Dibasic phosphorylation sites in the R domain of CFTR have stimulatory and inhibitory effects on channel activation

To better understand the mechanisms by which PKA-dependent phosphorylation regulates CFTR channel activity, we have assayed open probabilities (P_o), mean open time, and mean closed time for a series of CFTR constructs with mutations at PKA phosphorylation sites in the regulatory (R) domain. Forskolin-stimulated channel activity was recorded in cell-attached and inside-out excised patches from transiently transfected Chinese hamster ovary cells. Wild-type CFTR and constructs with a single Ser-to-Ala mutation as well as octa (Ser-to-Ala mutations at 8 sites) and constructs with one or two Ala-to-Ser mutations were studied. In cell-attached patches, Ser-to-Ala mutations at amino acids 700, 795, and 813 decreased P_o, whereas Ser-to-Ala mutations at 737 and 768 increased P_o. In general, differences in P_o were due to differences in mean closed time. For selected constructs with either high or low values of P_o, channel activity was measured in excised patches. With 1 mM ATP, P_o was similar to that observed in cell-attached patches, but with 10 mM ATP, all constructs tested showed elevated P_o values. ATP-dependent increases in P_o were due to reductions in mean closed time. These results indicate that R-domain phosphorylation affects ATP binding and not the subsequent steps of hydrolysis and channel opening. A model was developed whereby R-domain phosphorylation, in a site-dependent manner, alters equilibrium between forms of CFTR with low and high affinities for ATP. site-directed mutagenesis; kinase-dependent activation; cell-attached patch clamp; open probability; mean open time     

THE CRUCIFORM MUSCLE AND ITS ASSOCIATED SENSE ORGAN IN SCROBICULARIA PLANA (DA COSTA)

Long dendritic processes from neurones of the sense organ ganglionpenetrate the epithelial lining of the narrow and short ductsituated between the intramuscular slit and a large posteriorcavity which opens to the siphonal space through a narrow canalin the sense organ papilla. Motor nerve fibres innervate theanterior ends of the cruciform muscle strands. The latter becomesstretched during opening of animal and contracted during closure.The slit is widened during opening and narrowed during closure.The sense organ responds to foul water suggesting that it isa chemoreceptor for this stimulus. (Received 10 July 1977;     

The Difference Between Open and Closed Meristems

An open and a closed root meristem have been compared by investigatingthe cell kinetics of small regions of the apices of Helianthusand Zea. The cells of the stelar pole are quiescent in both and thereis no exchange of cells between stele and cortex or stele andcap. The immediately distal cells in the closed meristem (Zea)are also quiescent and the few divisions that do occur can betransverse or longitudinal. In the open meristem (Helianthus)these cells are not quiescent, but they go out of cycle transiently,prolonging the potential cell-doubling time. Their divisionsare transverse. It is a consequence of these differences thatclosed meristems form root caps discrete from the cortex whereasopen meristems force instability in the boundary between theperipheral part of the cap and the cortex. Another consequencein roots with open meristems is a succession of columella complexestransversely displaced from each other by the state of fluxin the meristem during the non-cycling phase of the proximaltier of cells, those immediately distal to the stelar pole. The results are discussed in relation to the ontogenetic onsetof quiescence and the evidence for switches between open andclosed operation of meristems. meristem, root apex, Helianthus annuus, Zea mays L.     

Differences between Acer saccharum Leaves from Open and Wind-Protected Sites

Size-correlated variations of sugar maple (Acer saccharum L.)leaf anatomy and mechanical properties were determined for twosaplings (from open and wind protected sites) to examine theeffects of chronic wind-induced mechanical disturbance on leafsize, rigidity and flexibility. Based on a total sample of 78leaves, comparisons indicated that the mean size of the opensite leaves (n =37) was smaller in every measured respect comparedwith that of the closed site leaves (n =41). Open site leaveshad, on average, smaller lamina surface area, shorter and narrowerpetioles with a smaller volume fraction of lignified tissuesthan those from the closed site. Biomechanical comparisons alsoindicated that the petioles of open site leaves were significantlyless rigid and more flexible than the petioles of closed siteleaves. Despite differences in mean leaf size and petiolar rigidityand flexibility, allometric comparisons indicated the size-dependentvariations in leaf properties were continuous across the twosites. Also, the allocation of leaf biomass with respect tostem biomass along the lengths of the two saplings was statisticallyidentical and indistinguishable from an isometric relationship.However, the smaller diametered branches of the open site saplingbore smaller and fewer leaves with less stiff and rigid petiolesthan those of the closed site sapling. The differences betweenopen and closed site leaves are interpreted to be functionallyadaptive and to indicate that chronic mechanical disturbanceof developingAcer saccharum leaves prefigures mature leaf sizeand petiole properties that have the capacity to reduce winddrag. Results from petioles are contrasted with those of mechanicallydisturbed stems. Leaves; Acer saccharum ; biomechanics; wind-drag; allometry     

Physiological Controls on the Production of Cleistogamous and Chasmogamous Flowers in Lamium amplexicaule L

Plants of Lamium amplexicaule, grown under short-day field conditionsin Northern California produce predominantly closed flowers.Under long-day field conditions, plants may produce up to 50per cent open flowers, the same total number of flowers beingproduced under each daylength regime. A 20 ml drop of GA₃ or GA₇ (100µM) was applied to themain shoot apex of plants growing under short-day conditions,and all subsequently produced flowers opened. CCC, an inhibitorof gibberellin synthesis, was applied (0.06 per cent) to thesoil of seedlings grown under long-day conditions. The CCC-treatedplants were dwarfed and bore only closed flowers. With GA₇ appliedexogenously to the CCC-treated shoots, inhibition was released,resulting in elongated internodes and open flower production.The timing of flower production and internodal extension inLamium amplexicaule are positively correlated. When floral primordiawere removed from main shoots, the average internode lengthsdecreased. The number of nodes produced in treated plants wasincreased as a result of flower removal. Exogenous GA₇ (10 µM)applied to the nodes from which flowers were removed resultedin internodal extension but had no effect on node number. Twoprocesses that may contribute to the control of the productionof open and closed flowers in Lamium amplexicaule are: (1) anincreasing anther sac size from lower to upper node flowersthat may exert control locally, via GN production, on corollaexpansion, and (2) a photoperiodic control. Lamium amplexicaule L, henbit, cleistogamy, chasmogamy, gibberellins, gibberellic acid (GA₃), (2 chloroethyl) trimethylammonium chloride (cycocel)     

Direct Measurements of Turgor Pressure Potentials of Guard Cells, I.

Measurements were made of pressures applied in subsidiary andguard cells which caused closure and opening of stomatal pores.These pressures were lower than would be expected from plasmolyticallydetermined osmotic potentials of guard cell saps. The pressuresneeded in guard cells to open almost closed stomata were practicallythe same as those required to close open stomata when appliedin adjacent subsidiary cells. Completely closed stomata couldnot be opened by this technique. The implications for the understandingof the mechanism of guard cell deformation, the Spannungsphase,and the pressure potentials of guard cells are discussed.     

An Electron Microscope Investigation into the Structure of the Floridean Pit

Ultra-thin sections of the red algae Rhodymenia palmata andLaurencia caespitosa were examined in the electron microscopein order to determine the fine structure of the Floridean pits.The pits found in the two species examined are shown to be quitedifferent from each other, in Rhodymenia the pit is open sothat the cytoplasm is continuous through the cell wall, whilstin Laurencia the pit is completely closed by a lamella of thecell wall. These two types are shown to be equivalent to thosedescribed by Jungers (1933) on the basis of light microscopeobservations.     

Major Element Composition of Epidermal and Mesophyll Tissues2Commelina Communis L. and Vicia Faba L.: Some Further Considerations of the Role of Ions in Stomatal Functioning

Epidermal and mesophyll tissues of Commelina communis L. andVicia faba L. were analysed by atomic absorption spectrometryfor the major plant inorganic cations and anions (K, Na, Ca,Mg, P, NO₃-N, Cl) when stomata of the leaf were open and closed.Water-soluble and residual levels of the elements were estimatedand a charge balance of the soluble fraction made. The major portion of K, Na, Cl, and P was extracted in the water-solublefraction of the epidermal and mesophyll tissues of both species.In both species the bulk of Ca remained in the insoluble residueof the epidermis whereas in mesophyll tissue it was equallydistributed be-between the two fractions in C. communis butmainly in the insoluble residue in V. faba. Magnesium was predominantlyfound in the water-soluble fraction of V. faba mesophyll tissueand distributed approximately equally between the two fractionsin the epidermal tissue. In C. communis Mg was slightly moreabundant in the water-soluble fraction of both mesophyll andepidermis. In both species no statistically significant differences inthe levels of the elements could be detected between epidermaland mesophyll tissues from leaves with open stomata and thesame tissues from leaves with closed stomata, suggesting thatthere was no major flux of ions between mesophyll and epidermisduring stomatal movements. Regardless of whether the stomata were open or closed, therewere considerably more water-soluble inorganic cations thananions present in all tissues of both species with K being themajor cation and Cl being the major anion. In V.faba and C-communis epidermis there was 49–53 per cent and 56%68per cent excess cation respectively. In the mesophyll tissuethe excess cation was 63–75 per cent and 75%78 per centin V.faba and C. communis respectively. When the partitioning of the levels of the elements betweenepidermis and mesophyll of a leaf is considered, except forNO₃-N in both species and Na in V. faba, 20 per cent or lessof each element was present in the epidermis.     

S-adenosyl-L-homocysteine hydrolase is necessary for aldosterone-induced activity of epithelial Na(+) channels

The A6 cell line was used to study the role ofS-adenosyl-L-homocysteine hydrolase (SAHHase) inthe aldosterone-induced activation of the epithelial Na⁺channel (ENaC). Because aldosterone increases methylation of severaldifferent molecules, and because this methylation is associated withincreased Na⁺ reabsorption, we tested the hypothesis thataldosterone increases the expression and activity of SAHHase protein.The rationale for this work is that general methylation may be promotedby activation of SAHHase, the only enzyme known to metabolize SAH, apotent end-product inhibitor of methylation. Although aldosteroneincreased SAHHase activity, steroid did not affect SAHHase expression.Antisense SAHHase oligonucleotide decreased SAHHaseexpression and activity. Moreover, this oligonucleotide, as well as apharmacological inhibitor of SAHHase, decreased aldosterone-inducedactivity of ENaC via a decrease in ENaC open probability. The kineticsof ENaC in cells treated with antisense plus aldosterone were similarto those reported previously for the channel in the absence of steroid. This is the first report showing that active SAHHase, in part, increases ENaC open probability by reducing the transition rate fromopen states in response to aldosterone. Thus aldosterone-induced SAHHase activity plays a critical role in shifting ENaC from a gatingmode with short open and closed times to one with longer open andclosed times.

     

An Electrophysiological Study of the Stomatal Complex of Tradescantia virginiana

Measurements of electrical potential difference (PD) and potassiumactivity were carried out on the intact leaf of Tradescantiavirginiana. PD gradients across the stomatal complex were observedwith both open and closed stomata. The guard cell PD appearedto be linearly related to stomatal aperture. With the stomataopen a gradient of potassium activity across the stomatal complexwas observed which became reversed on stomatal closure. Calculationof the driving forces on potassium suggested that it was distributedpassively between the vacuoles of the cells of the stomatalcomplex. The electrophysiological data obtained from this investigationenabled potassium activity in the apoplast to be calculated.The results showed that on stomatal closure there was a massiveincrease in the potassium activity in the guard cell wall. Key words: Stomata, Ionic gradients, Electrical potentials     

Vascular and General Anatomy of the Rootstocks of Three Stemless Viola Species

The primary vascular system of the rootstock of Viola rotundifolia,V. odorata and V. cucullata consists of an open system of threesympodia, corresponding to three orthostichies (parastichies)of leaves in the 1 /3 phyllotaxy. Between the major, collateralbundles there are vascular strands consisting of only primaryphloem. The vascular supply to axillary buds developing in thefirst season of growth is the same in flowering and vegetativebuds and there is a homology between the bracts on the pedunclesof axillary flowers and the prophylls of the axillary vegetativebranches. The overwintering portions of the rootstocks are somewhatwoody. The uneven development of secondary xylem correspondsto the locations of the sympodia. Secondary xylem closes a leafgap of a median trace only slowly, but any gap in the phloicsystem is closed quickly by secondary growth. Viola rotundifolia, V. odorata, V. cucullata, rootstock, vascular anatomy     

Hunting Behavior in the Ctenizidae

The most primitive way of hunting in Mygalomorph spiders seemsto be the free roaming and catching of encountered prey Theraphosidae.The trap-door spiders Ctenizidae, Actinopodidae and Barychelidawhich are entirely sedentary, lie in wait behind the trap-doorand leap at prey that happens to pass close to the door. Somespecies spin radial silk threads outside the door which functionas stumbleor signal-lines, and some Australian species use grassblades and other litter in the same way. A further evolutionarystep leads to species which do not build a trap-door but crowntheir burrow by a funnel-shaped web. The Dipluridae finallyare real web-builders, which depend on a sheet-web to catchtheir prey. Generally they do not dig a burrow but hide in asmall retreat from which a funnel-web leads to the net. With a few exceptions Ctenizidae are entirely nocturnal. Theirrhythm of activity has been analyzed. The "Zeitgeber" is thedaylight during the last half hour before sunset. Most trapdoorspiders never leave their burrow during their whole life. Theyneed three to four years from hatching to become adults. Adultmales die during or at the end of the restricted mating-season;they take no food when adult. Females, which undergo post-adultmolts, can probably live for 15–20 years. Nemesia caementaria, like most other species, hunts during thewhole night. The mean time of activity is about Si/o hours,consisting of periods of lying in wait and periods of intermediaterests. The spiders make an average of three leaps per nightto catch a prey, but only about 10% of all bounds are successful.A hungry animal, lurking in vain, shows unmotivated leaps. The effects of light, moisture, and temperature on hunting activityare analyzed. Ctenizidae hunt during autumn, winter, and springbut interrupt their activity in summer for an estivation whichlasts usually two months. The females of some species capture the male after mating andeat him; others never attack him. This difference in behaviorlias repercussions on reproduction. Among certain species the young nymphs of the third instar refuseall food until they have made their own burrow and can huntby themselves. The young of other species stay with the motherfor one year and leave her at the fifth or sixth instar to maketheir own burrow. The young of Nemesia caementaria can remainin the burrow of the mother until they are almost adult, i.e.,for two or three years, and feed on the prey the mother hascaught. The behavior of Ctenizidae can be grouped. Some species lurkbehind the closed or almost closed trap-door. Another groupopens the trap-door and puts the pedipalpi and the two anteriorpairs of legs radially out onto the rim of the burrow, whilethe cephalothorax is hidden behind the trap-door. A third groupspins threads of silk radially about the entrance and uses themas signal-lines. A fourth group can come out and pursue theprey, then drags it to the trap-door and into the burrow. The nature of the prey depends on the bio tope. The Ctenizidaefeed almost exclusively on insects, mainly ants and beetles;in the laboratory crickets are accepted as prey. Ctenizidae have no tarsal organ. They have different types oftrichobothria, transversal and longitudinal slit-organs, andlyriform organs. With these three kinds of sense-organs, thefunctions of which are not clearly understood, the trap-doorspiders are able to perceive the approach of prey, to judgeits distance from the trap-door, and to locate it in direction.They seem to have an organ of smell, since certain groups ofinsects are repulsive to them. Sight is not used for hunting.     

Evolution of sensory structures in basal metazoa

Cnidaria have traditionally been viewed as the most basal animalswith complex, organ-like multicellular structures dedicatedto sensory perception. However, sponges also have a surprisingrange of the genes required for sensory and neural functionsin Bilateria. Here, we: (1) discuss "sense organ" regulatorygenes, including; sine oculis, Brain 3, and eyes absent, thatare expressed in cnidarian sense organs; (2) assess the sensoryfeatures of the planula, polyp, and medusa life-history stagesof Cnidaria; and (3) discuss physiological and molecular datathat suggest sensory and "neural" processes in sponges. We thendevelop arguments explaining the shared aspects of developmentalregulation across sense organs and between sense organs andother structures. We focus on explanations involving divergentevolution from a common ancestral condition. In Bilateria, distinctsense-organ types share components of developmental-gene regulation.These regulators are also present in basal metazoans, suggestingevolution of multiple bilaterian organs from fewer antecedentsensory structures in a metazoan ancestor. More broadly, wehypothesize that developmental genetic similarities betweensense organs and appendages may reflect descent from closelyassociated structures, or a composite organ, in the common ancestorof Cnidaria and Bilateria, and we argue that such similaritiesbetween bilaterian sense organs and kidneys may derive froma multifunctional aggregations of choanocyte-like cells in ametazoan ancestor. We hope these speculative arguments presentedhere will stimulate further discussion of these and relatedquestions.     

Effect of upper airway negative pressure on inspiratory drive during sleep

To determine the effect of upper airway(UA) negative pressure and collapse during inspiration on regulation ofbreathing, we studied four unanesthetized female dogs duringwakefulness and sleep while they breathed via a fenestratedtracheostomy tube, which was sealed around the permanent trachealstoma. The snout was sealed with an airtight mask, thereby isolatingthe UA when the fenestration (Fen) was closed and exposing the UA tointrathoracic pressure changes, but not to flow changes, when Fen wasopen. During tracheal occlusion with Fen closed, inspiratory time(TI) increased duringwakefulness, non-rapid-eye-movement (NREM) sleep and rapid-eye-movement(REM) sleep (155 ± 8, 164 ± 11, and 161 ± 32%,respectively), reflecting the removal of inhibitory lung inflationreflexes. During tracheal occlusion with Fen open (vs. Fen closed):1) the UA remained patent;2)TI further increased duringwakefulness and NREM (215 ± 52 and 197 ± 28%, respectively) but nonsignificantly during REM sleep (196 ± 42%);3) mean rate of rise of diaphragmEMG (EMGdi/TI) and rate offall of tracheal pressure(Ptr/TI) were decreased,reflecting an additional inhibitory input from UA receptors; and4) bothEMGdi/TI andPtr/TI were decreasedproportionately more as inspiration proceeded, suggesting greaterreflex inhibition later in the effort. Similar inhibitory effects ofexposing the UA to negative pressure (via an open tracheal Fen) wereseen when an inspiratory resistive load was applied over severalbreaths during wakefulness and sleep. These inhibitory effectspersisted even in the face of rising chemical stimuli. This inhibitionof inspiratory motor output is alinear within an inspiration andreflects the activation of UA pressure-sensitive receptors by UAdistortion, with greater distortion possibly occurring later in theeffort.

     

The Function of the Hilum in Some Papilionaceae in Relation to the Ripening of the Seed and the Permeability of the Testa

In seeds of Trifolium repens, T. pratense, and Lupinus arboreus,the hilum is a hygroscopically activated valve in the impermeableepidermis of the testa. When relative humidity was low the fissurein the hilum opened permitting the seed to dry out; when therelative humidity was high the fissure closed obstructing theabsorption of moisture. During seed-ripening the moisture contentfell readily to approximately 25 per cent., and thereafter moreslowly until the epidermis became impermeable at approximately14 per cent, moisture content. Further drying of the seed tookplace only by diffusion of water vapour through the hilum. ‘Hard’seeds tended to have a moisture content in equilibrium withthe lowest relative humidity to which they had been exposed.They absorbed moisture under conditions of gradually increasingrelative humidity such that the hilar fissure remained open.The duration of the impermeable condition increased with thedegree of desiccation brought about by loss of water throughthe hilum.     

The First Record of Fossil Wood of Winteraceae from the Upper Cretaceous of Antarctica

Fossil wood of the Winteraceae from the Upper Cretaceous sedimentsof James Ross Island, Antarctic Peninsula, is described herefor the first time. The specimen is characterized by the absenceof vessels, rays of two distinct sizes and tracheids with one–threerows of circular bordered pits, mainly on the radial walls,grading to horizontally elongate and scalariform. Despite anatomicalconformity to the family Winteraceae, the fossil wood is notidentical to any one extant genus and therefore has been assignedto the fossil organ genus Winteroxylon Gottwald with which thefossil shows greatest similarity. Copyright 2000 Annals of BotanyCompany Antarctica, Cretaceous, angiosperm, wood, anatomy, Winteraceae, Winteroxylon, fossil, palaeoclimate     

Low-pressure Storage of Hibiscus Cuttings. Effect on Stomatal Opening and Rooting

Cuttings of Hibiscus rosa-sinensis L. cv. Moesiana were storedat 15 or 760 mmHg (2 or 101.3 kPa) for 3, 5, 8 or 11 weeks.No ethylene production could be detected regardless of treatment.No chlorophyll degradation, proline accumulation or respiratoryincrease were seen during storage. Stomata were open under lowpressure and closed under normal pressure. The open stomatacaused water loss from cuttings after removal from storage becausethey were unable to respond normally to low humidity. Rootingability decreased as storage time increased, but satisfactoryresults were obtained up to a storage period of five weeks. Adventitious rooting, cuttings, ethylene production, Hibiscus rosa-sinensis L., hypobaric storage, low-pressure storage, senescence, stomatal resistance     

光学活性拟除虫菊酯对棉铃虫神经细胞钠通道电流的影响

用全细胞膜片钳技术对比分析了alpha体氯氰菊酯与theta体氯氰菊酯对棉铃虫Helicoverpa armigera幼虫离体培养中枢神经细胞Na⁺通道门控过程的影响。结果表明,alpha体氯氰菊酯作用后,神经细胞Na⁺通道电流（I_Na）先增大,同时通道的激活电压向负电位方向移动约10 mV,提示alpha体氯氰菊酯使通道激活电位降低,通道更容易被激活。药剂作用约10 min后,I_Na又迅速降低,表明alpha体氯氰菊酯对开放状态的Na⁺通道有抑制作用。另外,alpha体氯氰菊酯使I_Na到达峰值的时间缩短,但对失活时间无明显影响。Theta体氯氰菊酯也使I_Na激活电位左移,幅值降低,但降低速率较慢。总的结果表明alpha体氯氰菊酯与theta体氯氰菊酯对棉铃虫中枢神经细胞处于关闭和开放状态的钠通道均有作用,且alpha体氯氰菊酯对钠通道电流的抑制作用强于theta体氯氰菊酯。     

The Complex Satellite DNA of Tropaeolum majus L.: Partial Characterization of Isolated and of Cloned Restriction Fragments

Satellite DNA (satDNA) was purified by repeated neutral CsCl,ethidium bromide/CsCl and actinomycin D/CsCl density centrifugationfrom total nuclear DNA of Tropaeolum majus. Digestion patternsof satDNA and main band DNA as generated with 22 different restrictionenzymes were partially characterized and hybridized by the Southernblot technique with Hpa II-generated satDNA fragments and aribosomal DNA fragment. Some restriction fragments were sequencedby the chemical method, either directly, or after cloning, andpossible secondary structures were determined. The results showthat the satellite is of high complexity and evidently composedof different, interspersed repeats, and is also interspersedwith ribosomal sequences. Some sequences of the satellite aredispersed throughout the Tropaeolum genome, and even found inthe chloroplast genome. Most of the restriction enzymes cutthe satDNA into many fragments of lengths between 14 bp and3000 bp. Hpa II cut the satellite DNA into at least 63 differentsized fragments (more than is known from any other plant satDNA).Sal I cut the satDNA into a single fragment of 1040 bp in size,which probably represented the superrepeat unit. Sequence analysisof satDNA restriction fragments and of cloned fragments isolatedat different times, indicated a rapid diversification in vivoand in vitro. Computer modeling of most likely secondary structuressuggest the occurrence of palindromic loops (cruciform structures).These could be the basis of rapid diversification in being sitesof preferred intragenomic recombination and rearrangement. Cloning, palindromic sequences, restriction analysis, satellite DNA complexity, Tropaeolum majus L, nasturtium     

Cell Division in Tetraspora

Cell division in Tetraspora sp. is described. The cell becomesimmotile some while before mitosis and the basal bodies withdrawfrom the cell surface. The preprophase nucleus migrates to thebasal body complex, around which increasing numbers of microtubulesgather. The spindle is closed with open polar fenestrae; a basalbody complex is always closely associated with at least onepole. No spindles were observed to have basal bodies at bothpoles, and the spindle may possibly be unicentric. During anaphase,spindle microtubules penetrate through the fenestrae. Aftertelophase, the nuclei come together as a phycoplast forms betweenthem; cytokinesis is effected by furrowing. Forming basal bodiesare frequently encountered in late telophase and cleaving cells;no evidence was obtained that the basal bodies replicated beforemitosis. The protoplast rotates inside the cell wall duringcleavage. Cell division is compared with that of other greenalgae, and in particular, Chlamydomonas.