpha subunits, revealed a 48 kDa Gs alpha, a 41 kDa Go alpha, a 34 kDa Gi alpha-3, and a unique 68 kDa G alpha subunit, with the 41 kDa and 34 kDa bands being the most intense. Immunoblots of osteoblast plasma membrane proteins had a substantially different profile with the most intense bands being a Gs alpha (48 kDa) and a Go alpha (36 and 38 kDa). The studies suggest the utilization of at least two different G-proteins in the parathyroid hormone regulation of acid formation by osteoclasts, a Gs and a pertussis toxin-sensitive G-protein (Go and/or Gi alpha-3).

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Identification of Band 3-like Proteins and Cl-/HCO3- Exchange in Isolated Cardiomyocytes

Cited by 51 publications

References 39 publications

γ-Aminobutyric acid type B receptors are expressed and functional in mammalian cardiomyocytes

γ-Aminobutyric acid type B receptors are expressed and functional in mammalian cardiomyocytes

Structural Model of the Anion Exchanger 1 (SLC4A1) and Identification of Transmembrane Segments Forming the Transport Site

Multiple G-protein involvement in parathyroid hormone regulation of acid production by osteoclasts

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