共查询到20条相似文献,搜索用时 31 毫秒
1.
《Gene》1998,221(1):11-16
Both the rpb9 gene and its cDNA encoding the subunit 9 of RNA polymerase II were cloned from the fission yeast Schizosaccharomyces pombe. From the DNA sequences, Rpb9 was predicted to consist of 113 amino acid residues with a molecular mass of 13 175. S. pombe Rpb9 is 47, 40 and 36% identical in amino acid sequence to the corresponding subunits from Saccharomyces cerevisiae, human and Drosophila melanogaster, respectively. Previously, we failed to detect Rpb9 in the purified RNA polymerase II by amino-terminal micro-sequencing of proteolytic fragments of subunits separated by SDS-gel electrophoresis. After Western blot analysis using antibodies raised against the protein product of the newly isolated rpb9 gene, we found that the purified RNA polymerase II contains Rpb9. 相似文献
2.
Hiromi Maekawa Tomoko Nakagawa Yoko Uno Kenji Kitamura Chikashi Shimoda 《Molecular & general genetics : MGG》1994,244(5):456-464
When the fission yeastSchizosaccharomyces pombe is starved for nitrogen, the cells are arrested in the G1 phase, enter the G0 phase and initiate sexual development. Theste13 mutant, however, fails to undergo a G1 arrest when starved for nitrogen and since this mutant phenotype is not suppressed by a mutation in adenylyl cyclase (cyr1), it would appear thatste13
+ either acts independently of the decrease in the cellular cAMP level induced by starvation for nitrogen, or functions downstream of this controlling event. We have used functional complementation to clone theste13
+ gene from anS. pombe genomic library and show that its disruption is not lethal, indicating that, while the gene is required for sexual development, it is not essential for cell growth. Nucleotide sequencing predicts thatste13
+ should encode a protein of 485 amino acids in which the consensus motifs of ATP-dependent RNA helicases of the DEAD box family are completely conserved. Point mutations introduced into these consensus motifs abolished theste13
+ functions. The predicted Ste13 protein is 72% identical to theDrosophila melanogaster Me31B protein over a stretch of 391 amino acids. ME31B is a developmentally regulated gene that is expressed preferentially in the female germline and may be required for oogenesis. Expression of ME31B cDNA inS. pombe suppresses theste13 mutation. These two evolutionarily conserved genes encoding putative RNA helicases may play a pivotal role in sexual development. 相似文献
3.
Georgina Rodríguez de Lores Arnaiz 《Neurochemical research》1993,18(6):655-661
In previous papers, the isolation of brain soluble fractions able to modify neuronal Na+, K+-ATPase activity has been described. One of those fractions-peak I-stimulates membrane Na+, K+-ATPase while another-peak II-inhibits this enzyme activity, and has other ouabain-like properties. In the present study, synaptosomal membrane Na+, K+-ATPase was analyzed under several experimental conditions, using ATP orp-nitrophenylphosphate (p-NPP) as substrate, in the absence and presence of cerebral cortex peak II. Peak II inhibited K+-p-NPPase activity in a concentration dependent manner. Double reciprocal plots indicated that peak II uncompetitively inhibits K+-p-NPPase activity regarding substrate, Mg2+ and K+ concentration. Peak II failed to block the known K+-p-NPPase stimulation caused by ATP plus Na+. At various K+ concentrations, percentage K+-p-NPPase inhibition by peak II was similar regardless of the ATP plus Na+ presence, indicating lack of correlation with enzyme phosphorylation. Na+, K+-ATPase activity was decreased by peak II depending on K+ concentration. It is postulated that the inhibitory factor(s) present in peak II interfere(s) with enzyme activation by K+. 相似文献
4.
Yasuhiro Anraku Naoyuki Umemoto Ryogo Hirata Yoshikazu Ohya 《Journal of bioenergetics and biomembranes》1992,24(4):395-405
The yeast vacuolar proton-translocating ATPase is a member of the third class of H+-pumping ATPase. A family of this type of H+-ATPase is now known to be ubiquitously distributed in eukaryotic vacuo-lysosomal organelles and archaebacteria. NineVMA genes that are indispensable for expression of the enzyme activity have been cloned and characterized in the yeastSaccharomyces cerevisiae. This review summarizes currently available information on theVMA genes and cell biological functions of theVMA gene products. 相似文献
5.
The Na+/H+ exchangers (NHEs) catalyze the transport of Na+ in exchange for H+ across membranes in organisms and are required for numerous physiological processes. Here we report the cloning and characterization
of a novel human NHEDC1 (Na+/H+ exchanger like domain containing 1) gene, which was mapped to human chromosome 4p24. This cDNA is 1859 bp in length, encoding
a putative protein of 515 amino acids. The NHEDC1 proteins are highly conserved in mammals including human, mouse, rat, and
Macaca fascicularis. One remarkable characteristic of human NHEDC1 gene is that it is exclusively expressed in the testis by RT-PCR analysis.
Western blot analysis showed that the molecular weight of NHEDC1 is about 56 KDa.
Guangming Ye and Cong Chen contributed equally to this work. 相似文献
6.
7.
The role of putative Na+/H+ antiporters encoded by nhaS1 (slr1727), nhaS3 (sll0689), nhaS4 (slr1595), and nhaS5 (slr0415) in salt stress response and internal pH regulation of the cyanobacterium Synechocystis PCC 6803 was investigated. For this purpose the mutants (single, double, and triple) impaired in genes coding for Na+/H+ antiporters were constructed using the method of interposon mutagenesis. PCR analyses of DNA demonstrated that mutations in nhaS1, nhaS4, and nhaS5 genes were segregated completely and the mutants contained only inactivated copies of the corresponding genes. Na+/H+ antiporter encoded by nhaS3 was essential for viability of Synechocystis since no completely segregated mutants were obtained. The steady-state intracellular sodium concentration and Na+/H+ antiporter activities were found to be the same in the wild type and all mutants. No differences were found in the growth rates of wild type and mutants during their cultivation in liquid media supplemented with 0.68 M or 0.85 M NaCl as well as in media buffered at pH 7.0, 8.0, or 9.0. The expression of genes coding for Na+/H+ antiporters was studied. No induction of any Na+/H+ antiporter encoding gene expression was found in wild type or single mutant cells grown under high salt or at different pH values. Nevertheless, in cells of double and triple mutants adapted to high salt or alkaline pH some of the remaining Na+/H+ antiporter encoding genes showed induction. These results might indicate that some of Na+/H+ antiporters can functionally replace each other under stress conditions in Synechocystis cells lacking the activity of more than one antiporter. 相似文献
8.
Georgina Rodríguez de Lores Arnaiz 《Molecular neurobiology》1992,6(4):359-375
The arrival of the nerve impulse to the nerve endings leads to a series of events involving the entry of sodium and the exit
of potassium. Restoration of ionic equilibria of sodium and potassium through the membrane is carried out by the sodium/potassium
pump, that is the enzyme Na+,K+-ATPase. This is a particle-bound enzyme that concentrates in the nerve ending or synaptosomal membranes. The activity of
Na+,K+-ATPase is essential for the maintenance of numerous reactions, as demonstrated in the isolated synaptosomes. This lends interest
to the knowledge of the possible regulatory mechanisms of Na+,K+-ATPase activity in the synaptic region. The aim of this review is to summarize the results obtained in the author's laboratory,
that refer to the effect of neurotransmitters and endogenous substances on Na+,K+-ATPase activity. Mention is also made of results in the field obtained in other laboratories.
Evidence showing that brain Na+,K+-ATPase activity may be modified by certain neurotransmitters and insulin have been presented. The type of change produced
by noradrenaline, dopamine, and serotonin on synaptosomal membrane Na+,K+-ATPase was found to depend on the presence or absence of a soluble brain fraction. The soluble brain fraction itself was
able to stimulate or inhibit the enzyme, an effect that was dependent in turn on the time elapsed between preparation and
use of the fraction.
The filtration of soluble brain fraction through Sephadex G-50 allowed the separation of two active subfractions: peaks I
and II. Peak I increased Na+,K+- and Mg2+-ATPases, and peak II inhibited Na+,K+-ATPase. Other membrane enzymes such as acetylcholinesterase and 5′-nucleotidase were unchanged by peaks I or II.
In normotensive anesthetized rats, water and sodium excretion were not modified by peak I but were increased by peak II, thus
resembling ouabain effects.3H-ouabain binding was unchanged by peak I but decreased by peak II in some areas of the CNS assayed by quantitative autoradiography
and in synaptosomal membranes assayed by a filtration technique. The effects of peak I and II on Na+,K+-ATPase were reversed by catecholamines. The extent of Na+,K+-ATPase inhibition by peak II was dependent on K+ concentration, thus suggesting an interference with the K+ site of the enzyme. Peak II was able to induce the release of neurotransmitter stored in the synaptic vesicles in a way similar
to ouabain. Taking into account that peak II inhibits only Na+,K+-ATPase, increases diuresis and natriuresis, blocks high affinity3H-ouabain binding, and induces neurotransmitter release, it is suggested that it contains an ouabain-like substance. 相似文献
9.
10.
We have already described the separation of two brain soluble fractions by Sephadex G-50, one of which stimulates (peak I) and the other inhibits (peak II) Na+, K+-ATPase and K+-p-nitrophenylphosphatase (K+-p-NPPase) activities. Here we examine the features of synaptosomal membrane p-NPPase activity in the presence and absence of brain peak I. It was observed that stimulation of Mg2+, K+-p-NPPase activity by peak I was concentration dependent, The ability of peak I to stimulate p-NPPase activity was lost by heat treatment followed by brief centrifugation. Pure serum albumin also stimulated enzyme activity. K+-p-NPPase stimulation by peak I proved dependent on K+ concentration but independent of Mg2+ and substrate p-nitrophenylphosphate concentrations. Since our determinations were performed in a non-phosphorylating condition reflecting the Na+, K+-ATPase Na+ site, it is suggested that peak I may stimulate the Na+-dependent enzyme phosphorylation known to take place from the internal cytoplasmic side. 相似文献
11.
Isolation and characterization of a Na+/H+ antiporter gene from the halophyte Atriplex gmelini 总被引:25,自引:0,他引:25
Hamada Akira Shono Mariko Xia Tao Ohta Masaru Hayashi Yasuyuki Tanaka Akira Hayakawa Takahiko 《Plant molecular biology》2001,46(1):35-42
With a homologous gene region we successfully isolated a Na+/H+ antiporter gene from a halophytic plant, Atriplex gmelini, and named it AgNHX1. The isolated cDNA is 2607 bp in length and contains one open reading frame, which comprises 555 amino acid residues with a predicted molecular mass of 61.9 kDa. The amino acid sequence of the AgNHX1 gene showed more than 75% identity with those of the previously isolated NHX1 genes from glycophytes, Arabidopsis thaliana and Oryza sativa. The migration pattern of AgNHX1 was shown to correlate with H+-pyrophosphatase and not with P-type H+-ATPase, suggesting the localization of AgNHX1 in a vacuolar membrane. Induction of the AgNHX1 gene was observed by salt stress at both mRNA and protein levels. The expression of the AgNHX1 gene in the yeast mutant, which lacks the vacuolar-type Na+/H+ antiporter gene (NHX1) and has poor viability under the high-salt conditions, showed partial complementation of the NHX1 functions. These results suggest the important role of the AgNHX1 products for salt tolerance. 相似文献
12.
The effect of the putative K+/H+ ionophore, nigericin on the internal Na+ concentration ([Na
i
]), the internal pH (pH
i
), the internal Ca2+ concentration ([Ca
i
]) and the baseline release of the neurotransmitter, GABA was investigated in Na+-binding benzofuran isophtalate acetoxymethyl ester (SBFIAM), 2′,7′-bis(carboxyethyl)-5(6) carboxyfluorescein acetoxymethyl
ester (BCECF-AM), fura-2 and [3H]GABA loaded synaptosomes, respectively. In the presence of Na+ at a physiological concentration (147 mM), nigericin (0.5 μM) elevates [Na
i
] from 20 to 50 mM, increases thepH
i
, 0.16 pH units, elevates four fold the [Ca
i
] at expense of external Ca2+ and markedly increases (more than five fold) the release of [3H]GABA. In the absence of a Na+ concentration gradient (i.e. when the external Na+ concentration equals the [Na
i
]), the same concentration (0.5 μM) of nigericin causes the opposite effect on thepH
i
(acidifies the synaptosomal interior), does not modify the [Na
i
] and is practically unable to elevate the [Ca
i
] or to increase [3H]GABA release. Only with higher concentrations of nigericin than 0.5 μM the ionophore is able to elevate the [Ca
i
] and to increase the release of [3H]GABA under the conditions in which the net Na+ movements are eliminated. These results clearly show that under physiological conditions (147 mM external Na+) nigericin behaves as a Na+/H+ ionophore, and all its effects are triggered by the entrance of Na+ in exchange for H+ through the ionophore itself. Nigericin behaves as a K+/H+ ionophore in synaptosomes just when the net Na+ movements are eliminated (i.e. under conditions in which the external and the internal Na+ concentrations are equal). In summary care must be taken when using the putative K+/H+ ionophore nigericin as an experimental tool in synaptosomes, as under standard conditions (i.e. in the presence of high external
Na+) nigericin behaves as a Na+/H+ ionophore. 相似文献
13.
Georgina Rodríguez de Lores Arnaiz 《Neurochemical research》1990,15(3):289-294
Previous evidence from this laboratory indicated that catecholamines and brain endogenous factors modulate Na+, K+-ATPase activity of the synaptosomal membranes. The filtration of a brain total soluble fraction through Sephadex G-50 permitted the separation of two fractions-peaks I and II-which stimulated and inhibited Na+, K+-ATPase, respectively (Rodríguez de Lores Arnaiz and Antonelli de Gomez de Lima, Neurochem. Res.11, 1986, 933). In order to study tissue specificity a rat kidney total soluble was fractionated in Sephadex G-50 and kidney peak I and II fractions were separated; as control, a total soluble fraction prepared from rat cerebral cortex was also processed. The UV absorbance profile of the kidney total soluble showed two zones and was similar to the profile of the brain total soluble. Synaptosomal membranes Na+, K+- and Mg2+-ATPases were stimulated 60–100% in the presence of kidney and cerebral cortex peak I; Na+, K+-ATPase was inhibited 35–65% by kidney peak II and 60–80% by brain peak II. Mg2+-ATPase activity was not modified by peak II fractions. ATPases activity of a kidney crude microsomal fraction was not modified by kidney peak I or brain peak II, and was slightly increased by kidney peak II or brain peak I. Kidney purified Na+, K+-ATPase was increased 16–20% by brain peak I and II fractions. These findings indicate that modulatory factors of ATPase activity are not exclusive to the brain. On the contrary, there might be tissue specificity with respect to the enzyme source. 相似文献
14.
Characterization of K+-dependent and K+-independentp-nitrophenylphosphatase activity of synaptosomes
M. Guerra Marichal A. Rodríguez Del Castillo P. Martín Vasallo E. Battaner Arias 《Neurochemical research》1993,18(7):751-758
These experiments examined effects of several ligands on the K+
p-nitrophenylphosphatase activity of the (Na+,K+)-ATPase in membranes of a rat brain cortex synaptosomal preparation. K+-independent hydrolysis of this substrate by the synaptosomal preparation was studied in parallel; the rate of hydrolysis in the absence of K+ was approximately 75% less than that observed when K+ was included in the incubation medium. The response to the H+ concentrations was different: K+-independent activity showed a pH optimum around 6.5–7.0, while the K+-dependent activity was relatively low at this pH range. Ouabain (0.1 mM) inhibited K+-dependent activity 50%; a concentration 10 times higher did not produce any appreciable effect on the K+-independent activity. Na+ did not affect K+-independent activity at all, while the same ligand concentration inhibited sharply the K+-dependent activity; this inhibition was not competitive with the substrate,p-nitrophenyl phosphate. K+-dependent activity was stimulated by Mg2+ with low affinity (millimolar range), and 3 mM Mg2+ produced a slight stimulation of the activity in absence of K+, which could be interpreted as Mg2+ occupying the K+ sites. Ca2+ had no appreciable effect on the activity in the absence of K+. However, in the presence of K+ a sharp inhibition was found with all Ca2+ concentrations studied. ATP (0.5 mM) did not affect the K+-independent activity, but this nucleotide behaved as a competitive inhibitor top-nitrophenylphosphate. Pi inhibited activity in the presence of K+, competively to the substrate, so it could be considered as the second product of the reaction sequence.Abbreviations used
p-NPP
p-nitrophenylphosphate
-
p-NPPase
rho-nitrophenylphosphatase activity 相似文献
15.
Horst Helbig Christoph Korbmacher Michael Wiederholt 《The Journal of membrane biology》1987,99(3):173-186
Summary Using intracellular microelectrode technique, we investigated the changes in membrane voltage (V) of cultured bovine pigmented ciliary epithelial cells induced by different extracellular solutions. (1)V in 213 cells under steady-state conditions averaged –46.1±0.6 mV (sem). (2) Increasing extracellular K+ concentration ([K+]
o
) depolarizedV. Addition of Ba2+ could diminish this response. (3) Depolarization on doubling [K+]
o
was increased at higher [K+]
o
(or low voltage). (4) Removing extracellular Ca2+ decreasedV and reduced theV amplitude on increasing [K+]
o
. (5)V was pH sensitive. Extra-and intracellular acidification depolarizedV; alkalinization induced a hyperpolarization.V responses to high [K+]
o
were reduced at acidic extracellular pH. (6) Removing K
o
+
depolarized, K
o
+
readdition after K+ depletion transiently hyperpolarizedV. These responses were insensitive to Ba2+ but were abolished in the presence of ouabain or in Na+-free medium. (7) Na+ readdition after Na+ depletion transiently hyperpolarizedV. This reaction was markedly reduced in the presence of ouabain or in K+-free solution but unchanged by Ba2+. It is concluded that in cultured bovine pigmented ciliary epithelial cells K+ conductance depends on Ca2+, pH and [K+]
o
(or voltage). An electrogenic Na+/K+-transport is present, which is stimulated during recovery from K+ or Na+ depletion. This transport is inhibited by ouabain and in K+-or Na+-free medium. 相似文献
16.
Steven P. Weinstein Adele B. Kostellow David H. Ziegler Gene A. Morrill 《The Journal of membrane biology》1982,69(1):41-48
Summary Progesterone initiates the resumption of the meiotic divisions in the amphibian oocyte. Depolarization of theRana pipiens oocyte plasma membrane begins 6–10 hr after exposure to progesterone (1–2 hr before nuclear breakdown). The oocyte cytoplasm becomes essentially isopotential with the medium by the end of the first meiotic division (20–22 hr). Voltage-clamp studies indicate that the depolarization coincides with the disappearance of an electrogenic Na+, K+-pump, and other electrophysiological studies indicate a decrease in both K+ and Cl– conductances of the oocyte plasma membrane. Measurement of [3H]-ouabain binding to the plasma-vitelline membrane complex indicates that there are high-affinity (K
d-4.2×10–8
m), K+-sensitive ouabain-binding sites on the unstimulated (prophase-arrest) oocyte and that ouabain binding virtually disappears during membrane depolarization. [3H]-Leucine incorporation into the plasma-vitelline membrane complex increased ninefold during depolarization with no significant change in uptake or incorporation into cytoplasmic proteins or acid soluble pool(s). This together with previous findings suggests that progesterone acts at a translational level to produce a cytoplasmic factor(s) that down-regulates the membrane Na+, K+-ATPase and alters the ion permeability and transport properties of both nuclear and plasma membranes. 相似文献
17.
Wenhui Wang Robert M. Henderson John Geibel Stanley White Gerhard Giebisch 《The Journal of membrane biology》1989,112(3):277-289
Summary Isolated early distal tubule cells (EDC) of frog kidney were incubated for 20–28 hr in the presence of aldosterone and then whole-cell K+ currents were measured at constant intracellular pH by the whole-cell voltage-clamp technique. Aldosterone increased barium-inhibitable whole-cell K+ conductance (gK+) threefold. This effect was reduced by amiloride and totally abolished by ouabain. However, aldosterone could still raisegK+ in ouabain-treated cells in the presence of furosemide.We tested whether changes in intracellular pH (pH
i
) could be a signal for cells to regulategK+. After removal of aldosterone, the increase ingK+ was preserved by subsequent incubation for 8 hr at pH 7.6 but abolished at pH 6.6. In the complete absence of aldosterone, incubation of cells at pH 8.0 for 20–28 hr raised pH
i
and doubledgK+.Using the patch-clamp technique, three types of K+-selective channels were identified, which had conductances of 24, 45 and 59 pS.Aldosterone had no effect on the conductance or open probability (P
o) of any of the three types of channels. However, the incidence of observing type II channels was increased from 4 to 22%. Type II channels were also found to be pH sensitive,P
o was increased by raising pH.These results indicate that prolonged aldosterone treatment raises pH
i
and increasesgK+ by promoting insertion of K+ channels into the cell membrane. Channel insertion is itself triggered by raising both pH
i
and increasing the activity of the Na+/K+ pump in early distal cells of frog kidney.
Present address: Department of Physiology, The University of Leeds, Leeds, LS2 9NQ, England 相似文献
18.
19.
Functional analysis of amino acids of the Na+/H+ exchanger that are important for proton translocation 总被引:3,自引:0,他引:3
Katiyar S Thelma BK Murthy NS Hedau S Jain N Gopalkrishna V Husain SA Das BC 《Molecular and cellular biochemistry》2003,254(1-2):117-124
The Na+/H+ exchanger is an integral membrane protein found in the plasma membrane of eukaryotic and prokaryotic cells. In eukaryotes it functions to exchange one proton for a sodium ion. In mammals it removes intracellular protons while in plants and fungal cells the plasma membrane form removes intracellular sodium in exchange for extracellular protons. In this study we used the Na+/H+ exchanger of Schizosaccharomyces pombe (Sod2) as a model system to study amino acids critical for activity of the protein. Twelve mutant forms of the Na+/H+ exchanger were examined for their ability to translocate protons as assessed by a cytosensor microphysiometer. Mutation of the amino acid Histidine 367 resulted in defective proton translocation. The acidic residues Asp145, Asp178, Asp266 and Asp267 were important in the proton translocation activity of the Na+/H+ exchanger. Mutation of amino acids His98, His233 and Asp241 did not significantly impair proton translocation by the Na+/H+ exchanger. These results confirm that polar amino acids are important in proton flux activity of Na+/H+ exchangers. 相似文献
20.
Summary To study the physiological role of the bidirectionally operating, furosemide-sensitive Na+/K+ transport system of human erythrocytes, the effect of furosemide on red cell cation and hemoglobin content was determined in cells incubated for 24 hr with ouabain in 145mm NaCl media containing 0 to 10mm K+ or Rb+. In pure Na+ media, furosemide accelerated cell Na+ gain and retarded cellular K+ loss. External K+ (5mm) had an effect similar to furosemide and markedly reduced the action of the drug on cellular cation content. External Rb+ accelerated the Na+ gain like K+, but did not affect the K+ retention induced by furosemide. The data are interpreted to indicate that the furosemide-sensitive Na+/K+ transport system of human erythrocytes mediates an equimolar extrusion of Na+ and K+ in Na+ media (Na+/K+ cotransport), a 1:1 K+/K+ (K+/Rb+) and Na+/Na+ exchange progressively appearing upon increasing external K+ (Rb+) concentrations to 5mm. The effect of furosemide (or external K+/Rb+) on cation contents was associated with a prevention of the cell shrinkage seen in pure Na+ media, or with a cell swelling, indicating that the furosemide-sensitive Na+/K+ transport system is involved in the control of cell volume of human erythrocytes. The action of furosemide on cellular volume and cation content tended to disappear at 5mm external K+ or Rb+. Thein vivo red cell K+ content was negatively correlated to the rate of furosemide-sensitive K+ (Rb+) uptake, and a positive correlation was seen between mean cellular hemoglobin content and furosemide-sensitive transport activity. The transport system possibly functions as a K+ and waterextruding mechanism under physiological conditiosin vivo. The red cell Na+ content showed no correlation to the activity of the furosemide-sensitive transport system. 相似文献