Calcium, ATP and nuclear pore channel gating

Bustamante, J. O.; Michelette, E.R.F.; Geibel, John P.; Dean, David A.; Hanover, John A.; McDonnell, Timothy J.

doi:10.1007/s004249900189

Cited by 33 publications

(27 citation statements)

References 47 publications

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“…The latter maintain the membrane potential at hyperpolarizing potentials and the driving force for Cl Ϫ efflux thereby ensuring efficient ion and water movement (12) (32)(33)(34)(35). Several reports have suggested that these observations are due to modulation of the nuclear pore complex (36,37), differential nuclear Ca 2ϩ buffering (35,38), or even nuclear dye artifacts (17). Although we cannot completely rule out these effects, an important observation from the present study is that inhibition of mitochondrial Ca 2ϩ uptake completely abolishes the nuclear [Ca 2ϩ ] delay.…”

Section: Discussionmentioning

confidence: 99%

Modulation of [Ca2+] Signaling Dynamics and Metabolism by Perinuclear Mitochondria in Mouse Parotid Acinar Cells

Bruce

Giovannucci²,

Blinder

et al. 2004

Journal of Biological Chemistry

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[Ca 2ϩ ] i signals may be tuned to evoke specific physiological responses in these cells. The major function of pancreatic acinar cells is the exocytosis of zymogen granules that can be activated by apically confined [Ca 2ϩ ] i signals (6), suggested to be the major physiological [Ca 2ϩ ] i signal evoked by threshold agonist concentrations (7). These apically confined [Ca 2ϩ ] i signals are in part because of the distribution of mitochondria, which form a belt around the apically located zymogen granules (8, 9). These perigranular mitochondria likely serve two main functions. First, mitochondrial Ca 2ϩ uptake provides a buffer barrier thereby preventing the spread of [Ca 2ϩ ] i waves to the basal part of the cell (8 -10). Second, the Ca 2ϩ taken up by mitochondria drives ATP production locally for the energy-consuming exocytotic process (11). The major function of parotid acinar cells is fluid secretion, and this is most effectively activated by rapid global [Ca 2ϩ ] i signals. We proposed that rapid global [Ca 2ϩ ] i signals facilitate the almost simultaneous activation of Ca 2ϩ -dependent Cl Ϫ channels on the apical membrane and Ca 2ϩ -dependent K ϩ channels on the basolateral membrane (5). The latter maintain the membrane potential at hyperpolarizing potentials and the driving force for Cl Ϫ efflux, thereby ensuring efficient ion and water movement (12). Interestingly, even at threshold stimulation parotid acinar cells exhibit rapid global [Ca 2ϩ ] i signals, and apically confined [Ca 2ϩ ] i signals were never observed (5). Given the many similarities between parotid and pancreatic acinar cells, the differences in the spatial and temporal patterns of [Ca 2ϩ ] i signaling between these two cell types 1 The abbreviations used are: [Ca 2ϩ ] i , intracellular calcium concentration; CCh, carbamylcholine (carbachol); FCCP, carbonyl cyanide 4-trifluoro-methoxyphenylhydrazone; OGB2, Oregon Green BAPTA 2; InsP 3 , inositol 1,4,5-trisphosphate; caged InsP 3 , D-myo-InsP 3 P-4(5)-1-(2-nitrophenyl)-ethyl ester; CaM, calmodulin; RuRed, ruthenium red; NPC, nuclear pore complex; rhod, rhodamine; TRITC, tetramethylrhodamine isothiocyanate.

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Section: Discussionmentioning

confidence: 99%

Modulation of [Ca2+] Signaling Dynamics and Metabolism by Perinuclear Mitochondria in Mouse Parotid Acinar Cells

Bruce

Giovannucci²,

Blinder

et al. 2004

Journal of Biological Chemistry

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“…However, quantitative comparison of structural AFM data with the functional NEER experiments indicates that most likely a certain percentage of NPCs (about 20%) become electrically tight for a short period (a few minutes) and then are again open for inorganic ion movements. A similar model was developed by Bustamante and coworkers (14,36) applying patch-clamp techniques to the NE. The early NEER peak was followed by a marked decrease in the envelope resistance.…”

Section: Discussionmentioning

confidence: 99%

Aldosterone signaling pathway across the nuclear envelope

Schäfer

Shahin

Albermann

et al. 2002

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

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We describe the route by which aldosterone-triggered macromolecules enter and exit the cell nucleus of Xenopus laevis oocyte. Oocytes were microinjected with 50 fmol aldosterone and then enucleated 2-30 min after injection. After isolation, nuclear envelope electrical resistance (NEER) was measured in the intact cell nuclei by using the nuclear hourglass technique. We observed three NEER stages: an early peak 2 min after injection, a sustained depression after 5-15 min, and a final late peak 20 min after injection. Because NEER reflects the passive electrical permeability of nuclear pores, we investigated with atomic force microscopy aldosterone-induced conformational changes of individual nuclear pore complexes (NPCs). At the early peak we observed small (Х100 kDa) molecules (flags) attached to the NPC surface. At the sustained depression NPCs were found free of flags. At the late peak large (Х800 kDa) molecules (plugs) were detected inside the central channels. Ribonuclease or actinomycin D treatment prevented the late NEER peak. Coinjection of aldosterone (50 fmol) and its competitive inhibitor spironolactone (500 fmol) eliminated the electrical changes as well as flag and plug formation. We conclude: (i) The genomic response of aldosterone can be electrically measured in intact oocyte nuclei. (ii) Flags represent aldosterone receptors on their way into the cell nucleus whereas plugs represent ribonucleoproteins carrying aldosterone-induced mRNA from the nucleoplasm into the cytoplasm. (iii) Because plugs can be mechanically harvested with the atomic force microscopy stylus, oocytes could serve as a bioassay system for identifying aldosterone-induced early genes.nuclear pore complex ͉ atomic force microscopy ͉ RNA export ͉ mineralocorticoid receptor S ignals such as an increase in intracellular calcium concentration or intracellular alkalinization are the early responses of target cells to the mineralocorticoid hormone aldosterone. The intracellular calcium signal can be detected a few seconds after hormone exposure (1-3), whereas the pH signal occurs later but usually lasts longer (4). Although there are no doubts about the existence of these early hormone responses, the physiological relevance is still unknown. For almost 40 years it has been postulated that inorganic ions could play a crucial role in steroid hormone-induced gene activation (5). This suggestion has been supported by more recent findings in our own laboratory (6, 7). The signaling pathway of steroid hormones involving intracellular macromolecules is well established. Aldosterone is known to bind to mineralocorticoid receptors located in the cytosol. Hormone binding changes protein conformation and translocates the receptor into the nucleus where it attaches to specific DNA. Transcription occurs and finally mRNA is exported to be translated into aldosterone-induced proteins at the ribosomes. Although this macromolecular signal pathway has been described in detail, there are no data available that directly show both functionally and structural...

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“…With the exception of the details for the new preparation, the experiments followed procedures developed earlier [5,10,11,12]. Briefly, the experiments consisted of monitoring with fluorescence microscopy the expression of enhanced GFP (EGFP) plasmids and the nuclear translocation of various fluorescent probes (e.g.…”

Section: Methodsmentioning

confidence: 99%

Nuclear pore ion channel activity in live syncytial nuclei

Bustamante

2002

Pflügers Arch - Eur J Physiol

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Nuclear pore complexes (NPCs) are important nanochannels for the control of gene activity and expression. Most of our knowledge of NPC function has been derived from isolated nuclei and permeabilized cells in cell lysates/extracts. Since recent patch-clamp work has challenged the dogma that NPCs are freely permeable to small particles, a preparation of isolated living nuclei in their native liquid environment was sought and found: the syncytial nuclei in the water of the coconut Cocos nucifera. These nuclei have all properties of NPC-mediated macromolecular transport (MMT) and express foreign green fluorescent protein (GFP) plasmids. They display chromatin movement, are created by particle aggregation or by division, can grow by throwing filaments to catch material, etc. This study shows, for the first time, that living NPCs engaged in MMT do not transport physiological ions - a phenomenon that explains observations of nucleocytoplasmic ion gradients. Since coconuts are inexpensive (less than US$1/nut per litre), this robust preparation may contribute to our understanding of NPCs and cell nucleus and to the development of biotechnologies for the production of DNA, RNA and proteins.

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Calcium, ATP and nuclear pore channel gating

Cited by 33 publications

References 47 publications

Modulation of [Ca2+] Signaling Dynamics and Metabolism by Perinuclear Mitochondria in Mouse Parotid Acinar Cells

Modulation of [Ca2+] Signaling Dynamics and Metabolism by Perinuclear Mitochondria in Mouse Parotid Acinar Cells

Aldosterone signaling pathway across the nuclear envelope

Nuclear pore ion channel activity in live syncytial nuclei

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