Brief irradiation with red or blue light induces orientational movement of chloroplasts in dark-adapted prothallial cells of the fernAdiantum

Orientational movement of chloroplasts was induced by a brief irradiation with red light (R) or blue light (B) in dark-adapted prothallial cells ofAdiantum, whose chloroplasts had gathered along the cell dividing wall (i.e., the anticlinal wall). When the whole dark-adapted prothallia were irradiated from a horizontal direction (i.e., from their lobes) with horizontally vibrating polarized R (H pol. R) for 10 or 3 min, the chloroplast left the anticlinal walls and spread over the prothallial surface (i.e., the periclinal walls) within 1–2 hr after the onset of irradiation, returning to the anticlinal wall (dark-position) within 10 hr. However, vertically vibrating polarized R (V pol. R) for 10 min did not induce the movement towards periclinal walls. The R effect was cancelled by non-polarized far-red light (FR) irradiation just after the R irradiation. Irradiation with H pol. B for 10 or 3 min but not with V pol. B could also induce a similar movement of chloroplasts, although the chloroplasts returned within 4 hr. The effect of H pol. B, however, was not cancelled by the subsequent FR irradiation. When a part of the dark-adapted cell at the prothallial surface was irradiated from above with a microbeam of R or B for 1 min, chloroplasts of the cell in the dark-position moved towards the irradiated locus in subsequent darkness. However, in the neighboring cells, orientational movement was not induced by either R or B microbeams. These results show that in dark-adapted prothallial cells, both brief irradiation with R and B can induce chloroplast photo-orientation and that the photoreceptors are phytochrome and blue light-absorbing pigment, respectively. It is also clear that effects of both R and B irradiation do not transfer to neighboring cells.     

Phytochrome- and blue-light-absorbing pigment-mediated directional movement of chloroplasts in dark-adapted prothallial cells of fernAdiantum as analyzed by microbeam irradiation

When prothalli ofAdiantum capillus-veneris L. were kept for 2 d in the dark, chloroplasts gathered along the anticlinal walls (Kagawa and Wada, 1994, J Plant Res 107: 389–398). In these dark-adapted prothallial cells, irradiation with a microbeam (10 gm in diameter) of red (R) or blue light (B) for 60 s moved the chloroplasts towards the irradiated locus during a subsequent dark period. Chloroplasts located less than 20 gm from the center of the R microbeam (18 J·m^–2) moved towards the irradiated locus. The higher the fluence of the light, the greater the distance from which chloroplasts could be attracted. The B microbeam was less effective than the R microbeam. Chloroplasts started to move anytime up to 20 min after the R stimulus, but with the B microbeam the effect of the stimulus was usually apparent within 10 min after irradiation. The velocity of chloroplast migration was independent of light-fluence in both R and B and was about - 0.3 m·min^–1 between 15 min and 30 min after irradiation. Whole-cell irradiation with far-red light immediately after R- and B-microbeam irradiations demonstrated that these responses were mediated by phytochrome and a blue-light-absorbing pigment, respectively. Sequential treatment with R and B microbeams, whose fluence rates were less than the threshold values when applied separately, resulted in an additive effect and induced chloroplast movement, strongly suggesting that signals from phytochrome and the blue-light-absorbing pigment could interact at some point before the induction of chloroplast movement.Abbreviations B blue light - FR far-red light - IR infrared light - R red light     

Low temperature-induced chloroplast relocation mediated by a blue light receptor,phototropin 2, in fern gametophytes

Chloroplast movement in response to light has been known more than 100 years. Chloroplasts move towards weak light and move away from strong light. Dark-induced relocation, called dark positioning, has also been shown. However, the effects of other stimuli on chloroplast movement have not been well characterized. Here we studied low temperature-induced chloroplast relocation (termed cold positioning) in prothallial cells of the gametophytes of the fern Adiantum capillus-veneris. Under weak light chloroplasts in prothallial cells accumulated along the periclinal wall at 25 degrees C, but they moved towards anticlinal walls when the prothalli were subsequently transferred to 4 degrees C. A temperature shift from 25 degrees to 10 degrees C or lower was enough to induce cold positioning, and high-intensity light enhanced the response. Nuclei also relocated from the periclinal position (a position along periclinal walls) to the anticlinal position (a position along anticlinal walls) under cold temperature, whereas mitochondria did not. Cold positioning was not observed in mutant fern gametophytes defective of the blue light photoreceptor, phototropin 2.     

Phototropins and neochrome1 mediate nuclear movement in the fern Adiantum capillus-veneris

In gametophytic cells (prothalli) of the fern Adiantum capillus-veneris, nuclei as well as chloroplasts change their position according to light conditions. Nuclei reside on anticlinal walls in darkness and move to periclinal or anticlinal walls under weak or strong light conditions, respectively. Here we reveal that red light-induced nuclear movement is mediated by neochrome1 (neo1), blue light-induced movement is redundantly mediated by neo1, phototropin2 (phot2) and possibly phot1, and dark positioning of both nuclei and chloroplasts is mediated by phot2. Thus, both the nuclear and chloroplast photorelocation movements share common photoreceptor systems.     

Phytochrome-mediated polarotropism of Adiantum capillus-veneris L. protonemata as analyzed by microbeam irradiation with polarized light

Summary Perception of polarized light inducing phytochrome-mediated polarotropism in protonemata of the fern Adiantum capillus-veneris L. was analyzed using brief microbeam irradiation with polarized red (R) or far-red light (FR). The polarotropic response inducible by irradiation of the subapical 10–30-m part with polarized R vibrating parallel to the cell axis was nullified by subsequently giving R at the apical 0–2.5-m region. This inhibitory effect of R showed an action dichroism, that is, polarized R vibrating normal to the cell axis was effective but the parallel-vibrating R was not. On the other hand, FR irradiation of the extreme tip after irradiation of the whole cell with depolarized R effectively induced a tropic response. This FR effect also showed action dichroism, with parallel-vibrating polarized FR being more effective than FR vibrating normal to the cell axis. When the apical-dome region and the adjacent subapical 10–20-m region were sequentially irradiated with polarized R vibrating obliquely in different directions, polarotropism took place depending on the vibrating direction of the light given to the apical-dome region. Obliquely vibrating polarized FR given to the apical dome after irradiation of the whole cell with depolarized R also induced polarotropism. Thus, the difference in amount (or percent) of the far-redabsorbing form of phytochrome (Pfr) between the extreme tip and the subapical region appears to be crucial in regulating the direction of apical growth; the difference in Pfr level between the two sides of the protonemal apex may occur mainly at the apical dome. Furthermore, the transition moments of the red-absorbing form of phytochrome (Pr) and Pfr seem to be aligned parallel and normal, respectively, to the cell surface at the periphery of the apical hemisphere.Abbreviations FR far-red light - Pfr far-red-absorbing form of phytochrome - Pr red-absorbing form of phytochrome - R red light     

Actin-based mechanisms for light-dependent intracellular positioning of nuclei and chloroplasts in Arabidopsis

The plant organelles, chloroplast and nucleus, change their position in response to light. In Arabidopsis thaliana leaf cells, chloroplasts and nuclei are distributed along the inner periclinal wall in darkness. In strong blue light, they become positioned along the anticlinal wall, while in weak blue light, only chloroplasts are accumulated along the inner and outer periclinal walls. Blue-light dependent positioning of both organelles is mediated by the blue-light receptor phototropin and controlled by the actin cytoskeleton. Interestingly, however, it seems that chloroplast movement requires short, fine actin filaments organized at the chloroplast edge, whereas nuclear movement does cytoplasmic, thick actin bundles intimately associated with the nucleus. Although there are many similarities between photo-relocation movements of chloroplasts and nuclei, plant cells appear to have evolved distinct mechanisms to regulate actin organization required for driving the movements of these organelles.Key words: actin, Arabidopsis, blue light, chloroplast positioning, phototropin, nuclear positioning     

Effects of blue and red light on unrolling of rice leaves

Unrolling of the second leaf of 8-day-old rice (Oryza sativa L.) seedlings was promoted by weak blue light (B), but not by red light (R). The effect of B was counteracted by irradiation with R just before or after the B. The counteracting effect of R was reversed by subsequent irradiation with far-red light but not by B, even if B was applied for 10 h. The B was effective when the region 0.5–2 cm from the tip of the leaf was irradiated. These results indicate that in rice photoreceptors for blue light located in the region 0.5–2 cm from the tip of the leaf play a key role in leaf unrolling and that a B-absorbing pigment and phytochrome participate in leaf unrolling in a closely related manner.Abbreviations B blue light - R red light - FR far-red light - W white light - D dark This work was presented at the Annual Meeting of the Japanese Society of Plant Physiologists on April 4, 1978, in Hiroshima     

Development of the endosperm in rice (Oryza sativa L.): Cellularization

The syncytial endosperm of rice undergoes cellularization according to a regular morphogenetic plan. At 3 days after pollination (dap) mitosis in the peripheral synctium ceases. Radial systems of microtubules emanating from interphase nuclei define nuclear-cytoplasmic domains (NCDs) which develop axes perpendicular, to the embryo sac wall. Free-growing anticlinal walls between adjacent NCDs compart-mentalize the cytoplasm into open-ended alveoli which are overtopped by syncytial cytoplasm adjacent to the central vacuole. At 4 dap, mitosis resumes as a wave originating adjacent to the vascular bundle. The spindles are oriented parallel to the alveolar walls and cell plates formed in association with interzonal phragmoplasts result in periclinal walls that cut off a peripheral layer of cells and an inner layer of alveoli displaced toward the center. Polarized growth of the newly formed alveoli and elongation of the anticlinal walls occurs during interphase. The next wave of cell division in the alveoli proceeds as the first and a second cylinder of cells is cut off inside the peripheral layer. The periods of polarized growth/anticlinal wall elongation alternating with periclinal cell division are repeated 3–4 times until the grain is filled by 5 dap.     

Action spectra for the inhibition of growth in radish hypocotyls

In etiolated seedlings of Raphanus sativus L. the inhibition of hypocotyl elongation by continuous light showed a major bimodal peak of action in the red and far-red, and two minor peaks in the blue regions of the spectrum. It is argued that, under conditions of prolonged irradiation, phytochrome is the pigment controlling the inhibition of hypocotyl elongation by red and far-red light, but that its mode of action in far-red is different from that in red. A distinct pigment is postulated for blue light.Abbreviations B blue - FR far red - G green - R red - HIR high irradiance reaction - P_r and P_fr red and far red absorbing forms of phytochrome - R red     

Regulation of the orientation movement of chloroplasts in epidermal cells of Vallisneria: cooperation of phytochrome with photosynthetic pigment under low-fluence-rate light

In epidermal cells of the leaves of the aquatic angiosperm Vallisneria gigantea Graebner, the chloroplasts accumulate in the outer periclinal layer of cytoplasm (P side) under light at low fluence rates. The nature of such intracellular orientation of chloroplasts was investigated in a semiquantitative manner. Time-lapse video microscopy revealed that, while irradiation with red light (650 nm, 0.41 W · m^–2) rapidly accelerated the migration of chloroplasts, not only from the anticlinal layers of cytoplasm (A sides) to the P side but also from the P side to the A sides, the increased rate of migration in both directions returned to the control rate upon subsequent irradiation with far-red light (746nm, 0.14W · m^–2). These effects of red and far-red light could be observed repeatedly, both in the presence and in the absence of inhibitors of photosynthesis, suggesting the involvement of phytochrome as the photoreceptor. After saturating irradiation with red light, the increased rate of migration of chloroplasts from the P side to the A sides declined more rapidly than the increased rate of migration in the opposite direction. This imbalance in the migration of chloroplasts between the two opposing directions resulted in the accumulation of chloroplasts on the P side. The more rapid decline in the rate of migration of chloroplasts from the P side to the A sides than in the opposite direction was not observed in the presence of an inhibitor of photosynthesis. It appears, therefore, that phytochrome and photosynthetic pigment cooperatively regulate the accumulation of chloroplasts on the P side through modulation of the nature of the movement of the chloroplasts.Abbreviations A side cytoplasmic layer that faces the anticlinal wall - DCMU 3-(3,4-dichlorophenyl)-1,1-dimethylurea - Pfr farred-light-absorbing form of phytochrome - Pr red-light-absorbing form of phytochrome - P side cytoplasmic layer that faces the outer periclinal wall This work was supported in part by Grants-in-Aid from the Japanese Ministry of Education, Science and Culture to S.T. and R.N. The authors are indebted to the Osaka branch of Kashimura Inc. for their kind cooperation in preparing the GREEN software.     

Polarized microtubule deposition coincides with wall ingrowth formation in transfer cells ofVicia faba L. cotyledons

Summary The epidermal transfer cells in developingVicia faba L. cotyledons are highly polarized. Extensive wall ingrowths occur on their outer periclinal walls and extend part way down both anticlinal walls. This ingrowth development serves to increase the surface area of the plasma membrane and thus maximize porter-dependent uptake of sugars from the seed apoplasm. In contrast, the inner periclinal walls of these transfer cells do not form wall ingrowths. We have commenced a study of the mechanisms responsible for establishing this polarity by first analysing the microtubule (MT) cytoskeleton in developing transfer cells. Thin sections of fixed cotyledons embedded in methacrylate resin were processed for immunofluorescence microscopy using monoclonal anti--tubulin and counterstained with Calcofluor White to visualize wall ingrowths. In epidermal cells of young cotyledons where wall ingrowths were yet to develop, MT labelling was detected around all cortical regions of the cell. However, in cells where wall ingrowths were clearly established, MT labelling was detected almost exclusively in cortical regions adjacent to the wall ingrowths. Little, if any, MT labelling was detected on the anticlinal or inner periclinal walls of these cells. This distribution of MTs was most prominent in cells with well developed wall ingrowths. In these cells, a subpopulation of MTs were also detected emanating from the subcortex and extending towards the wall ingrowth region. The possible role of MT distribution in establishing transfer cell polarity and wall ingrowth formation is discussed.Abbreviations MT microtubule     

Blue- and red-light action in photoorientation of chloroplasts in Adiantum protonemata

An action spectrum for the low-fluencerate response of chloroplast movement in protonemata of the fern Adiantum capillus-veneris L. was determined using polarized light vibrating perpendicularly to the protonema axis. The spectrum had several peaks in the blue region around 450 nm and one in the red region at 680 nm, the blue peaks being higher than the red one. The red-light action was suppressed by nonpolarized far-red light given simultaneously or alternately, whereas the bluelight action was not. Chloroplast movement was also induced by a local irradiation with a narrow beam of monochromatic light. A beam of blue light at low energy fluence rates (7.3·10^-3-1.0 W m^-2) caused movement of the chloroplasts to the beam area (positive response), while one at high fluence rates (10 W m^-2 and higher) caused movement to outside of the beam area (negative response). A red beam caused a positive response at fluence rates up to 100 W m^-2, but a negative response at very high fluence rates (230 and 470 W m^-2). When a far-red beam was combined with total background irradiation with red light at fluence rates causing a low-fluence-rate response in whole cells, chloroplasts moved out of the beam area. When blue light was used as background irradiation, however, a narrow far-red beam had no effect on chloroplast distribution. These results indicate that the light-oriented movement of Adiantum chloroplasts is caused by red and blue light, mediated by phytochrome and another, unidentified photoreceptor(s), respectively. This movement depends on a local gradient of the far-red-absorbing form of phytochrome or of a photoexcited blue-light photoreceptor, and it includes positive and negative responses for both red and blue light.Abbreviations BL blue light - FR far-red light - Pfr far-red-absorbing form of phytochrome - Pr red-absorbing form of phytochrome - R red light - UV ultraviolet     

Dynamic changes in the organization of microfilaments associated with the photocontrolled motility of chloroplasts in epidermal cells ofVattisneria

Summary Using time-lapse video microscopy, we performed a semiquantitative investigation of the movement of chloroplasts on the cytoplasmic layer that faces the outer periclinal wall (P side) of epidermal cells of leaves of the aquatic angiospermVallisneria gigantea Graebner. Under continuous irradiation with red light (650 nm, 0.41 W/m²), the movement of chloroplasts on the P side was transiently accelerated within 5 min. The increased movement began to decrease at around 20 min and fell below the original level after 40 to 60 min of irradiation with red light. The acceleration and deceleration of movement of chloroplasts on the P side seemed to lead directly to the increase and the subsequent decrease in the rate of migration of chloroplasts from the P side to the anticlinal layers of cytoplasm, which are responsible for the accumulation of chloroplasts on the P side, as we demonstrated previously. In the presence of inhibitors of photosynthesis, the accelerated movement of chloroplasts was maintained for as long as the chloroplasts were irradiated with red light. The rapid acceleration and deceleration of the movement of chloroplasts could be observed repeatedly with sequential irradiation with red and then far-red light (746 nm, 0.14 W/m²). Concomitantly with the loss of motility of chloroplasts on the P side, a dynamic change in the configuration of microfilaments, from a network to a honeycomb, occurred on the P side.Abbreviations APW artificial pond water - A side cytoplasmic layer that faces the anticlinal wall - ATP adenosine triphosphate - DCMU 3-(3,4-dichlorophenyl)-1,1-dimethylurea - F-actin fibrous actin - FITC fluorescein isothiocyanate - PBS phosphate-buffered saline - Pfr farred-light-absorbing form of phytochrome - Pr red-light-absorbing form of phytochrome - P side cytoplasmic layer that faces the outer periclinal wall Dedicated to Professor Eldon H. Newcomb in recognition of his contributions to cell biology     

Effects of prolonged omnilateral gravistimulation on the ultrastructure of statocytes and on the graviresponse of roots

Statocytes of vertically growing roots of Lepidium sativum L. exhibit a strict polarity: The nucleus is positioned near the proximal periclinal cell wall, amyloplasts are sedimented on a complex of rough endoplasmic reticulum (ER) consisting of parallel cisternae near the distal periclinal cell wall.When 24 h old, vertically grown roots are rotated for an additional 20 h on a horizontal clinostat, this polarity is destroyed. Furthermore, the prolonged omnilateral stimulation leads to a damage of the statocytes, which in some cases ends in the self-destruction of the sensitive cells. The different components of the ultrastructural respones of the statocytes are: Displacement of the nucleus; changes in amount and distribution of the ER; loss of amyloplast starch; confluence of lipid droplets to large aggregates: a considerable increase of the lytic compartment. In addition, even anticlinal cell walls may be lysed up to small stumps. As all these effects are clearly restricted to the statocytes, only these cells are able to respond to the continuously changing direction of the gravity vector, thus perceiving gravity as such.After being exposed horizontally, the graviresponse of rotated roots is delayed as compared to the controls. About 20% of the rotated roots do not respond (curve) at all, but grow perpendicular in relation to the gravity vector. Perception of gravity is inevitably correlated with the polarity and the integrity of the statocytes.Abbreviation ER endoplasmic reticulum A preliminary report was presented at the Fall Meeting of the German Society for Cell Biology in Salzburg, Austria, September 1979 (Hensel and Sievers 1979)This paper represents part of a dissertation (D 5) of W. H.     

Blue light-induced chloroplast relocation in Arabidopsis thaliana as analyzed by microbeam irradiation

Chloroplast relocation in mesophyll cells of Arabidopsis thaliana was observed microscopically and analyzed by microbeam irradiation. Chloroplasts located along the anticlinal walls in dark-adapted cells. When part of a cell was irradiated with a microbeam of high fluence rate blue light (B) simultaneously with background red light (R) on the whole cell, the chloroplasts moved towards the B-irradiated area, but did not enter the beam. The background R illumination activated cytoplasmic motility as well as chloroplast movement. Without R illumination, there was little chloroplast relocation. In light-adapted cells in which the chloroplasts were spread over the cell surface perpendicular to the incident light, R-illumination had the same effect. Under background R, the chloroplasts moved out of the area irradiated with a B microbeam of 8 or 30 W m(-2) (avoidance response), but chloroplasts outside the beam moved towards the area irradiated with the B microbeam (accumulation response). These results suggest that the signals for accumulation and avoidance responses were generated in a single cell by high fluence rate B. cry1cry2, npq1 and nph1 mutants showed B-induced chloroplast relocation. Both the accumulation and avoidance responses were observed in all the mutants, although in the nph1 mutant, the sensitivity of accumulation movement was slightly lower than that of the wild type. We discuss the possible photoreceptor for B-induced chloroplast relocation.     

Microtubules and morphogenesis in stomata of the water fernAzolla: An unusual mode of guard cell and pore development

Summary A mature stomate of the water fernAzolla consists of a single apparently unspecialized annular guard cell (GC) with two nuclei surrounding an elongated pore aligned longitudinally in the leaf. During development, the guard mother cell develops a preprophase band (PPB) of microtubules (MTs) oriented transverse to the leaf axis. This is followed by a cell plate which fuses with the parental walls at the PPB site. Subsequently only the central part of the cell plate is consolidated, while the parts to either side become perforated and tenuous and may disperse completely, forming a single composite GC.Meanwhile, a dense array of MTs appears along both faces of the central part of the new wall, oriented normal to the leaf surface. Further MT arrays radiate out across the periclinal walls from the region of the consolidated cell plate. Putative MT nucleating sites are seen along the cell edges between these anticlinal and periclinal arrays. Polarized light microscopy reveals cellulose deposition parallel to the periclinal MT arrays. At the same time lamellar material is deposited within the new anticlinal wall. As the GC complex elongates, a split appears in these lamellae creating an initially transverse slit which then opens up to become first circular and ultimately an elongated pore aligned in the long axis of the leaf,i.e., at right angles to the wall in which it originated. The radiating pattern of cellulose microfibrils in the periclinal walls contributes to the shaping of the pore. Elongation at the apical and basal ends of the GC is restricted by longitudinal microfibril orientation, while that at the sides is facilitated by transverse alignment.     

Plastid movement impaired 2, a new gene involved in normal blue-light-induced chloroplast movements in Arabidopsis

Chloroplasts move in a light-dependent manner that can modulate the photosynthetic potential of plant cells. Identification of genes required for light-induced chloroplast movement is beginning to define the molecular machinery that controls these movements. In this work, we describe plastid movement impaired 2 (pmi2), a mutant in Arabidopsis (Arabidopsis thaliana) that displays attenuated chloroplast movements under intermediate and high light intensities while maintaining a normal movement response under low light intensities. In wild-type plants, fluence rates below 20 micromol m(-2) s(-1) of blue light lead to chloroplast accumulation on the periclinal cell walls, whereas light intensities over 20 micromol m(-2) s(-1) caused chloroplasts to move toward the anticlinal cell walls (avoidance response). However, at light intensities below 75 micromol m(-2) s(-1), chloroplasts in pmi2 leaves move to the periclinal walls; 100 micromol m(-2) s(-1) of blue light is required for chloroplasts in pmi2 to move to the anticlinal cell walls, indicating a shift in the light threshold for the avoidance response in the mutant. The pmi2 mutation has been mapped to a gene that encodes a protein of unknown function with a large coiled-coil domain in the N terminus and a putative P loop. PMI2 shares sequence and structural similarity with PMI15, another unknown protein in Arabidopsis that, when mutated, causes a defect in chloroplast avoidance under high-light intensities.     

Phytochrome-mediated branch formation in protonemata of the mossCeratodon purpureus

We have analyzed light induction of side-branch formation and chloroplast re-arrangement in protonemata of the mossCeratodon purpureus. After 12 hr of dark adaptation, the rate of branch formation was as low as 5%. A red light treatment induced formation of side branches up to 75% of the dark-adapted protonema. The frequency of light induced branch formation differed between cells of different ages, the highest frequency being found in the 5th cell, the most distal cell studied from the apex. We examined the effect of polarized light given parallel to the direction of filament growth. The position of branching within the cell depended on the vibration plane of polarized red light. Branch formation was highest when the electric vector of polarized light vibrates parallel to the cell surface and is fluence rate dependent. The positional effect of polarized red light could be nullified to some extent by simultaneous irradiation with polarized far-red light. An aphototropic mutant,ptr116, shows characteristics of deficiency in biosynthesis of the phytochrome chromophore and exhibits no red-light induced branch formation. Biliverdin, a precursor of the phytochrome chromophore, rescued the red-light induced branching when added to the medium, supporting the conclusion that phytochrome acts as photoreceptor for red light induced branch formation. The light effect on chloroplast re-arrangement was also analyzed in this study. We found that polarized blue light induced chloroplast re-arrangement in wild-type cells, whereas polarized red light was inactive. This result suggests that chloroplast re-arrangement is only controlled by a blue light photoreceptor, not by phytochrome inCeratodon.     

Phytochrome-Mediated Photoorientation of Chloroplasts in Protonemal Cells of the Fern Adiantum Can be Induced by Brief Irradiation with Red Light

Measuring the ratio of the number of photooriented chloroplaststo the total number of chloroplasts, we found that photoorientationof chloroplasts in protonemata of the fern Adiantum capillus-veneriscould be induced by brief irradiation with polarized red light.After irradiation with red light (R) of 3 or 10 min, orientationalmovement was detected as early as 10 min after the irradiation;it continued during the subsequent dark period for 30–60min, after which chloroplasts gradually dispersed again. WhenR-treated protonemata were irradiated briefly with a second10-min pulse of R, 60 min after the onset of the first irradiation,the orientational response of chloroplasts was again observed.Typical red/far-red photoreversibility was apparent in the response,indicating the involvement of phytochrome. By contrast, irradiationwith polarized blue light for 10 min was ineffective, whileirradiation with blue light (B) at the same fluence for a longerperiod of time clearly induced the photoorientation of chloroplasts.It is likely that longterm irradiation is necessary for theresponse mediated by a blue-light receptor. When protonemata were irradiated with far-red light (FR) immediatelyafter R or after a subsequent dark period of 10 min, the magnitudeof the orientational response was smaller and chloroplasts dispersedmore quickly than those exposed to R alone. When FR was appliedat 50 min, when the response to R had reached the maximum level,chloroplasts again dispersed rapidly to their dark positions.These results indicate that P_FR not only induces the photoorientationmovement of chloroplasts but also fixes the chloroplasts atthe sites to which they have moved as a result of photoorientation. (Received June 2, 1993; Accepted January 11, 1994)     

Development of endosperm in Arabidopsis thaliana

 The process of endosperm development in Arabidopsis was studied using immunohistochemistry of tubulin/microtubules coupled with light and confocal laser scanning microscopy. Arabidopsis undergoes the nuclear type of development in which the primary endosperm nucleus resulting from double fertilization divides repeatedly without cytokinesis resulting in a syncytium lining the central cell. Development occurs as waves originating in the micropylar chamber and moving through the central chamber toward the chalazal tip. Prior to cellularization, the syncytium is organized into nuclear cytoplasmic domains (NCDs) defined by nuclear-based radial systems of microtubules. The NCDs become polarized in axes perpendicular to the central cell wall, and anticlinal walls deposited among adjacent NCDs compartmentalize the syncytium into open-ended alveoli overtopped by a crown of syncytial cytoplasm. Continued centripetal growth of the anticlinal walls is guided by adventitious phragmoplasts that form at interfaces of microtubules emanating from adjacent interphase nuclei. Polarity of the elongating alveoli is reflected in a subsequent wave of periclinal divisions that cuts off a peripheral layer of cells and displaces the alveoli centripetally into the central vacuole. This pattern of development via alveolation appears to be highly conserved; it is characteristic of nuclear endosperm development in angiosperms and is similar to ancient patterns of gametophyte development in gymnosperms. Received: 21 September 1998 / Revision accepted: 17 November 1998