Cyclic AMP regulation of glucose transport in germinating Pilobolus longipes spores

Pilobolus longipes spores were activated by either glucose or 6-deoxyglucose. Glucose-induced spore activation was previously shown to follow an increase in intracellular cyclic AMP. Concurrent with glucose-induced spore activation, were shifts in 6-deoxyglucose transport kinetics towards higher V _max and K _m values. Cyclic AMP derivatives also caused spore activation and similar changes in the kinetic parameters of 6-deoxyglucose transport. The time course of activation was paralleled by changes in transport activity. Inhibition of phosphodiesterase alone did not cause activation or induce changes in transport activity, but in combination with sub-optimal levels of either 6-deoxyglucose or cAMP derivatives, it amplified the germination signals to produce large increases in both spore activation and 6-deoxyglucose transport activity. These results support the conclusion that glucose transport in germinating spores is regulated by cAMP.Abbreviations IBMX 3-isobutyl-1-methylxanthine; monobutyryl cyclic AMP - N⁶ monobutyryladenosine 3:5-cyclic monophosphate - 8-bromo cyclic AMP 8-bromoadenosine 3:5-cyclic monophosphate     

Effects of light and temperature on sporangiophore initiation in Pilobolus crystallinus (Wiggers) Tode

Sporangiophore initiation in Pilobolus crystallinus grown in white light was induced by either a dark or a low-temperature treatment. The period of darkness necessary to induce sporangiophore initiation was shortened by lowering the temperature. Arrhenius plots for the sporangiophore-suppressing reaction in both light and darkness consisted of two straight lines with a Q₁₀ of about 2 at lower temperatures and 8–11 at higher temperatures. The temperature at which the Q₁₀ changed was the lower, the higher the fluence rate: 14° C at 8 W/m², 19.5° C at 0.24 W/m² and 24.5° C in darkness. Possible interpretations of these results are briefly presented.Abbreviations %SP percentage of trophocysts initiating sporangiophores - D50% duration of treatment required to 50% sporangiophore initiation     

Physiology and fine structure of sporangiospore germination in Piptocephalis unispora prior to infection

Germination of the sporangiospore of Piptocephalis unispora Benjamin, observed by means of light and electron microscopy, involved the formation of a new inner wall which became continous with the inner layer of the wall of the germ tube. The outer wall layer of the germ tube was continous with the original inner wall layer of the dormant spore. Preliminary details of appressorium structure were noted. Nutritional experiments indicated that sporangiospores required external sources of utilisable nitrogen and carbon compounds for maximal swelling and germ tube production. Limited development occurred when either nutrient was supplied singly. Comparison of germination of the asexual spore with that in other Mucorales, especially the Kickxellaceae, has been made, and the merosporangial status in P. unispora discussed.Non-Standard Abbreviations CH casein hydrolysate - Q spore quotient     

Light germination ofAnthoceros miyabeanus spores

The effects of light on the spore germination of a hornwort species,Anthoceros miyabeanus Steph., were investigated. Spores of this species were photoblastic, but their sensitivities to light quality were different. Under either continuous white, red or diffused daylight, more than 80% of the spores germinated, but under blue light none or a few of them germinated. Under continuous far-red light or in total darkness, the spores did not germinate at all.Anthoceros spores required red light irradiation for a very long duration, i.e., over 12–24 hr of red light for saturated germination. However, the spore germination showed clear photo-reversibility by repeated irradiation of red and far-red light. The germination pattern clearly varied with the light quality. There were two fundamental patterns; (1) cell mass type in white or blue light: spores divide before germination, and the sporelings divide frequently and form 1–2 rhizoids soon after germination, and (2) germ tube type in red light: spores germinate without cell division, and the single-cell sporelings elongate without cell division and rhizoid formation.     

Blue-light-induced shift of the phototropic fluence-response curve in Pilobolus sporangiophores

The effects of preirradiation with blue light on the shift of the fluence-response curve for the first and the second positive curvatures were examined in Pilobolus crystallinus (Wiggers) Tode sporangiophores. A 1-min preirradiation with blue light at 47 or 960 nmol·m^-2 lowered the fluence-response curve for the first positive curvature and shifted the peak to a higher fluence. The fluence-response curve was shifted back to a lower fluence when a dark period was inserted between the preirradiation and the curvature-inducing light. This shift back to lower fluence was biphasic when the fluence was high (960 nmol · m^-2), indicating the participation of two components in the phototropic reaction for the first positive curvature.The fluence-response curve for the second positive curvature did not seem to be shifted to a higher fluence region when fluence was varied by varying exposure time. However, the fluence-response curve obtained by varying the fluence rate of a 20-min irradiation period indicated that the second positive curvature was also shifted to a higher-fluence region by preirradiation with blue light. A small shoulder appeared on the fluence-response curve when preirradiation at a high fluence rate was given.Abbreviations BL blue light - CIL curvature-inducing light     

Phototropic fluence-response curves for Pilobolus crystallinus sporangiophore

Fluence-response relationships were examined for positive and negative phototropism induced by blue (450 nm) and ultraviolet-B (UV-B, 280 nm) light, respectively, in the Pilobolus crystallinus sporangiophore. Fluence-response curves for both blue and UV-B light obtained by changing the fluence by varying exposure time only showed the classical first and second positive bending. However, fluence-response curves obtained by varying the fluence rate were bell-shaped irrespective of the length of the exposure time. With increasing exposure time the peak became higher along the ascendant arm and the descendant arm was shifted toward the higher fluence. The Bunsen-Roscoe reciprocity law was valid only when the fluence was less than approx. 400 pmol·m^-2 for both blue and UV-B light. Because the shapes of the fluence-response curves for blue and UV-B light were nearly the same, the photoreceptor systems for both blue and UV-B light are considered to be the same.Abbreviation UV-B ultraviolet-B     

Bioprospecting for novel oxylipins in fungi: The presence of 3-hydroxy oxylipins in Pilobolus

As previously found in various members of the Mucorales, 3-hydroxy oxylipins in Mucor genevensis are associated with the sporangia, i.e. mainly the columella structure and between aggregating sporangiospores. To determine if this phenomenon is also true in distantly related members, the mucoralean fungus Pilobolus was examined. This fungus is characterized by relatively large sub sporangial-columella structures which actively eject sporangia in a sticky liquid for attachment onto herbage surrounding its growth medium – in this case horse dung. Strikingly, this fungus produced a novel oxylipin i.e. a 3-hydroxy monounsaturated fatty acid, possibly a nonenoic acid, which is mainly associated with the sub sporangial-columella structure and aggregating sporangiospores. The specificity of the antibody against 3-hydroxy oxylipins used in immunofluorescence mapping of the mucoralean fungi, was further confirmed in the yeast, Saccharomycopsis malanga which produces 3-hydroxy palmitate in crystal form. These crystals occur between aggregating yeast cells. On the basis of the available data, we hypothesize that 3-hydroxy oxylipins probably function as adhesives, attaching fungal cells to each other or to other surfaces through entropic based hydrophobic forces and/or hydrogen bonds.     

The arrangement of F-actin and microtubules during germination of Mucor rouxii sporangiospores

The distribution of F-actin microfilaments and microtubules was analyzed in germinating sporangiospores of Mucor rouxii by labeling with rhodamine-tagged phalloidin and by immunofluorescence microscopy. The transition from isodiametrical to apical growth was accompanied by a switch from uniform distribution of F-actin patches to a polarized accumulation of F-actin material at the germ tube tips. Immunoblotting of cell-free extracts of M. rouxii with a monoclonal anti-porcine -tubulin antibody (TU-01) disclosed two discrete bands of -tubulin suggesting the existence of two -tubulin genes in this fungus. Immunofluorescence microscopy of germinating cells stained with the same antibody revealed an elaborate network of cytoplasmic microtubules that persisted during the entire germination process and extended into the apex of the germ tube. Although their precise roles remain undetermined, the observed arrangement of cytoskeletal elements during germination is consistent with their presumed involvement in cell wall morphogenesis: the long axial microtubules serving as long-distance conveyors of wall-building vesicles to the apical region while the concentrated F-actin patches mark the participation of microfilaments in the zone of intense vesicle exocytosis at the hyphal apex.Abbreviations DAPI 4,6-diamidino-2-phenylindole - DTT dithiothreitol - EGTA Ethylene glycol-bis (beta-aminoethyl ether) - N,N,N,N tetraacetic acid - F-actin Filamentous actin - MES 2-(N0morpholino)-ethanesulfonic acid - PIPES Piperazine-N,N-bis-2-ethanesulfonic acid - PMSF Phenyl-methylsulphonyl fluoride - TBS Tris-buffered saline     

Analysis of growth and rotational behavior of sporangiophores in Pilobolus crystallinus (Wiggers) Tode

The growth and rotation of the sporangiophore of Pilobolus crystallinus, which are important factors in its phototropic behavior, were analyzed throughout its development. The sporangiophore initial emerged from the trophocyst and elongated at the extreme tip without rotating. The elongation rate of the sporangiophore apex then gradually decreased and the apex expanded radially to produce the sporangium, but no rotation occurred. A transient cessation of elongation after sporangium development was followed by resumption of both elongation and radial expansion in the region beneath the sporangium developing the subsporangial vesicle. Rotation was not obvious at this stage. Radial expansion of the subsporangial vesicle continued at a decreasing rate until full size was reached. Elongation then recommenced in the newly established growth zone in the upper region of the sporangiophore just beneath the subsporangial vesicle. During this period of growth, the sporangiophore rotated in a clockwise direction as viewed from above. All growth and rotation ceased about 1 h before ejection of the sporangium into the air. Based on these results, a modified classification of the developmental stages has been proposed.This work was carried out under the Joint Research Program of the Institute of Genetic Ecology, Tohoku University, Japan (892006). The authors please to thank Kaori Koga and Hiroko Kikuchi for their helpful assistance.     

Effect of light and temperature on the germination of lettuce seeds

A short period (15–30 min) at 30° C promotes germination of seeds of Lactuca sativa L. cv. Repolhuda in darkness. Far-red light reverses this stimulation, and the escape curves for phytochrome and high-temperature action are quite similar, indicating that the two factors act at a common point in the chain of events leading to germination. It is suggested that high temperature acts by decreasing the threshold of the active, far-red absorbing, form of phytochrome (Pfr) needed to promote germination.Abbreviations FR far-red light - Pfr far-red-absorbing form of phytochrome - R red light     

环境因子对蕨类植物孢子萌发的影响

蕨类植物通过孢子萌发形成独立生活的配子体,配子体能够形成精子器和颈卵器,进而通过受精作用形成新的孢子体。孢子萌发是蕨类植物生活史过程中配子体世代向孢子体世代转变的关键步骤。同时,此过程不仅受到多种环境因子的影响,也是研究细胞核极性移动、细胞不对称分裂、假根极性生长等独特的细胞学事件的良好模型。迄今为止,人们已经研究发现多种环境因子对约200余种蕨类植物孢子萌发有影响。总结了环境因子对蕨类植物孢子萌发影响的规律如下:(1)孢子萌发除了受到光照强度影响外,主要受光质的影响,光质的影响主要表现为4种方式:①孢子萌发受红光刺激与远红光抑制像开关一样调控;②孢子萌发不受远红光抑制;③孢子萌发受蓝光抑制;④孢子只能在黑暗条件下萌发。(2)重力作用会影响孢子细胞核移动,进而影响孢子细胞发育的极性。(3)赤霉素(GA)能增加孢子萌发率或帮助孢子打破休眠。成精子囊素与GA作用相似,启动或促进孢子萌发。而脱落酸(ABA)、茉莉酸(JA)和乙烯等其它激素对孢子萌发的影响相对较小。(4)不同植物孢子有着各自最适的萌发培养基条件,如不同种类孢子对MS培养基中无机盐含量、蔗糖含量、pH值的要求不同。孢子外被中的Ca2+、Mn2+和Mg2+,培养基中的Cd2+和La3+,以及孢子接种密度、萌发空间CO2含量也会对孢子萌发造成影响。(5)多数蕨类植物孢子在15-30℃可以萌发,最适萌发温度为25℃。(6)4℃和液氮储藏可以延长孢子寿命并保持较高萌发率。     

Effects of nutrients and light pretreatment on phytochrome-mediated fern-spore germination

Spores of the ferns, Dryopteris filix-mas, D. paleacea and Polystichum minutum, sown on plain agar in quartz-distilled water, required several hours of red light in order to germinate. When, however, water agar was replaced by agar made up with a mineral nutrition medium, a single pulse of red light (about 1 min) was able fully to induce germination. Under these conditions spores became light-sensitive a few minutes after sowing. Thus, zero germination in dark controls was obtained only when all light was excluded immediately after sowing or when saturating far-red was given thereafter. The effect of the mineral medium was also obtained using low ion concentrations with an osmolality of less than 100 mol l^–1. Thus, a specific ion effect appears more probable than an unspecific osmotic effect. Species differences in light sensitivity and in dark-germination levels, as reported in the literature, might partly be the consequence of different culture media and of light acting at a very early stage after sowing, which hitherto was assumed to be still insensitive to light. On water agar as well as on mineral agar, the inducing effect of a single red pulse could be increased by the appropriate pretreatment, i.e. by preirradiation with red light for several hours, followed by a saturating pulse of far-red, the latter abolishing the direct inducing effect of the red preirradiation. The nature of both the ion-phytochrome interaction and the phytochrome-phytochrome interaction has not yet been analysed.Abbreviations FR saturating far-red light - Pfr far-red absorbin form of phytochrome - R broad-band red light, acting continuously during several hours This work was performed at the Department of Plant Physiology, University of Lund, Sweden, during a sabbatical leave     

Spore germination in Dictyostelium discoideum

Mutant spores of Dictyostelium discoideum, strain SG-10, differ from wild type spores in their ability to spontaneously germinate, to be activated with 5% dimethyl Sulfoxide (DMSO), and to be deactivated with 0.2 M sucrose. Both heat-activated wild type and mutant spores began to swell after a lag of 60–75 min at ambient temperature. Suspension of heat activated spores in 5% DMSO resulted in blockage of spore swelling and a concomitant severe inhibition of respiration; removal of 5% DMSO allowed resumption of respiration and the spores began to swell after a lag of only 15 min. It was concluded that 5% DMSO allowed the early reactions (M) to proceed but blocked the later reactions (R) of post-activation lag.Treatment of one day old spores with 20% DMSO solution for 30–120 min quantitatively activated the population. The post-activation lag time was directly dependent on the time of 20% DMSO treatment. Spores activated with 20% DMSO treatment could be deactivated by incubation at 0°C; the spores most quickly deactivated at 0°C were those within 10 min of swelling. Mitochondrial transport inhibitors such as azide and cyanide caused deactivation in an analogous manner. It is hypothesized that spores proceed to the second portion of the lag phase called (R) before the environment determines if dormancy is reimposed or if germination will proceed. The sensitive strain (SG-10) showed a greater degree of damage than the wild type after supraoptimal treatment with 40% DMSO. The spores became more resistant with age to the damaging action of 40% DMSO. All the observed effects of DMSO treatment were compatible with our multistate model of activation which suggests that the early portion of the lag phase (M) may involve a relative uncoupling of oxidative phosphorylation while the later portion (R) may require tight coupling.     

Low temperature seed germination of several tomato genotypes

Environmental stresses at particularly vulnerable stages during crop development may severely diminish productivity. At temperature of 10 °C or below cultivated tomato germinate slowly if at all. In this study, seven tomato genotypes bred at the Research Institute of Vegetable Crops were evaluated for germination time at 10 °C. Analysis identified that one genotype which has L. chilense in its pedigree, germinated most rapidly while four other genotypes germinated slower. After 21 days, four out of five of the genotypes resulted in seed germination from 81 to 98 %.     

Storage reserve mobilization during low temperature germination and early seedling growth in Brassica napus

In western Canada, oilseed rape (Brassica napus L. var. oleifera cv. Westar) is seeded during the early months of spring, when ambient temperatures are well below the optimum. This can result in poor seedling emergence. The objectives of the present study were to determine which developmental stages are sensitive to low temperature and whether the effects are thermal or developmental in nature. Seed was germinated at 22, 10 and 6 °C. Fresh weight changes and seedling growth were assessed on the basis of equal accumulated heat units, and the mobilization of storage reserves was assessed by employing antibodies against isocitrate lyase (ICL; EC 4.1.3.1), oleosin and cruciferin. Additionally, de novo protein synthesis was determined by quantifying the incorporation of methionine via in vivo labelling. Low temperature resulted in poor germination and early seedling growth with phase II of germination being most sensitive. At 10 °C, there was a temporal delay in germination that did not affect the overall success of germination. This was a thermal effect as seed at the lower temperatures required the equivalent of 16–24 degree days before germination occurred. Also, seedling growth at 10 °C was lower in comparison to seedlings grown at 22 °C. Seed at 6 °C displayed slow and incomplete germination and poor seedling growth as a result of both thermal and developmental effects.     

Interaction of light and temperature on seed germination of Rumex obtusifolius L.

Germination of Rumex obtusifolius L. seeds (nutlets) is low in darkness at 25° C. Germination is stimulated by exposure to 10 min red light (R) and also by a 10-min elevation of temperature to 35° C. A 10-min exposure to far-red light (FR) can reverse the effect of both R (indicating phytochrome control) and 35° C treatment. Fluence-response curves for this reversal of the effect of R and 35° C treatments are quantitatively identical. Treatment for 10 min with light of wavelenght 680, 700, 710 and 730 nm, after R and 35° C treatment, demonstrates that germination induced by 35° C treatment results from increased sensitivity to a pre-existing, active, far-red-absorbing form of phytochrome (Pfr) in the seeds.Abbreviations FR far-red light - P phytochrome - Pr red-absorbing form of P - Pfr far-red-absorbing form of P - R red light     

Calcium requirement of phytochrome-mediated fern-spore germination: No direct phytochrome-calcium interaction in the phytochrome-initiated transduction chain

Phytochrome-mediated germination of fern spores of Dryopteris paleacea Sw. was initiated by a saturating red-light (R) irradiation after 20 h of imbibition. For its realization external Ca²⁺ was required, with a threshold at a submicromolar concentration, and an optimum was reached around 10^-4 M. At concentrations 10^-1 M only a reduced response was obtained, based probably on an unspecific osmotic or ionic effect. The germination response was inhibited by La³⁺, an antagonist of Ca²⁺. From these results it is concluded that Ca²⁺ influx from the medium into the spores may be an important event in phytochrome-mediated germination. In the absence of Ca²⁺ the R-stimulated system remained capable of responding to Ca²⁺, added as late as 40 h after R. Moreover, Ca²⁺ was effective even if added after the active form of phytochrome, Pfr, had been abolished by far-red (FR) 24 h after R. Thus, the primary effect of Pfr, that initiates the transduction chain, does not require calcium. Coupling of Pfr to subsequent dark reactions has been investigated by R-FR irradiations with various dark intervals. The resulting escape kinetics were characterized by a lag phase (6 h) and half-maximal escape from FR reversibility (19 h). These kinetics were not significantly changed by the presence or absence of calcium. Thus, direct interaction of Pfr and calcium is not a step in the transduction chain initiated by the active form of photochrome.Abbreviations EGTA ethyleneglycol-bis(-aminoethyl ether)-N,N,N,N-tetraacetic acid - FR far-red light - Pr red-light-absorbing form of phytochrome - Pfr far red-light-absorbing form of phytochrome - Pipes piperazine-1,4-bis(2-ethanesulfonic acid) - R red light A preliminary report of this work was presented at the XIV Int. Bot. Congr., Berlin (West), Germany, Book of Abstracts, 2-116a-5 (1987)     

两种云杉种子萌发和幼苗生长对环境因子的适应性

青海云杉是我国沙区通过种子育苗引种成功的物种。通过对不同储藏时间青海云杉和沙地云杉种子生活力测定、种子萌发对温度、光照、水盐胁迫以及幼苗生长对水盐胁迫和沙埋的响应实验,比较两种种子萌发和幼苗生长对环境因子适应性,为沙地云杉在我国沙区广泛引种提供参考。结果表明:1)在2年的储藏过程中,沙地云杉和青海云杉种子生活力分别由79%和72%下降了19%和5%; 2)沙地云杉和青海云杉种子适宜萌发温度分别为15—30℃和10—30℃,最适萌发温度分别为25℃(72%)和25/15℃(69%),除10℃和10/30℃外,两种种子萌发率在各温度下没有显著差异; 3)沙地云杉种子萌发光照条件为14 h光照/8 h黑暗交替(67%),青海云杉为24 h光照(61%)或24 h黑暗(61%); 4)水势在-2.7—0 MPa时,2种云杉的IGR(初始萌发率)、RGR(恢复萌发率)、ISL(初始幼苗长度)和RSL(恢复幼苗长度)均没有显著差异; 5) NaCl浓度在200 mmol/L和250 mmol/L,青海云杉种子IGR显著大于沙地云杉,NaCl浓度在0—450 mmol/L,青海云杉和沙地云杉种子RGR没有显著差异,当NaCl浓度为50 mmol/L和100 mmol/L,青海云杉ISL显著大于沙地云杉; 6)在0.5—2.0 cm沙埋深度时,青海云杉出苗率显著高于沙地云杉,沙地云杉最适沙埋深度0.5 cm,青海云杉为0.5—1.5 cm。因此,青海云杉种子萌发和幼苗生长比沙地云杉有更强的环境适应性,但只要采取合理的播种时间、播种深度和水分管理等措施,沙地云杉会和青海云杉一样在我国沙区大面积引种育苗。     

Influence of salinity and temperature on the germination of Kochia scoparia

Kochia scoparia is one of the most common annual halophytes foundin the Great Basin. Seeds were collected from a population growing in asalt playa at Faust, Utah and were germinated at 5 temperature regimes(12 h night/12 h day, 5–15 ^°C, 10–20 ^°C, 15–25 ^°C,20–30 ^°C and 25–35 ^°C) and 6 salinities (0, 200, 400,600, 800 and 1000 mM NaCl) to determine optimal conditions forgermination and recovery of germination from saline conditions after beingtransferred to distilled water. Maximum germination occurred in distilledwater, and an increase in NaCl concentration progressively inhibited seedgermination. Few seeds germinated at 1000 mM NaCl. A temperatureregime of 25 ^°C night and 35 ^°C day yielded maximumgermination. Cooler temperature 5–15 ^°C significantly inhibited seedgermination. Rate of germination decreased with increase in salinity.Germination rate was highest at 25–35 ^°C and lowest at5–15 ^°C. Seeds were transferred from salt solutions to distilled waterafter 20 days and those from high salinities recovered quickly at warmertemperature regimes. Final recovery germination percentages in high salttreatments were high, indicating that exposure to high concentration ofNaCl did not inhibit germination permanently.