Swelling and Ca2+-activated Anion Conductances in C127 Epithelial Cells Expressing WT and ΔF508-CFTR

CFTR is a chloride channel that is required for fluid secretion and salt absorption in many exocrine epithelia. Mutations in CFTR cause cystic fibrosis. CFTR expression influences some ion channels, but the range of channels influenced, the mechanism of the interaction and the significance for cystic fibrosis are not known. Possible interactions between CFTR and other ion channels were studied in C127 mouse mammary epithelial cell lines stably transfected with CFTR, ΔF508-CFTR, or vector. Cell lines were compared quantitatively using an ¹²⁵I efflux assay and qualitatively using whole-cell patch-clamp recording. As expected, ¹²⁵I efflux was significantly increased by forskolin only in the CFTR line, and forskolin-stimulated whole-cell currents were time- and voltage independent. All three lines responded to hypotonic challenge with large ¹²⁵I efflux responses of equivalent magnitude, and whole-cell currents were outwardly rectified and inactivated at positive voltages. Unexpectedly, basal ¹²⁵I efflux was significantly smaller in the ΔF508-CFTR cell line than in either the CFTR or control cell lines (P < 0.0001), and the magnitude of the efflux response to ionomycin was largest in the vector cell line and smallest in the cell line expressing ΔF508-CFTR (P < 0.01). Whole-cell responses to ionomycin had a linear instantaneous I-V relation and activated at depolarizing voltages. Forskolin responses showed simple summation with responses to ionomycin or hypotonic challenge. Thus, we found no evidence for interactions between CFTR and the channels responsible for swelling-mediated responses. Differences were found in basal and ionomycin-stimulated efflux, but these may arise from variations in the clonally selected cell lines that are unrelated to CFTR expression. Received: 15 November 1995/Revised: 16 February 1996     

Role of protein kinase A and the serine-rich region of herpes simplex virus type 1 ICP4 in viral replication.

Efficient expression of herpes simplex virus genes requires the synthesis of functional ICP4, a nuclear phosphoprotein that contains a prominent serine-rich region between amino acids 142 and 210. Residues in this region not only are potential sites for phosphorylation but also are involved in the functions of ICP4. By comparing the growth of a virus in which this region is deleted (d8-10) with wild-type virus (KOS) in PC12 cells or PC12 cells that are deficient in cyclic AMP-dependent protein kinase (PKA), two observations were made: (i) the growth of wild-type virus was impaired by 1 to 2 orders of magnitude in the PKA-deficient cells, indicating the involvement of PKA in the growth cycle of herpes simplex virus type 1, and (ii) while the growth of d8-10 was impaired by almost 2 orders of magnitude in wild-type cells, it was not further impaired (as was that of wild-type virus) in PKA-deficient cells, implicating the region deleted in d8-10 as a possible target for cellular PKA. In trigeminal'ganglia of mice, the d8-10 mutant virus grew poorly; however, it established latency in nearly 90% of ganglia tested. Studies of the phosphorylation of wild-type and d8-10 ICP4 proteins revealed that the serine-rich region is a major determinant for phosphorylation of ICP4 in vivo and that the phosphorylation state could change as a function of the PKA activity. Consistent with this observation, the serine-rich region of ICP4 was shown to be a target for PKA in vitro. While intact ICP4 was readily phosphorylated by ICP4 in vitro, the d8-10 mutant ICP4 was not. Moreover, a synthethic peptide representing a sequence in the serine tract that is predicted to be a substrate for PKA was phosphorylated by PKA in vitro, having a Km within the physiological range. These data suggest that PKA plays a role in viral growth through phosphorylation of one or more sites on the ICP4 molecule.     

中国野豌豆属的分类研究

本文报道了国产野豌豆属43种,4变种及6变型,其中包括4个新种(多叶野豌豆,三尖野豌豆,武山野豌豆,长齿野豌豆);一个新变种(三叶歪头菜)及一个新等级(千山野豌豆)。     

Calcium binding to the subunit c ofE. coli ATP-synthase and possible functional implications in energy coupling

The 8-kDa subunit c of theE. coli F₀ ATP-synthase proton channel was tested for Ca⁺⁺ binding activity using a⁴⁵Ca⁺⁺ ligand blot assay after transferring the protein from SDS-PAGE gels onto polyvinyl difluoride membranes. The purified subunit c binds⁴⁵Ca⁺⁺ strongly with Ca⁺⁺ binding properties very similar to those of the 8-kDa CF₀ subunit III of choloroplast thylakoid membranes. The N-terminal f-Met carbonyl group seems necessary for Ca⁺⁺ binding capacity, shown by loss of Ca⁺⁺ binding following removal of the formyl group by mild acid treatment. The dicyclohexylcarbodiimide-reactive Asp-61 is not involved in the Ca⁺⁺ binding, shown by Ca⁺⁺ binding being retained in twoE. coli mutants, Asp61Asn and Asp61Gly. The Ca⁺⁺ binding is pH dependent in both theE. coli and thylakoid 8-kDa proteins, being absent at pH 5.0 and rising to a maximum near pH 9.0. A treatment predicted to increase the Ca⁺⁺ binding affinity to its F₀ binding site (chlorpromazine photoaffinity attachment) caused an inhibition of ATP formation driven by a base-to-acid pH jump in whole cells. Inhibition was not observed when the Ca⁺⁺ chelator EGTA was present with the cells during the chlorpromazine photoaffinity treatment. An apparent Ca⁺⁺ binding constant on the site responsible for the UV plus chlorpromazine effect of near 80–100 nM was obtained using an EGTA-Ca⁺⁺ buffer system to control free Ca⁺⁺ concentration during the UV plus chlorpromazine treatment. The data are consistent with the notion that Ca⁺⁺ bound to the periplasimic side of theE. coli F₀ proton channel can block H⁺ entry into the channel. A similar effect occurs in thylakoid membranes, but the Ca⁺⁺ binding site is on the lumen side of the thylakoid, where Ca⁺⁺ binding can modulate acid-base jump ATP formation. The Ca⁺⁺ binding to the F₀ and CF₀ complexes is consistent with a pH-dependent gating mechanism for control of H⁺ ion flux across the opening of the H⁺ channel.This work was supported in part by grants from the Department of Energy and the U.S. Department of Agriculture.On leave from the Institute of Soil Science and Photosynthesis, Russian Academy of Science, Pushchino, Russia.     

48例原发性闭经患者的细胞遗传学分析

本文报告对48例原发闭经患者的临床和细胞遣传学分析,共发现染色体异常17例,占35.4%,其中包括45,X,7例;45,X/46,XX,2例;X染色体结构异常5例;核型中有Y染色体3例。讨论了原发闭经的细胞遗传学病因及异常核型与表型的关系。     

多年生黑麦草草地生态系统中放牧强度对草地结构及组织转化的影响

研究了放牧强度对多年生黑麦草人工草地蘖的形态、密度、草地生产率及组织转化的影响。结果表明,重牧条件下蘖密度大于轻牧,而轻牧的单株蘖重大于重牧。重牧划地净生产率大于轻牧,主要是由于轻和手条件下,草地的高的生长率被更高的枯死率所抵消。春夏之交,采用灵活的管理措施,转换放牧强度可以提高草地的生产率。     

中国的草蛉及其地理分布（脉翅目：草蛉科）

本文讨论了中国草蛉科已知属种的地理分布及其特点,指出：属的分布以东洋成分比重较大,高山高原属分化明显;种类分布呈现东洋成分由东部沿海向北挺进、古北种类通过西北地区向西南延伸的格局,丰富的高山、高原种类及狭布种类,是我国草蛉区系的一大特点。     

中国无患子科的地理分布

无患子科是一个乏热带分布科,全世界有１４４属,１６８０种,本文着重阐述：１．无患子科的分类系统及演化;２．世界无患子科的地理分布;３．中国无患子科的地理分布;４．无患子科的早期化地点及散布途径。结果表明无患子科现代分布变异中心在亚洲（印度－马西亚）－非洲－南美洲。中国无患子科有２５属,５４种,特有成分外,均与印度一马来西亚共有,大多数位于分布区的边缘。根据化石记录及现代及分布推断,无患子科可能在白     

L-异亮氨酸和甘氨酸对大肠杆菌表达与分泌邻苯二酚2,3-双加氧酶的作用

证实了甘氨酸与L-异亮氨酸对大肠杆菌表达邻苯二酚2,3-双加氧酶(CatO_2ase)的促进作用和甘氨酸促使该酶分泌至胞外培养基中的作用.产酶量高低和分泌量多少与培养基种类、甘氨酸和L-异亮氨酸的浓度以及培养时间等因素有关.在甘氨酸存在的情况下,胞壁对溶菌酶的敏感性有所增加,超微形态似有变化,还存在其他物质的伴随分泌,故甘氨酸可能是引起细胞壁结构的改变而导致邻苯二酚2,3-双加氧酶等胞内容物被动分泌至胞外.