Molecular characterization of a K+ channel blocker in the scorpionButhus martensii Karsch

K⁺ channel blockers of scorpion venoms are of important value in studying pharmacology and physiology of specific K⁺ channel of cells. Based on the amino acid sequences of BmP01 previously characterized as a small-conductance Ca²⁺-activated K⁺ channel blocker, two “back to back” degenarate primers have been designed and synthesized for inverse PCR strategy, its full-length cDNA has been cloned from the venom gland of the Chinese scorpionButhus martensii. The cDNA is composed of 3 parts: 5′ UTR, ORF and 3′ UTR. The flanking sequence of translation initiation codon ATG is AAAATGA, which is highly conserved in scorpion Na⁺ channel toxin and protozoan genes, suggesting that these genes may have followed a common mechanism for translation initiation. The 3′ UTR contains poly(A) signal AATAAA. The open reading frame encodes a precursor of 57 residues with a signal peptide of 28 residues and a mature peptide of 29 residues. The signal peptide is rich in hydrophobic amino acid residues and its length is significantly different from that of the determined scorpion Na⁺ channel toxin. The deduced amino acid sequence of mature peptide is completely consistent with BmP01 previously determined by primary structure analysis.     

Reduced Na+ and K+ permeability of K+ channel in plasma membrane isolated from roots of salt-tolerant mutant of wheat

The Na⁺ and K⁺ permeability of K⁺ channel in plasma membrane, isolated from roots of the salt-tolerant mutant of wheat, was lower than that of wild type in 100 mmol/L KCl and NaCl solution. The opening frequency of K⁺ channel of the mutant reduced more significantly than that of wild type in two kinds of solution mentioned above. It is assumed that the reduction of opening frequency mainly contributes to the Na⁺ and K⁺ permeability of K⁺ channel of the mutant. The electric conductance of single-channel of the mutant was similar to that of wild type and the main difference between them was exhibited as the opening frequency. Their K⁺/Ka⁺ selectivity of K⁺ channel had no significant difference. The K⁺/Na⁺ selectivity of the mutant and wild type was 3.35 and 3.18 respectively.     

Effect of fatty acids on polyamine contents and Na+/H+ antiport activity in PM vesicles isolated from roots of barley under salt stress

With 200 mmol/L NaCl treatment on barley cultivar “Jian 4” (Hordeum vulgare L. cv. J4) seedlings for 6 d, the contents of covalently and noncovalently conjugated polyamines (PAs) and activities of H⁺-ATPase in plasma membrane (PM) vesicles isolated from the roots decreased remarkably. Moreover, the activity of Na⁺/H⁺ antiport was detected first in PM vesicles. The results showed that the decrease in the contents of membrane phospholipid, noncovalently conjugated PAs and activity of H⁺-ATPase caused by NaCl could be restored partially by application of 1 mmol/L stearie acid (C_16:0) and linoleic acid (C_18:2), and C_18:2 was more effective than C_16:0 In addition, a reduction in the contents of covalently conjugated PAs was only reversed partially in the presence of C_18:2 Furthermore, Na⁺/H⁺ antiport activity was strengthened by exogenous C_16:0 and C_18:2 and C_18:2 was more effective than C_16:0. The correlative analysis suggested that, after application of C_16:0 and C_18:2 under salt stress, there was a significant positive correlation existing among phospholipid content, noncovalently conjugated PA levels, H⁺-ATPase activities and Na⁺/H⁺ antiport activities, indicating that one of the mitigative mechanisms of exogenous fatty acids on salt injury was to improve membrane phospholipid and PA contents, leading to an enhance in membrane integrity and a change in charge status of PM vesicles, so the activity of membrane-associated enzyme H⁺-ATPase was increased and synthesis of Na⁺/H⁺ antiport protein was activated.     

The short C-terminal hydrophilic domain of NhaH Na^＋/H^＋ antiporter from Halobacillus dabanensis with roles in resistance to salt and in pH sensing

The Na^＋/H^＋ antiporter plays key roles in maintaining low cytoplasmic NaNa^＋ level and pH homeostasis, while little is known about the Carboxyl-terminal hydrophilic tails of prokaryotic antiporters. In our previous study, the first Na^＋/H^＋ antiporter gene nhaH from moderate halophiles was cloned from Halobacillus dabanensis D-8 by functional complementation. A topological model suggested that only nine amino acid residues （^395PLIKKLGMI403） existed in the hydrophilic C-terminal domain of NhaH. The C-terminal truncated mutant of NhaH was constructed by PCR strategy and designated as nhaH△C. Salt tolerance experiment demonstrated that the deletion of hydrophilic C-terminal nine amino acid residues significantly inhibited the complementation ability of E. coil KNabc, in which three main Na^＋/H^＋ antiporters nhaA, nhaB and chaA were deleted. Everted membrane vesicles prepared from E. coil KNabc/nhaHAC decreased both Na^＋/H^＋ and Li^＋/H^＋ exchange activities of NhaH, and also resulted in an acidic shift of its pH profile for Na^＋, indicating a critical role of the short C-terminal domain of NhaH antiporter in alkali cation binding/translocation and pH sensing.     

IS4 peptide forms ion channel in rat skeletal muscle cell membrane

A 22-mer peptide, identical to the primary sequence of domain I segment 4 (IS4) of rat brain sodium channel I, has been synthesized. IS4 peptide can incorporate into cultured rat skeletal myotube membranes and form ion channels. With patch clamp cell-attached technique single channel currents through IS4 channels can be recorded. The single channel conductances of IS4 channels are distributed heterogeneously. With different holding potentials, the mean open time, the mean closed time and the mean open probability are different respectively. IS4 channels are selective for Na⁺, Li⁺ and K⁺, but not for Cl⁻.     

Impact of gamma radiation and salinity on growth and K+/Na+ balance in two populations of Medicago sativa (L.) cultivar Gabès

The perennial Medicago sativa cv. Gabès is widely grown on saline soils in Tunisian oases. Morphological and physiological analyses of two populations (Mareth and Gannouch) were conducted in order to study the effect of salinity and gamma radiation (350 Gy) interaction on two populations of this species. It has been shown that the two irradiated populations of Medicago sativa are fairly tolerant to salt at growth phase compared to the non-irradiated. Exposure to gamma irradiation (350 Gy), alone or in combination with salt stress, increased significantly (p<0.001) shoot number, stem height and chlorophyll b pigment especially for the Gannouch population, while no change occurred for the Mareth population. The presence of salt to 9g/l affected significantly the root biomass and induced a reduction of shoot development of both control and irradiated alfalfa populations. For all treatments, plants of two populations remained able to produce and to allocate dry matter to the different organs. The survey of Na⁺ / K⁺ ratio showed that the growth of the aerial organs of two non-irradiated populations was at least determined by a selectivity in favour of the K+ ions (r² = 0.97 and r² = 0.59 for Mareth and Gannouch irradiated populations, respectively). However, the rather weak correlation detected for the irradiated populations, particularly for the irradiated Gannouch, seems to be the consequence of the effect of irradiation that improved potassium availability, essential element for growth and development. Results also showed that the two irradiated populations, especially the Mareth, accumulated Na⁺ ions in its photosynthetic organs. This accumulation was associated with an improvement of foliar water content at a level of salinity arrounding 5g/l. Such a mechanism reflects probably an inclusive behaviour of the plants and a good aptitude to use the dominant ions (Na⁺) for the osmotic adjustment. However, the non-irradiated populations are unable to adjust their internal osmotic potential. Consequently, irradiated plants have probably adapted to the osmotic stress by either closing their stomata or increasing the osmotic pressure of the leaf cells.     

H-ATPase in the plasma membrane of Arabidopsis pollen cells is involved in extracellular calmodulin-promoted pollen germination

In this paper, the role of the plasma membrane (PM) H⁺-ATPase in extracellular calmodulin (CaM)-promoted pollen germination and in tube growth of Arabidopsis thaliana was investigated. Pollen germination, pollen tube growth rate, free cytosolic Ca²⁺ concentration ([Ca²⁺]_cyt) and Ca²⁺ channel activity in the PM of pollen cells were measured. In response to fusicoccin or CaM treatment, in vitro pollen germination and pollen tube growth rate, [Ca²⁺]_cyt and activity of a hyperpolarization-activated Ca²⁺-permeable channel increased. Sodium vanadate inhibited the promotion of these parameters by extracellular CaM. The results suggest that H⁺-ATPase may be involved in extracellular CaM-regulated pollen germination and in tube growth by modulation of the hyperpolarization-activated Ca²⁺ channel in the PM of A. thaliana pollen cells.     

Na+-K+-ATP酶的π-演算语义及验证

给出了Na⁺-K⁺-ATP酶跨越细胞膜同时主动向胞内运转钾离子和向胞外运转钠离子这一生化过程的π-演算模型及该模型的Spin验证. 证明了用过程代数的方法表示以“相互通讯”和“可移动”为主要特征的生物系统并模拟其行为的可行性.      

Studies on the overexpression of the soybean GmNHX1 in Lotus corniculatus： The reduced Na^＋ level is the basis of the increased salt tolerance

Soil salinity is one of the major factors reducing plant growth and productivity. The detrimental effects of salt on plants are a consequence of both a water deficit resulting in osmotic stress and the excess so- dium ions on critical biochemical processe…     

The seasonal variations of δ~(18)O , Cl~- , Na~+ , NO_3~- and Ca~(2+) in the snow and firn recovered from Princess Elizabeth Land, Antarctica

Snow and firn samples recovered from two snow pits (2.5 and 4.5m deep) and one 50-m firn core along the route of the 1996/1997 Chinese First Antarctic Inland Traverse Expedition in Princess Elizabeth Land, East Antarctica, have been measured for chemical composition and oxygen isotope ratio. In the two snow pits, the variations of NO3- are partly in phase with that of δ18O, while the variations of Cl~ and Na+ are in inverse phase with that of δ18O. The variations of CI- , Na+ , NO3- and δ18O show obvious seasonal variations and annual stratag-raphy. However, with the depth increasing, the seasonal variations of δ18O are gradually smoothed below 3 m (corresponding to about 10-year mass accumulation) in depth while the seasonal variations of Cl- , Na+ and NO3- are kept fairly well in the whole profile of the 50-m firn core (corresponding to about 250-year mass accumulation). The results provide a useful tool for dating the snow stratum in this region. On the contrary, no obvious seasonal variations of Ca2 + are found in the profiles.     

Sodium current of neurosecretory cells in the eyestalk from the shrimp Penaeus japonicus

The properties of the inward current of medulla terminalis-X-organ (MTXO) cells isolated from the Penaeus japonicus eyestalk were studied with the whole-cell clamp technique in the presence of Ca²⁺ and K⁺ channel blockers. The inward currents had a threshold at about −50 mV and peaked at −10 mV. The reversed potential (V_rev) was very close to V_Na, the theoretical Nernst equilibrium potential for Na⁺. V_rev followed V_Na when the external Na⁺ concentration was varied and the currents were entirely suppressed by 30 nM tetrodotoxin (TTX), indicating that it was carried by Na⁺. The smooth line of concentration-dependent inhibition of sodium currents by TTX represented the best fit with the Hill equation, yielding an IC₅₀ of 2.1 ± 0.1 nM. The values of the half-maximal activation voltage V_h were −20.6 ± 0.5 and −19.3 ± 0.5 mV, respectively, in the absence and presence of 2 nM TTX. TTX had no significant effect on the voltage dependence of steady-state activation and inactivation of I_Na. Taken together, the results suggest that the inward current recorded under our experimental conditions was carried by sodium ions flowing through fast voltage-dependent Na⁺ channels.     

Circular dichroism spectroscopic studies on structures formed by telomeric DNA sequencesin vitro

Telomere plays an important role in cellular processes, such as cell aging, death and carcinogenisis. Having special sequences, it can form quadruplex structurein vitro. Circular dichroism (CD) spectroscopic studies show that TTAGGG, (TTAGGG)₂ and (TTAGGG)₄ can all form quadruplexin vitro and exist mainly as parallel quadruplex without metal ions. Both K⁺ and Na⁺ can stabilize the tetrameric structure and facilitate the forming of anti-parallel conformation. Furthermore, the conformations of quadruplex can also be affected by sequence length, the nature and concentration of metal ions.     

Effect of Cl- on photosynthetic bicarbonate uptake in two cyanobacteria Microcystis aeruginosa and Synechocystis PCC6803

Photosynthetic inorganic carbon utilization was investigated in two cyanobacteria Microcystis aeruginosa and Synechocystis PCC6803 grown in standing culture. Photosynthetic rates for the two algae reached about 10 times the theoretical CO₂ supply rate at low dissolved inorganic carbon (DIC) of 100 μmol/L, and the rates were unaffected by the addition of 20 mmol/L Na⁺, indicating that the two algae possessed Na+-independent HCO^-₃ utilization for photosynthesis under low DIC. Their photo- synthetic rates at low DIC were inhibited by higher Cl^- and the degrees of inhibition were increased with the rise of CI^- concentration, and in the presence of Diphenylamine-2-carboxylate (DPC), a reported Cl^- channel inhibitor, the rates decreased by 74%－82%, implying that putative DPC-sensitive Cl^- channels participate in Na+-independent HCO₃^- uptake for photosynthesis. The experiment of intracellular 14C-DIC accumulation for photosynthesis showed that internal DIC pools decreased by about 80% with 200 μmol/L DPC and by 64%－70% with 100 mmol/L Cl^-. The experiment of chlorophyll a fluorescence quenching showed that initial rates and extents of fluorescence quenching obviously decreased with 200 μmol/L DPC or 100 mmol/L Cl^-. The two experiments gave further evidence that putative DPC-sen- sitive Cl^- channels participate in Na+-independent HCO^-₃ uptake for photosynthesis in the two algae grown in standing culture.     

新型LED红色荧光材料Ca_xSr_(1-x-y-z)MoO_4:yEu~(3+);zNa~+的制备和表征

利用Na+为电荷补偿剂制备CaxSr1-x-y-zMoO4:yEu3+,zNa+红色LED荧光粉体.研究了电荷补偿剂的用量、基质材料中Ca2+的含量、合成温度、反应时间及发光中心Eu3+摩尔分数对荧光材料晶体结构和发光性能的影响.结果表明:最佳工艺条件为电荷补偿剂摩尔分数为6%,Eu3+摩尔分数为8%,Ca2+摩尔分数为60%,反应温度900℃和反应时间2 h.光谱测试结果表明,该荧光材料可被311,395或465 nm有效激发,发射峰在616 nm.     

黑鲷NKCC1分子特征及其对急性盐度胁迫的表达响应

为了解黑鲷(Acanthopagrus schlegelii)在盐度胁迫过程中的适应机制,本研究利用基因扩增、聚类分析、荧光定量PCR等技术对NKCC1基因进行生物信息学分析以及在不同组织中的表达特征研究,并探讨其在急性盐度胁迫下的表达机制。结果表明NKCC1基因的开放阅读框(Open Reading Frame, ORF)为3 504 bp,共编码1 167个氨基酸,预测蛋白分子量为127.19 kDa,理论等电点为5.75。结构域分析表明黑鲷NKCC1分子为跨膜蛋白,并包含一个典型的Na~+-K~+-Cl~-共同转运蛋白SLC12A结构域,且在不同物种中具有很高的保守性;序列比对分析表明黑鲷NKCC1氨基酸序列与金头鲷NKCC1a的同源性最高,为97.8%;组织表达分析表明黑鲷NKCC1基因在所检测的12个组织,即肝脏、肾脏、心、脑、眼、鳃、鳍条、脾脏、皮肤、肠、性腺和白肌中均有表达,其中在肾脏、鳃和肠中表达量最高,并显著高于其他组织(P0.05);在低盐度胁迫下,NKCC1基因在鳃组织中的表达量对盐度变化响应十分迅速,而在肾脏组织中,低盐度和高盐度胁迫都会在不同时间点抑制NKCC1基因的表达量。综上所述,本研究为解析NKCC1基因在黑鲷调节渗透压和离子平衡过程中的功能机制奠定了基础,并为促进黑鲷养殖产业的可持续发展提供一定的技术理论。     

Cesium accumulation by bacterium Thermus sp. TibetanG7： hints for biomineralization of cesiumbearing geyserite in hot sprinss in Tibet, China

The bacterium Thermus sp. TibetanG7, isolated from hot springs in Tibet, China, was examined for the ability to accumulate cesium from solutions. Environmental conditions were simulated and the effects of pH, K^＋, Na^＋and K^＋-regimes were then studied to determine the possible role of the bacterium in the formation of cesium-bearing geyserite around these hot springs. In despite of the inhibition of K^＋ and Na^＋, the bacterium Thermus sp. TibetanG7 revealed noticeable accumulation of cesium from solutions, with maximum accumulations of 53.49 and 40.41 pmol Cesium/g cell dry weight in Na^＋ and K^＋ inhibition experiments, respectively. The accumulation of cesium by this microorganism is rapid, with 40% --50% accumulated within the first 5 min. K^＋-deficient cells showed a much higher capacity of cesium accumulation compared with K^＋-sufficient cells. It is evident that the bacteria within the genus thermus play a significant role in the cesium assembly. The formation of cesium-bearing geyserite is also considered.     

Phenotypic analysis of the medullary-type CD4−CD8+ thymocytes in mice

Phenotypic analysis of the medullary-type CD4 CD8⁺ (CD8SP) thymocytes has revealed phenotypic heterogeneity within this cell population. The phenotype of mature peripheral CDS⁺T cells is TCRαβ⁺CD3⁺Qa-2⁺HSA 3G11⁻6C10, whereas in the medullary-type CD8SP thymocytes, 20% are Qa-2⁺; 33%, HAS; 30%, 3G11; and 70% are 6C10. The disparate expression patterns of these four cell surface markers suggest that medullary-type CD8SP thymocytes may undergo phenotypic maturation process. According to the distribution of these four cell surface markers, six subgroups of CD8SP thymocytes have been identified. The precursor-progeny relationship along with developmental pathway is postulated as follows: 6C10⁺HSA⁺3G11 Qa-2→ 6C10⁺HSA⁺ 3G11⁺Qa-2 → 6C10 HSA⁺3G11⁺Qa-2 → 6C10⁻HSA⁻3G11⁺Qa-2 → 6C10⁻HSA⁻3G11 Qa-2 → 6C10⁻HA S 3G11 Qa-2⁺, the cells in the last subgroup exit the thymus and home into periphery.     

Nitric oxide suppresses stomatal opening by inhibiting inward-rectifying K<Stack><Subscript>in</Subscript><Superscript>+</Superscript></Stack> channels in <Emphasis Type="Italic">Arabidopsis</Emphasis> guard cells

We explore nitric oxide （NO） effect on K^＋in, channels in Arabidopsis guard cells. We observed NO inhibited K^＋in, currents when Ca^2＋ chelator EGTA （Ethylene glycol-bis（2-aminoethylether）-N,N,N′,N;tetraacetic acid） was not added in the pipette solution; K^＋in currents were not sensitive to NO when cytosolic Ca^2＋ was chelated by EGTA. NO inhibited the Arabidopsis stomatal opening, but when EGTA was added in the bath solution, inhibition effect of NO on stomatal opening vanished. Thus, it implies that NO elevates cytosolic Ca^2＋ by activating plasma membrane Ca^2＋ channels firstly, then inactivates K^＋in, chartnels, resulting in stomatal opening suppressed subsequently.     

Mechanism for chronic pain generation

Neuropathic pain and the other abnormalities of sensation induced by axon injury or by peripheral nerve inflammation should result from functional compensations of the injured neurons during their regeneration. Ectopic distribution of proteins related to Na⁺, K⁺ and Ca²⁺ channels as well as of receptors on both membranes of injured axon and its cell body becomes a main pacemaker from which spontaneous ectopic afferent of primary sensatory neurons and crosstalk between neurons occur. Abnormal ectopic afferent activities lead to disorders of the sensation, such as hyperalgesia, allodynia, spontaneous pain and paraesthesia. Administration of some ion channel agents and/or α2-adrenergic blockers has shown efficiency in preventing neuropathic pain development and in relieving neuropathic pain.     

Effect of MTEC1 on the functional expression of TCRaβ+ 3G11− 6C10− CD4+ CD8− thymocyte subset

A murine CD4⁺ thymocyte subset with phenotype of TCRαβ⁺ 3G11⁻ 6C10⁻ CD4⁺ CD8⁻ CD69^+/- HSA^med/lo contains the cells in relatively functional matured status. The functional property of the cells in this subset is characterized by the unique pattern of cytokine production at transitional stage from Th0 to Th2 type with the latter being the dominant type. After being co-cultured with murine thymic medullary epithelial cell line (MTEC1) cells, a murine thymic medullary type epithelial cell line, the TCRαβ⁺ 3G11⁻ 6C10⁻ CD4⁺ CD8⁻ CD69^+/- HSA^med/lo thymocytes, has exhibited significantly higher levels of proliferation capability and IL-6 production, whereas the production of IL-4 and IL-10 is suppressed after co-culturing with MTECl. By contrast, MTECl could not induce thymocytes to secrete Thl type of cytokines. The results suggest that MTECl can regulate functional status of this thymocyte subset and induce them to develop into a specialized Th2 subset.