Regulation of a heart potassium channel by protein kinase A and C

The enzymes adenosine 3',5'-monophosphate (cAMP)-dependent protein kinase (protein kinase A) and protein kinase C regulate the activity of a diverse group of cellular proteins including membrane ion channel proteins. When protein kinase A was stimulated in cardiac ventricular myocytes with the membrane-soluble cAMP analog 8-chlorphenylthio cAMP (8-CPT cAMP), the amplitude of the delayed-rectifier potassium current (IK) doubled when recorded at 32 degrees C but was not affected at 22 degrees C. In contrast, modulation of the calcium current (ICa) by 8-CPT cAMP was independent of temperature with similar increases in ICa occurring at both temperatures. Stimulation of protein kinase C by phorbol 12,13-dibutyrate also enhanced IK in a temperature-dependent manner but failed to increase ICa at either temperature. Thus, cardiac delayed-rectifier potassium but not calcium channels are regulated by two distinct protein kinases in a similar temperature-dependent fashion.     

Calcium as a mediator of adrenocorticotrophic hormone action on adrenal protein synthesis

Calcium stimulates leucine incorporation into protein during incubations of sections and cell-free preparations of the rat adrenal. Like adrenocorticotrophic hormone (ACTH) action, calcium enhances the transfer of amino acid from transfer RNA to protein. Stimulation of leucine incorporation by ACTH and cyclic adenosine monophosphate is best observed when sections are incubated in limiting calcium concentrations.     

A spatial-temporal model of cell activation

A spatial-temporal model of calcium messenger function is proposed to account for sustained cellular responses to sustained stimuli, as well as for the persistent enhancement of cell responsiveness after removal of a stimulus, that is, cellular memory. According to this model, spatial separation of calcium function contributes to temporal separation of distinct phases of the cellular response. At different cellular sites, within successive temporal domains, the calcium messenger is generated by different mechanisms and has distinct molecular targets. In particular, prolonged cell activation is brought about by the interaction of calcium with another spatially confined messenger, diacylglycerol, to cause the association of protein kinase C with the plasma membrane. Activity of the membrane-associated protein kinase C is controlled by the rate of calcium cycling across the plasma membrane. In some instances, a single stimulus induces both protein kinase C activation and calcium cycling and thus causes prolonged activation; but in others, a close temporal association of distinct stimuli brings about cell activation via interaction of these intracellular messengers. Persistent enhancement of cell responsiveness after removal of stimuli is suggested to be due to the continued association, or anchoring, of protein kinase C to the membrane.     

A potential second messenger role for unsaturated fatty acids: activation of Ca2+-dependent protein kinase

Arachidonate and other unsaturated long-chain fatty acids were found to activate protein kinase C from human neutrophils. Kinase activation by arachidonate required calcium and was enhanced by diolein but did not require exogenous phosphatidylserine. Submaximal levels of arachidonate also enhanced the affinity of the kinase for calcium during activation by phosphatidylserine. Thus the release of arachidonate, which is triggered in many cell types by ligand-receptor interactions, could play a second messenger role in the regulation of cellular function by activation of protein kinase C.     

Juvenile hormone induced vitellogenin synthesis in the monarch butterfly

The vitellogenin (yolk protein) of the monarch butterfly has been identified by electrophoretic and immunochemical techniques. In freshly emerged female adults ligature of the neck prevents appearance of this protein in the hemolymph; it prevents yolk deposition and egg maturation as well. These processes are restored by injection of juvenile hormone; the restoration involves induction of vitellogenin synthesis, as shown by incorporation of [(3)H]leucine.     

Calcium modulation activates Epstein-Barr virus genome in latently infected cells

In many viral infections the host cell carries the viral genome without producing viral particles, a phenomenon known as viral latency. The cellular mechanisms by which viral latency is maintained or viral replication is induced are not known. The modulation of intracellular calcium concentrations by calcium ionophores induced Epstein-Barr viral antigens in lymphoblastoid cell lines that carry the virus. When calcium ionophores were used in conjunction with direct activators of protein kinase C (12-O-tetradecanoyl phorbol-13-acetate and a synthetic diacylglycerol), a greater induction of viral antigens was observed than with either agent alone. Activation of protein kinase C may be required for the expression of the viral genome.     

Female specific protein: biosynthesis controlled by corpus allatum in Leucophaea maderae

A specific protein is Present in serums of all of Leucophaea maderae that mature eggs but not in serums of males, nymphs of either sex, or females that are reproductively inactive. Ablations, microsurgery and remplantation showed that this protein is produced de novo under the influence of the corpus allatum hormone. This protein, essential for egg maturation, is synthesized in isolated abdimens of allatectomized females treated with 0.08 to 0.12 microgram of the t,t,t-isomer of the authentic juvenile hormone. The crops allatum hormone also stimulates toa similar degree the synthesis of other proteins which appear to be essential for egg maturation as well; these are not specific to females.     

甲壳动物生长发育的激素调控研究概述

蜕皮贯穿甲壳动物生长发育的始终。概述了调控甲壳动物蜕皮的主要内分泌腺(Y-器官、大颚器官、X-器官窦腺复合体等)以及外源性激素(高等动物性激素、脊椎动物生长激素、外源性蜕皮激素和外源性保幼激素拮抗物),其中保幼激素拮抗物作为甲壳动物促生长剂,高效、低成本、相对安全。     

Generation of calcium oscillations in fibroblasts by positive feedback between calcium and IP3

A wide variety of nonexcitable cells generate repetitive transient increases in cytosolic calcium ion concentration ([Ca2+]i) when stimulated with agonists that engage the phosphoinositide signalling pathway. Current theories regarding the mechanisms of oscillation disagree on whether Ca2+ inhibits or stimulates its own release from internal stores and whether inositol 1,4,5-trisphosphate (IP3) and diacylglycerol (DG) also undergo oscillations linked to the Ca2+ spikes. In this study, Ca2+ was found to stimulate its own release in REF52 fibroblasts primed by mitogens plus depolarization. However, unlike Ca2+ release in muscle and nerve cells, this amplification was insensitive to caffeine or ryanodine and required hormone receptor occupancy and functional IP3 receptors. Oscillations in [Ca2+]i were accompanied by oscillations in IP3 concentration but did not require functional protein kinase C. Therefore, the dominant feedback mechanism in this cell type appears to be Ca2+ stimulation of phospholipase C once this enzyme has been activated by hormone receptors.     

Cell cycle-dependent coupling of the calcitonin receptor to different G proteins.

Calcitonin is a calcium regulating peptide hormone with binding sites in kidney and bone as well as in the central nervous system. The mechanisms of signal transduction by calcitonin receptors were studied in a pig kidney cell line where the hormone was found to regulate sodium pumps. Calcitonin receptors activated the cyclic adenosine monophosphate (cAMP) or the protein kinase C (PKC) pathways. The two transduction pathways required guanosine triphosphate (GTP)-binding proteins (G proteins) (the choleratoxin sensitive Gs and the pertussis toxin sensitive Gi, respectively) and led to opposite biological responses. Moreover, selective activation of one or the other pathway was cell cycle-dependent. Therefore, calcitonin may induce different biological responses in target cells depending on their positions in the cell cycle. Such a modulation of ligand-induced responses could be of importance in rapidly growing cell populations such as during embryogenesis, growth, and tumor formation.     

Intracellular calcium release mediated by sphingosine derivatives generated in cells

Soluble and hydrophobic lipid breakdown products have a variety of important signaling roles in cells. Here sphingoid bases derived in cells from sphingolipid breakdown are shown to have a potent and direct effect in mediating calcium release from intracellular stores. Sphingosine must be enzymically converted within the cell to a product believed to be sphingosine-1-phosphate, which thereafter effects calcium release from a pool including the inositol 1,4,5-trisphosphate-sensitive calcium pool. The sensitivity, molecular specificity, and reversibility of the effect on calcium movements closely parallel sphingoid base-mediated inhibition of protein kinase C. Generation of sphingoid bases in cells may activate a dual signaling pathway involving regulation of calcium and protein kinase C, comparable perhaps to the phosphatidylinositol and calcium signaling pathway.     

Feedback control of juvenile hormone synthesis in cockroaches: possible role for ecdysterone

Inactive female corpora allata implanted into adult males become active and continue to synthesize juvenile hormone at high rates. However, when an ovary is implanted together with the corpora allata, rates of juvenile hormone synthesis decline as the oocytes complete maturation. Injections of ecdysterone mimic the effect of an implanted ovary.     

Primary structure of the beta subunit of the DHP-sensitive calcium channel from skeletal muscle

Complementary DNAs for the beta subunit of the dihydropyridine-sensitive calcium channel of rabbit skeletal muscle were isolated on the basis of peptide sequences derived from the purified protein. The deduced primary structure is without homology to other known protein sequences and is consistent with the beta subunit being a peripheral membrane protein associated with the cytoplasmic aspect of the sarcolemma. The protein contains sites that might be expected to be preferentially phosphorylated by protein kinase C and guanosine 3',5'-monophosphate-dependent protein kinase. A messenger RNA for this protein appears to be expressed in brain.     

化学放射法测定蜜蜂保幼激素酯酶活性

蜜蜂保幼激素(JH)是由蜜蜂咽侧体(CA)合成和分泌的一种氧化物。保幼激素酯酶是对蜜蜂体内呈游离态或结合态的保幼激素进行专化性降解的特异性酯酶。采用放射化学法(RC)测定了西方蜜蜂(Apis mellifera L．)不同日龄工蜂体内保幼激素酯酶(JHE)的活性。结果表明，工蜂体内保幼激素酯酶活性随着日龄的增大而下降。     

内蒙古白绒山羊蛋白激酶B/AKT基因的克隆及表达模式分析

 【目的】克隆内蒙古白绒山羊AKT基因cDNA并分析其基本表达模式。【方法】RT-PCR克隆AKT基因 cDNA。通过在线软件BLAST进行核酸序列分析,用SMART与Psite进行氨基酸序列分析。半定量RT-PCR检测AKT基因在绒山羊组织中的表达特异性。Western blotting检测绒山羊胎儿成纤维细胞中AKT表达。【结果】克隆到的内蒙古白绒山羊AKT基因cDNA片段长 1 443 bp,包含了编码480个氨基酸残基的全长ORF,氨基酸序列与绵羊（NM_001161857.1）同源性为97%。SMART分析表明,ORF编码的蛋白包含了可与3-磷酸肌醇结合的PH结构域及具有丝氨酸/苏氨酸激酶催化活性的S_TKc结构域。Psite分析表明,含有1个cAMP-/cGMP-依赖性蛋白激酶磷酸化位点、6个蛋白激酶C磷酸化位点、10个酪蛋白激酶Ⅱ磷酸化位点、2个蛋白激酶ATP结合区信号和1个丝氨酸/苏氨酸蛋白激酶活性区域。PSORT程序预测其定位于细胞质中。AKT基因mRNA丰度在睾丸、脑和肾中较高,在脾、肝、肺及乳腺组织中相对低。绒山羊胎儿成纤维细胞中抑制mTOR活性,AKT表达量降低。【结论】内蒙古白绒山羊AKT基因cDNA全长ORF的核苷酸序列与绵羊的AKT基因具有很高的同源性,AKT基因在脾、睾丸、脑、肝、肺、乳腺及肾组织中均有表达,其AKT的表达受mTOR信号通路的调控。     

PDGF-induced activation of phospholipase C is not required for induction of DNA synthesis

Platelet-derived growth factor (PDGF) induction of DNA synthesis is believed to involve activation of phospholipase C (PLC) and subsequent accumulation of inositol 1,4,5-triphosphate [I(1,4,5)P3], increase in intracellular Ca2+, activation of protein kinase C (PKC), and receptor down regulation. Generation of these events is triggered by the tyrosine protein kinase (TPK) activity of the PDGF receptor. The TPK inhibitor genistein blocked PDGF induction of these events, including DNA synthesis, with the exception of receptor down regulation. PDGF-induced phosphotyrosine phosphorylations, including receptor autophosphorylation, were inhibited by genistein. Removal of genistein and PDGF resulted in DNA synthesis without the occurrence of PLC activation. These findings indicate that these early events, with the exception of receptor down regulation, are not necessary for PDGF-induced DNA synthesis.     

Molecular mimicry regulates ABA signaling by SnRK2 kinases and PP2C phosphatases

Abscisic acid (ABA) is an essential hormone for plants to survive environmental stresses. At the center of the ABA signaling network is a subfamily of type 2C protein phosphatases (PP2Cs), which form exclusive interactions with ABA receptors and subfamily 2 Snfl-related kinase (SnRK2s). Here, we report a SnRK2-PP2C complex structure, which reveals marked similarity in PP2C recognition by SnRK2 and ABA receptors. In the complex, the kinase activation loop docks into the active site of PP2C, while the conserved ABA-sensing tryptophan of PP2C inserts into the kinase catalytic cleft, thus mimicking receptor-PP2C interactions. These structural results provide a simple mechanism that directly couples ABA binding to SnRK2 kinase activation and highlight a new paradigm of kinase-phosphatase regulation through mutual packing of their catalytic sites.     

黄河鲇幼鱼对饲料中钙、磷的需要量

为探明黄河鲇Silurus lanzhouensis幼鱼对饲料钙、磷的适宜需要量,选取总能、钙、粗蛋白质、非植酸磷、赖氨酸5个因素作为试验因素,每个因素各设6个水平,钙为0.50%~1.34%,非植酸磷为0.50%~1.00%,采用均匀设计法U*6(64)设计6种配合饲料,饲喂体质量为(170±5)g的黄河鲇幼鱼,分别记为A、B、C、D、E、F组,饲养时间为60 d。试验结束后,以增重率、特定生长率、肌肉和肝脏RNA/DNA值、粗蛋白质离体消化率为试验指标,采用偏最小二乘法建立回归方程,通过降维分析和优化求解,分析研究黄河鲇幼鱼对饲料中钙和非植酸磷的适宜需要量。结果表明:C组(钙0.56%、非植酸磷0.60%)试验鱼的增重率、特定生长率和肌肉RNA/DNA最高,分别为84.83%、1.02%和2.98,肠、肝、胃中粗蛋白质离体消化率也达到最大值,分别为67.53%、66.76%、51.52%;随着饲料中钙、非植酸磷含量的增加,鱼体增重率、特定生长率、肌肉RNA/DNA值和粗蛋白质离体消化率均呈现先上升后下降的趋势。研究表明,饲料中钙和非植酸磷含量对黄河鲇幼鱼生长、蛋白质合成能力、饲料粗蛋白质离体消化率影响明显,黄河鲇幼鱼对饲料中钙的适宜需要量为0.54%,对非植酸磷的适宜需要量为0.50%~0.53%。     

Parallel single-cell monitoring of receptor-triggered membrane translocation of a calcium-sensing protein module

Time courses of translocation of fluorescently conjugated proteins to the plasma membrane were simultaneously measured in thousands of individual rat basophilic leukemia cells. We found that the C2 domain---a calcium-sensing, lipid-binding protein module that is an essential regulator of protein kinase C and numerous other proteins---targeted proteins to the plasma membrane transiently if calcium was released from internal stores, and persistently in response to entry of extracellular calcium across the plasma membrane. The C2 domain translocation time courses of stimulated cells clustered into only two primary modes. Hence, the reversible recruitment of families of signaling proteins from one cellular compartment to another is a rapid bifurcation mechanism for inducing discrete states of cellular signaling networks.     

A calcium-dependent protein kinase with a regulatory domain similar to calmodulin.

Calcium can function as a second messenger through stimulation of calcium-dependent protein kinases. A protein kinase that requires calcium but not calmodulin or phospholipids for activity has been purified from soybean. The kinase itself binds calcium with high affinity. A complementary DNA clone for this kinase has been identified; it encodes a protein with a predicted molecular mass of 57,175 daltons. This protein contains a catalytic domain similar to that of calmodulin-dependent kinases and a calmodulin-like region with four calcium binding domains (EF hands). The predicted structure of this kinase explains its direct regulation via calcium binding and establishes it as a prototype for a new family of calcium-regulated protein kinases.