髙橋 秀幸

Background and Aims Cucumber (Cucumis sativus) seedlings develop a peg on the concave side of the gravitropically bending transition zone between the hypocotyl and the root after seed germination. Peg initiation occurs in response to auxin when its levels in the concave side of the transition zone exceed a particular threshold through the graviresponse. Ethylene also plays an important role in peg formation, but its relationship to auxin in this event is not understood. Here, the role ethylene plays in auxin-induced peg formation is studied. Methods Peg formation of cucumber seedlings exposed to ethylene at different stages of growth or during exogenous auxin treatment was observed. In addition, ethylene evolution from the concave and convex sides of the transition zone was compared and their transcription of CS-ACS (1-aminocyclopropane-1-carboxylic acid synthase) genes was analysed by RT-PCR and in situ hybridization. Key Results Seedlings treated with ethylene after peg initiation produced an enlarged peg, whereas ethylene treatment before peg initiation inhibited peg formation. Ethylene also promoted the development of the peg in the auxin-treated seedlings. Furthermore, the concave side of the transition zone at peg initiation produced more ethylene and CS-ACS1 mRNA than the convex side. Conclusions Since CS-ACS1 is an auxin-inducible gene, the greater abundance of auxin in the concave side of the transition zone causes peg initiation and increases CS-ACS1-mediated ethylene biosynthesis, which then facilitates peg development. (C) 2004 Annals of Botany Company.

Background and Aims Cucumber (Cucumis sativus) seedlings develop a peg on the concave side of the gravitropically bending transition zone between the hypocotyl and the root after seed germination. Peg initiation occurs in response to auxin when its levels in the concave side of the transition zone exceed a particular threshold through the graviresponse. Ethylene also plays an important role in peg formation, but its relationship to auxin in this event is not understood. Here, the role ethylene plays in auxin-induced peg formation is studied. Methods Peg formation of cucumber seedlings exposed to ethylene at different stages of growth or during exogenous auxin treatment was observed. In addition, ethylene evolution from the concave and convex sides of the transition zone was compared and their transcription of CS-ACS (1-aminocyclopropane-1-carboxylic acid synthase) genes was analysed by RT-PCR and in situ hybridization. Key Results Seedlings treated with ethylene after peg initiation produced an enlarged peg, whereas ethylene treatment before peg initiation inhibited peg formation. Ethylene also promoted the development of the peg in the auxin-treated seedlings. Furthermore, the concave side of the transition zone at peg initiation produced more ethylene and CS-ACS1 mRNA than the convex side. Conclusions Since CS-ACS1 is an auxin-inducible gene, the greater abundance of auxin in the concave side of the transition zone causes peg initiation and increases CS-ACS1-mediated ethylene biosynthesis, which then facilitates peg development. (C) 2004 Annals of Botany Company.

ISI 被引用数：4

In order to understand the microspore and pollen development, recently, we have isolated a number of anther-specific genes in the model legume, Lotus japonicus. From these anther-specific genes, we identified one novel microspore-specific gene, LjImfb-c82. In order to determine the molecular characterization of LjImfb-c82, full-length cDNA clone was first isolated and sequenced. It encoded a protein of 286 amino acids (LjHIR1), which had sequence similarity to Hypersensitive-Induced Response like protein. LjHIR1 was specifically expressed in microspore on the in situ hybridization experiment. From the sequence similarity to prohibitin-domain protein, the LjHIR1 might be related to ion channel regulation in microspore development.

The complicated genetic pathway regulates the developmental programs of male reproductive organ, anther tissues. To understand these molecular mechanisms, we performed cDNA microarray analyses and in situ hybridization to monitor gene expression patterns during anther development in rice. Microarray analysis of 4,304 cDNA clones revealed that the hybridization signal of 396 cDNA clones (271 non-redundant groups) increased more than six-fold in every stage of the anthers compared with that of leaves. Cluster analysis with the expression data showed that 259 cDNA clones (156 non redundant groups) were specifically or predominantly expressed in anther tissues and were regulated by developmental stage-specific manners in the anther tissues. These co-regulated genes would be important for development of functional anther tissues. Furthermore, we selected several clones for RNA in situ hybridization analysis. From these analyses, we found several novel genes that show temporal and spatial expression patterns during anther development in addition to anther-specific genes reported so far. These results indicate that the genes identified in this experiment are controlled by different programs and are specialized in their developmental and cell types.

Cucumber seedlings show positive gravitropism and bend in the transition zone between the hypocotyl and the root. The peg, a specialized protuberance, develops on the concave side of the bending transition zone. Auxin and the mRNA of an auxin-inducible gene (CslAA1) isolated from cucumber are differentially accumulated across the transition zone during the gravity-regulated peg formation. In this study, five cDNAs of Auxin Response Factors (ARFs) from cucumber were isolated and their mRNA accumulation was compared with that of CslAA1. The tissue specificity of CsARF2 mRNA accumulation was similar to that of CslAA1. Because the structural character of CsARF2 predicts that it is a transcriptional activator, CsARF2 may be involved in the activation of CslAA1 transcription, which plays a role in gravity-regulated peg formation. Neither gravity nor auxin affected mRNA accumulation of five CsARFs including CsARF2, suggesting that CsARF2 may be regulated at a post-transcriptional level to induce the asymmetric expression of the CslAA1 gene in response to gravistimulation and auxin in cucumber seedlings.

In the present study, root hydrotropism in an agravitropic mutant of Pisum sativum L. grown in vermiculite with a steep water potential gradient was examined. When wet and dry vermiculite were placed side by side, water diffused from the wet (-0.04 MPa) to the dry (-1.2 MPa) and a steep water potential gradient became apparent in the dry vermiculite close to the boundary between the two. The extent and location of the gradient remained stable between the fourth and sixth day after filling a box with vermiculite, and the steepest gradient (approx. 0.02 MPa mm(-1)) was found in the initially dry vermiculite between 60 and 80 mm from the boundary. When seedlings with 25-35 mm long roots were planted in the initially dry vermiculite near where the gradient had been established, each of the main roots elongated toward the wet vermiculite, i.e. toward the high water potential. Control roots elongated without curvature in both the wet and the dry vermiculite, in which no water potential gradient was detectable. These results show that pea roots respond to the water potential gradient around them and elongate towards the higher water potential. Therefore, positive hydrotropism occurs in vermiculite just as it does in air. Hydrotropism in soil may be significant when a steep water potential gradient is apparent, such as when drip irrigation is applied. (C) 2003 Annals of Botany Company.

Background: The RING-H2 finger protein RBX1 (ROC1/HRT1) is a common subunit of SKP1-CDC53/CUL1-F-box (SCF), other cullins and von Hippel-Lindau (VHL) tumour suppressor E3 ubiquitin ligase complexes. RBX1 protein sequences are highly conserved in various species, including yeasts, Drosophila melanogaster, mice and humans. In Saccharomyces cerevisiae, RBX1 is essential for the G1/S transition. Results: Caenorhabditis elegans RBX1 is strongly expressed in early embryos and in the gonad, including meiotic cells. Depletion of RBX1 by RNA-mediated interference (RNAi) caused pronounced defects in the first meiotic division. Several irregular phenotypes were identified in embryos that escaped from meiotic arrest: defects in mitotic chromosomal condensation and segregation, abnormal chromosome bridges, giant nuclei, abnormal cortical protrusion, multinucleate cells and defects in germ cell proliferation. Moreover, histone H3 phosphorylation at Ser(10) and Ser(28) was significantly reduced in these embryos. The histone H3 phosphorylation defect of embryos was rescued by the additional depletion of protein phosphatase 1 (GLC7alpha/beta) by RNAi. Conclusion: These results indicate that the RBX1 protein participates in diverse functions relevant to chromosome metabolism and cell cycle control.

Cucurbit seedlings potentially develop a peg on each side of the transition zone between the hypocotyl and root. Seedlings grown in a horizontal position suppress the development of the peg on the upper side of the transition zone in response to gravity. It is suggested that this suppression occurs due to a reduction in auxin levels to below the threshold value. We show in this study that the free indole-3-acetic acid (IAA) content is low, while IAA conjugates are significantly more abundant in the upper side of the transition zone of gravistimulated seedlings, compared to the lower side. A transient increase in mRNA of the auxin-inducible gene, CS-IAA1, was observed in the excised transition zone. The result suggests that the transition zone is a source of auxin. Cucumber seedlings treated with auxin-transport inhibitors exhibited agravitropic growth and developed a peg on each side of the transition zone. Auxin-transport inhibitors additionally caused an increase in CS-IAA1 mRNA accumulation at the transition zone, indicating a rise in intracellular auxin concentrations due to a block of auxin efflux. To study the involvement of the auxin transport system in peg formation, we isolated the cDNAs of a putative auxin influx carrier, CS-AUX1, and putative efflux carrier, CS-PIN1, from cucumber (Cucumis sativus L.) plants. Both genes (CS-AUX1 in particular) were auxin-inducible. Accumulation of CS-AUX1 and CS-PIN1 mRNAs was observed in vascular tissue, cortex and epidermis of the transition zone. A reduced level of CS-AUX1 mRNA was observed in the upper side of the gravistimulated transition zone, compared with the lower side. It is therefore possible that a balance in the activities of auxin influx and efflux carriers controls intracellular auxin concentration at the transition zone, which results in lateral placement of a peg in cucumber seedlings.

Sex differentiation in cucumber plants (Cucumis sativus L.) appears to be determined by the selective arrest of the stamen or pistil primordia. We investigated the influence of an ethylene-releasing agent (ethephon) or an inhibitor of ethylene biosynthesis (aminoethoxyvinyl glycine) on sex differentiation in different developmental stages of flower buds. These treatments influence sex determination only at the stamen primordia differentiation stage in both monoecious and gynoecious cucumbers. To clarify the relationships between the ethylene-producing tissues and the ethylene-perceiving tissues in inducing female flowers in the cucumber, we examined the localization of mRNA accumulation of both the ACC synthase gene (CS-ACS2) and the ethylene-receptor-related genes (CS-ETR1, CS-ETR2, and CS-ERS) in flower buds by in situ hybridization analysis. CS-ACS2 mRNA was detected in the pistil primordia of gynoecious cucumbers, whereas it was located in the tissues just below the pistil primordia and at the adaxial side of the petals in monoecious cucumbers. In flower buds of andromonoecious cucumbers, only CS-ETR1 mRNA was detected, and was located in the pistil primordia. The localization of the mRNAs of the three ethylene-receptor-related genes in the flower buds of monoecious and gynoecious cucumbers overlap but are not identical. We discuss the relationship between the mRNA accumulation patterns and sex expression in cucumber plants.

The effects of heavy ion particle irradiation on meiosis and reproductive development in the nematode Caenorhabditis elegans were studied. Meiotic pachytene nuclei are significantly resistant to particle irradiation by the heavy ions carbon and argon, as well as to X-rays, but not UV, whereas diplotene to diakinesis stage oocytes and early embryonic cells are not. Chromosomal abnormalities appear in mitotic cells and in maturing oocytes irradiated with heavy ion particles during the diplotene to the early diakinesis stages, but not in oocytes irradiated during the pachytene stage. The pachytene nuclei of ced-3 mutants, which are defective in apoptosis, are similarly resistant to ionizing radiation, but pachytene nuclei depleted for Ce-atl-1 (ataxia-telangiectasia like 1) or Ce-rdh-1/rad-51 are more sensitive. Pachytene nuclei thus appear to effectively repair heavy ion-induced DNA damage by the meiotic homologous recombination system.

The shoots of a Japanese strain of morning glory (Pharbitis nil) called 'Shidare-asagao' display agravitropic and weeping growth. It has been shown that this shoot agravitropism may be due to the defective differentiation of endodermal cells that contain statoliths. Roots of the weeping morning glory show normal responsiveness to gravity and the shoots are positively phototropic. Shoots of the morning glory cultivar Violet used as a wild type exhibited distinct circumnutation with circular movements that increase as the plants grow. In weeping morning glory, however, nutation was limited to slight back and forth or side to side movements. To determine whether endodermal cells participate in circumnutation through a function that is independent of their role in gravitropism, the nutational movements of various gravitropic mutants of Arabidopsis thaliana were compared. The inflorescences of wild-type Arabidopsis showed relatively large circular movements. Inflorescences of the pgm-1 mutant, which is defective in starch synthesis, showed reduced nutation. Even more seriously affected were the sgr1-1/scr-3 and sgr7-1/shr-2 mutants, which are defective in endodermal cell differentiation, and the auxin-resistant axr2-1 mutant showed no significant nutational movements at all. 1-N-naphthylphthalamic acid (NPA) could inhibit Violet circumnutation, supporting the notion that auxin participates in circumnutation. Thus, the gravitropic response is an essential component in plant shoot circumnutation. Endodermal cells are involved in such circumnutation possibly because of their role in inducing the gravitropic response.

In response to a moisture gradient, roots exhibit hydrotropism to control the orientation of their growth. To exhibit hydrotropism, however, they must overcome the gravitropism that is dominant on Earth. We found that moisture gradient or water stress caused immediate degradation of the starch anchors, amyloplasts, in root columella cells of Arabidopsis and radish (Raphanus sativus). Namely, development of hydrotropic response was accompanied by a simultaneous reduction in starch content in columella cells. Rapid degradation of amyloplasts in columella cells also occurred in the water-stressed roots with sorbitol or mannitol. Both hydrotropically stimulated and water-stressed roots showed a reduced responsiveness to gravity. Roots of a starchless mutant, pgm1-1, showed an enhanced hydrotropism compared with that of the wild type. These results suggest that the reduced responsiveness to gravity is, at least in part, attributable to the degradation of amyloplasts in columella cells. Thus, the reduction in gravitropism allows the roots to exhibit hydrotropism.

The Hong Mang Mai wheat cultivar is tolerant to deep-sowing conditions because it has an elongated first internode that is sensitive to gibberellin (GA3). The cells in the GA-treated first internodes were approximately 4.2 mm long, twice as long as the untreated Hong Mang Mai first internode cells. The elongation of the first internode of Hong Mang Mai, particularly when treated with GA3, was accompanied by remarkable spiral growth. In contrast, the first internodes of the GA-insensitive cultivar Norin 10 did not exhibit GA3-induced elongation or spiral growth. The walls of the first internode cells of GA3-treated Hong Mang Mai seedlings showed increased extensibility and higher (1→3), (1→4)-β-D-glucanase activity, autolysis and glucan contents than the cell walls of untreated Hong Mang Mai first internodes. The changes in the cell wall extensibility due to GA3 treatment correlated strongly with the GA3-induced changes in cell wall glucan content, autolysis, and glucanase activity. GA3-treated Hong Mang Mai seedlings showed elevated expression of Glucanase EI gene in the first internode compared to GA3-treated Norin 10. Thus, GA aids first internode elongation in Hong Mang Mai by enhancing glucan turnover and thus increasing cell wall loosening. The spiral growth of the first internode also helps the plant elongate against soil resistance, thereby promoting the deep-sowing tolerance of this cultivar.

Photoperiod and the plant hormone, ethylene, modify sex expression of flowers in cucumber (Cucumis sativus L.). In the present study, femaleness of cucumber occurred under short-day (8 h photoperiod) conditions compared to that under long-day (16 h photoperiod) conditions, although the effect of photoperiod was more pronounced in a monoecious than in an andromonoecious cucumber. Application of ethylene had a greater effect than photoperiod on the production of female and bisexual flowers in monoecious and andromonoecious cucumbers, respectively. Ethylene evolution and the expression of CS-ACS2, CS-ACS4 and CS-ERS genes in the shoot apices of both monoecious and andromonoecious cucumber plants had a diurnal rhythm with a peak in the middle of an 8 h or a 16 h light period. Peak ethylene evolution and expression of CS-ACS2 was greater under short-day conditions than under long-day conditions in a monoecious cucumber but not in an andromonoecious one. Expression of CS-ACS4 in monoecious and andromonoecious cucumber plants did not differ, but the level was higher under short-day conditions compared with that under long-day conditions. Thus, CS-ACS2 and CS-ACS4 might be involved in the basic diurnal rhythm of ethylene evolution in cucumber. Because exogenous ethylene increased the expression of CS-ACS2 and CS-ERS in monoecious cucumber possessing the M locus, but not in andromonoecious cucumber in which the function of the M locus was lost (Yamasaki et al. Plant and Cell Physiology 42, 608-619, 2001), the CS-ACS2 gene might also be involved in ethylene production by positive feedback via regulation of M locus under short-day conditions.

A fundamental study was conducted to develop a facility having an adequate air circulation system for growing healthy plants over a long-term under microgravity conditions in space. To clarify the effects of gravity on heat exchange between plant leaves and the ambient air, surface temperatures of sweet potato and barley leaves and replica leaves made of wet paper and copper were evaluated at gravity levels of 0.01, 1.0, 1.5 and 2.0 g for 20 s each during parabolic aeroplane flights. Thermal images were captured using infrared thermography at an air temperature of 26°C, a relative humidity of 18% and an irradiance of 260 W m-2. Mean leaf temperatures increased by 0.9-1.0°C with decreasing gravity levels from 1.0 to 0.01 g and decreased by 0.5°C with increasing gravity levels from 1.0 to 2.0 g. The increase in leaf temperatures was at most 1.9°C for sweet potato leaves over 20 s as gravity decreased from 1.0 to 0.01 g. The boundary layer conductance to sensible heat exchange decreased by 5% when the gravity decreased from 1.0 to 0.01 g at the air velocity of 0.2 m s-1. The decrease in the boundary layer conductance with decrease in the gravity levels was more significant in a lower air velocity. Heat exchange between leaves and the ambient air was more retarded at lower gravity levels because of less sensible and latent heat transfers with less heat convection.

根は水分屈性を発現する能力を有するが、地球上では重力屈性が水分屈性をマスクする。エンドウやキュウリの根では、したがって、突然変異、クリノスタット、宇宙実験などで重力屈性を消去させることによって水分屈性を発現させることができる。一方、われわれは、最近シロイヌナズナやダイコンの根が水分勾配の存在下で水分屈性を強く発現させるのは、水ストレスがコルメラ細胞中のデンプンを分解し、重力応答性を低下させるためであることを明らかにした。シロイヌナズナの根は、重力屈性や水分屈性だけでなく光屈性をも発現させる。そこで本研究では、シロイヌナズナの根における光屈性と重力屈性の相互作用をコルメラ細胞内デンプン量の変化に注目して解析した。その結果、コルメラ細胞のデンプン量が、光屈性に先立って有意に減少することを見出した。すなわち、根は、白色光および青色光の一方向照射によって負の光屈性を、赤色光の一方向照射によって正の光屈性を示した。この光屈性を誘導する一方向照射によってコルメラ細胞中のデンプン量は減少したが、同軸上双方向照射では光屈性が誘導されず、デンプン量も減少しなかった。これらのことから、シロイヌナズナの根は、水分屈性だけでなく光屈性を発現させる場合にも、コルメラ細胞中のデンプン量を減少させることによって重力感受性を低下させ、重力屈性による干渉を小さくする機構を持つものと考えられる。

根は，水分勾配刺激に応答して水分屈性を発現し，重力屈性等とともにその姿勢を制御している．近年，重力屈性については分子生物学的な解析が行われるようになってきた．しかし，水分屈性の発現機構については，根冠が刺激受容器官であること，オーキシン，アブシジン酸およびカルシウムイオンの関与することなどが知られているものの，その刺激受容やシグナル伝達に関する分子機構はほとんど解明されていない．そこで本研究では，モデル植物のシロイヌナズナを用いた根の水分屈性突然変異体スクリーニング法を開発し，EMS処理をしたM₂種子約2万個から，野生型に比較して水分屈性の低下した突然変異体の単離を試みた．その結果，これまでに12株の水分屈性突然変異体rhy (root hydrotropism) が見出された．それらのrhy突然変異体について水分屈性の経時的な発現，重力屈性，光屈性，波型成長等を解析した結果，水分屈性特異的な突然変異体（rhy1）や，水分屈性を含む複数の屈性に異常のある突然変異体（rhy2，rhy3，rhy4）が見出された．現在，Landsberg erectaと交配したF₂集団のDNAを用いて，変異遺伝子のマッピングを行っている．

During meiotic prophase 1, homologous recombination is accompanied by dynamic chromosomal changes. The Ce-rdh-1/rad-51 gene is the only bacterial recA-like gene in the nematode C. elegans genome. Upon depletion of Ce-rdh-1/rad-51 using the RNA interference method, abnormal 'kinked' chromosomes can be observed in mature oocytes at diakinesis, whereas synapsis between homologous chromosomes during the pachytene stage is normal. Following fertilization, Ce-rdh-1/rad-51-depleted embryos die early in embryogenesis, and their nuclei exhibit abnormal chromosome fragments and bridges. From epistasis analyses with Ce-spo-11 defective mutant and ionizing radiation, it is indicated that Ce-rdh-1/rad-51 functions after double-strand break (DSB) formation of meiotic recombination. Under the Ce-chk-2 defective condition, whose meiotic synapsis and meiotic recombination between homologous chromosomes are completely inhibited, the Ce-rdh-1/rad51 is normally expressed in the gonadal cells. Moreover, it seems that exogenous DSBs in the Ce-chk-2 defective nuclei at the pachytene stage can be repaired between sister chromatids in a Ce-rdh-1/rad-51-dependent manner. These results indicate that Ce-rdh-1/rad51 functions after both endogenous and exogenous DSB formation during meiosis, but not as 'pairing centers' for meiotic synapsis.

We have developed experimental systems to study hydrotropism in seedling roots of Arabidopsis thaliana (L.) Heynh. Arabidopsis roots showed a strong curvature in response to a moisture gradient, established by applying 1% agar and a saturated solution of KCl or K2CO3 in a closed chamber. In this system, the hydrotropic response overcame the gravitropic response. Hydrotropic curvature commenced within 30 min and reached 80-100.dogrees within 24 h of hydrostimulation. When 1 % agar and agar containing 1 MPa sorbitol were placed side-by-side in humid air, a water potential gradient formed at the border between the two media. Although the gradient changed with time, it still elicited a hydrotropic response in Arabidopsis roots. The roots curved away from 0.5-1.5 MPa of sorbitol agar. Various Arabidopsis mutants were tested for their hydrotropic response. Roots of aba1-1 and abi2-1 mutants were less sensitive to hydrotropic stimulation. Addition of abscisic acid restored the normal hydrotropic response in abal-I roots. In comparison, mutants that exhibit a reduced response to gravity and auxin, axr1-3 and axr2-1, showed a hydrotropic response greater than that of the wild type. Wavy mutants, wav2-1 and wav3-11 showed increased sensitivity to the induction of hydrotropism by the moisture gradient. These results suggest that auxin plays divergent roles in hydrotropism and gravitropism, and that abscisic acid plays a positive role in hydrotropism. Furthermore, hydrotropism and the wavy response may share part of a common molecular pathway controlling the directional growth of roots.

We have developed experimental systems to study hydrotropism in seedling roots of Arabidopsis thaliana (L.) Heynh. Arabidopsis roots showed a strong curvature in response to a moisture gradient, established by applying 1% agar and a saturated solution of KCl or K2CO3 in a closed chamber. In this system, the hydrotropic response overcame the gravitropic response. Hydrotropic curvature commenced within 30 min and reached 80-100.dogrees within 24 h of hydrostimulation. When 1 % agar and agar containing 1 MPa sorbitol were placed side-by-side in humid air, a water potential gradient formed at the border between the two media. Although the gradient changed with time, it still elicited a hydrotropic response in Arabidopsis roots. The roots curved away from 0.5-1.5 MPa of sorbitol agar. Various Arabidopsis mutants were tested for their hydrotropic response. Roots of aba1-1 and abi2-1 mutants were less sensitive to hydrotropic stimulation. Addition of abscisic acid restored the normal hydrotropic response in abal-I roots. In comparison, mutants that exhibit a reduced response to gravity and auxin, axr1-3 and axr2-1, showed a hydrotropic response greater than that of the wild type. Wavy mutants, wav2-1 and wav3-11 showed increased sensitivity to the induction of hydrotropism by the moisture gradient. These results suggest that auxin plays divergent roles in hydrotropism and gravitropism, and that abscisic acid plays a positive role in hydrotropism. Furthermore, hydrotropism and the wavy response may share part of a common molecular pathway controlling the directional growth of roots.

Primary roots of cucumber seedlings showed positive hydrotropism when exposed to a moisture gradient and rotated on a two-axis clinostat. To examine the role of auxin in the differential growth of the hydrotropically responding roots, we first examined the expression of auxin-inducible genes, CS-AUX/IAAs, in cucumber roots. After auxin starvation, mRNA levels of CS-IAA1 and CS-IAA3 decreased in the roots. Applying auxin to the auxin-starved roots resulted in accumulation of CS-1AA1 and CS-IAA3 mRNA. The level of expression of these genes increased when the auxin concentration was increased. CS-IAA1 mRNA accumulated in response to 10(-8) M auxin, and the level increased further, depending on the dose. Auxin starvation did not result in a decrease in the level of CS-IAA2 mRNA; however, adding exogenous auxin at concentrations higher than 10(-7) M increased its accumulation. In the primary roots responding hydrotropically or gravitropically, CS-IAA1 expression was greater on the concave side of the curving roots than on the convex side. The difference could be detected 30 min following stimulation by gravity or a moisture gradient, and that difference increased with time. These results support the idea that asymmetry of localization of auxin is associated with differential growth in hydrotropically responding roots.

Seedlings of Hong Mang Mai wheat (Triticum aestivum L.) emerge from much deeper in the soil than do other cultivars by elongating their first internode. We examined the roles of ethylene, gibberellins (GA) and potassium in the elongation of the first internode in wheat cultivars. Although ethylene stimulated the elongation of the first internode of Hong Mang Mai in the dark, the ethylene evolution and the expression level of ethylene-receptor-like gene of this cultivar were not greater than those of other cultivars. In Hong Mang Mai, gibberellin A(3) (GA(3)) substantially stimulated the elongation of the first internode in the: dark. Maximum elongation of the first internode was obtained by GA(3) at the concentrations of 2.89 X 10(-6) to 2.89 X 10(-7) M. This cultivar responded markedly to gibberellins A(1) and A(3) by elongating the first internode in the dark. A mixture of gibberellins A(4) and A(7) (GA(4+7)) was not effective in inducing the elongation. Haruhikari, a cultivar having no GA -insensitive-reducing-height (Rht) genes, substantially responded to GA(3), but other cultivars did not. The differences in the sensitivity to GA could account, at least in part, for the varietal differences in the elongation of the first internode in wheat. Takune-komugi, a cultivar whose first internode elongates in response to ethylene, did not respond to GA(3). Thus, there are different mechanisms for the GA- and ethylene-stimulated elongation of the first internode in wheat. Also, it was suggested that potassium uptake from the soil might function together with endogenous GA in inducing the elongation of the first internode in Hong Mang Mai wheat.

The water potential and hydraulic conductivity (Lp) of elongating cells in hydrotropically bending roots of the age-otropic mutant ageotropum of pea (Pisum sativum L.) were measured in situ. When agar blocks with water potentials of -0.03 and -0.8 MPa were unilaterally applied directly to a root tip, cells in the most rapidly elongating zone, 3-4 mm from the tip, showed marked differential growth. The rate of water uptake by a cell on the side treated with an agar block with a lower water potential was significantly larger in the outer first and second layers of cortex than on the other side. There were no differences in the values of turgor pressure, osmotic potential and calculated water potentiaI between the two sides either in elongating or in mature cells, indicating the absence of any difference in the growth-induced water potential on the two sides of the root. Lp was significantly larger on the side with the agar block with lower water potential. The results suggest that the difference in the rate of water uptake during the differential cell growth that occurs during root hydrotropism might be induced mainly by a change in Lp.

Sex determination in cucumber (Cucumis sativus ) plants is genetically controlled by the F and M loci. The F and M loci interact to produce three different sex phenotypes: gynoecious (M-F-), monoecious (M-ff), and andromonoecious (mmff). We found that the levels of ethylene production and the accumulation of CS-ACS2 mRNA in andromonoecious cucumber plants did not differ from those in monoecious plants and were less than the levels measured in gynoecious plants. Ethylene inhibited stamen development in monoecious cucumber but not in andromonoecious one. To investigate the action mechanism of ethylene in the induction of femaleness of cucumber flowers, we isolated three ethylene-receptor-related genes, CS-ETR1, CS-ETR2 and CS-ERS. Ethylene caused substantial increases in the accumulation of CS-ETR2, CS-ERS, and CS-ACS2 mRNA in monoecious cucumber plants, but not in andromonoecious plants. These results suggest that ethylene responses in andromonoecious cucumber plants are reduced compared to those in monoecious plants. This is the first evidence that ethylene signals may mediate the product of the M locus to inhibit stamen development in cucumber. The andromonoecious line provides novel material to study the function of the M locus during sex determination of flowers in cucumber plants.

Cucumber seedlings grown in microgravity developed a peg on each side of the transition zone (TR) between hypocotyl and root, whereas seedlings grown in a horizontal position on the ground developed a peg on the concave side of the gravitropically bending TR. The morphological features of the space-grown seedlings were similar to that of seedlings grown in a vertical position on the ground with their radicles pointing down; both became two-pegged seedlings. Morphogenesis of cucumber seedlings is thus negatively controlled by gravity. Analysis by in situ hybridization of an auxin-inducible gene, CS-IAA1, showed that its mRNA accumulated much more on the lower side of the TR in the horizontally-placed seedlings on the ground just prior to and at the initiation period of peg formation. On the other hand, when seedlings were grown in microgravity or in a vertical position on the ground, accumulation of CS-IAA1 mRNA was seen all around the transition zone. It appeared that the accumulation of CS-IAA1 mRNA decreased on the upper side of the transition. zone upon gravistimulation. Application of exogenous IAA to the seedlings in a horizontal position caused the development of a peg on each side of the transition zone, or a collar-like protuberance, depending on the concentration used. These results suggest that there is a threshold level of auxin concentration for peg formation in cucumber seedlings. Upon gravistimulation, auxin concentration on the upper side of the horizontally placed transition zone is possibly reduced to the level below the threshold value.

Sex determination in cucumber (Cucumis sativus L,) plants is genetically controlled by the F and M loci. These loci interact to produce three different sexual phenotypes: gynoecious (M-F-), monoecious (M-ff), and andromonoecious (mmff). Gynoecious cucumber plants produce more ethylene than do monoecious plants, We found that the levels of ethylene production and the accumulation of CS-ACS2 mRNA in andromonoecious cucumber plants did not differ from those in monoecious plants and were lower than the levels measured in gynoecious plants, Ethylene inhibited stamen development in gynoecious cucumbers but not in andromonoecious ones. Furthermore, ethylene caused substantial increases in the accumulation of CS-ETR2, CS-ERS, and CS-ACS2 mRNA in monoecious and gynoecious cucumber plants, but not in andromonoecious one. In addition, the inhibitory effect of ethylene on hypocotyl elongation in andromonoecious cucumber plants was less than that in monoecious and gynoecious plants. These results suggest that ethylene responses in andromonoecious cucumber plants are reduced from those in monoecious and gynoecious plants. This is the first evidence that ethylene signals may influence the product of the M locus and thus inhibit stamen development in cucumber. The andromonoecious line provides novel material for studying the function of the M locus during sex determination in flowering cucumbers.

The addition of gibberellin A(3) (GA(3)) to culture media induced a dramatic elongation of the first internode in Hong Mang Mai, a wheat variety tolerant to deep-seeding conditions. The length of the first internode in the GA(3)-treated seedlings reached approximately 45 cm, which is twice as long as that of the control, whereas first internodes of other varieties were 3 to 12 cm long and hardly affected by GA(3), The level of endogenous gibberellins (GA) in Hong Mang Mai was not greater than that of other wheat varieties. The expression of GAMyb, a transcription factor, was abundant in the first internode and substantially increased by GA(3) application in Hong Mang Mai, compared with other wheat varieties. These results suggest that the first internode of Hong Mang Mai is more sensitive to GA in inducing strong elongation. The presence of potassium in the culture media was indispensable for the first internode elongation. Application of GA(3) enhanced the uptake of potassium in Hong Mang Mai, Higher sensitivity of the first internode to GA could cause strong elongation by increasing the amount of osmotic solute, which plays an important role in the tolerance mechanism of Hong Mang Mai to deep-seeding conditions.

To clarify the effects of gravity on heat/gas exchange between plant leaves and the ambient air, the leaf temperatures and net photosynthetic rates of plant leaves were evaluated at 0.01, 1.0, 1.5 and 2.0 G of 20 seconds each during a parabolic airplane flight. Thermal images of leaves were captured using infrared thermography at an air temperature of 26degreesC, a relative humidity of 15% and an irradiance of 260 W m(-2). The net photosynthetic rates were determined by using a chamber method with an infrared gas analyzer at an air temperature of 20degreesC, a relative humidity of 50% and a photosynthetic photon flux of 0.5 mmol m(-2) s(-1). The mean leaf temperature increased by 1degreesC and the net photosynthetic rate decreased by 13% with decreasing gravity levels from 1.0 to 0.01 G. The leaf temperature decreased by 0.5degreesC and the net photosynthetic rate increased by 7% with increasing gravity levels from 1.0 to 2.0 G. Heat/gas exchanges between leaves and the ambient air were more retarded at lower gravity levels. A restricted free air convection under microgravity conditions in space would limit plant growth by retarding heat and gas exchanges between leaves and the ambient air. (C) 2001 COSPAR. Published by Elsevier Science Ltd. All rights reserved.

Researchers from 5 Japanese universities have developed a plant growth facility (Space Plant Box) for seed to seed experiments under microgravity. The breadboard model of the Space Plant Box was fabricated by assembling subsystems developed for microgravity. The subsystems include air conditioning and water recycle system, air circulation system, water and nutrient delivery system, lighting system and plant monitoring system. The air conditioning and water recycle system is simply composed of a single heat exchanger, two fans and hydrophilic fibrous strings. The strings allow water movement from the cooler fin in the Cooling Box to root supporting materials in the Plant Growth Chamber driven by water potential deficit. Relative humidity in the Plant Growth Chamber can be changed over a wide range by controlling the ratio of latent heat exchange to sensible heat exchange on the cooling fin of the heat exchanger. The transpiration rate was successfully measured by circulating air inside the Plant Growth Chamber only. Most water was recycled and a small amount of water needed to be added from the outside. The simple, air conditioning and water recycle system for the Space Plant Box showed good performance through a barley (Hordeum vulgare L.) growth experiment. (C) 2001 COSPAR. Published by Elsevier Science Ltd. All rights reserved.

The development of the inflorescence, microspores and anthesis were well synchronized among individuals or in the panicles of barley under controlled environmental conditions. To study the effects of high-temperature stress on the development of pollen mother cells (PMCs) and microspores, the plants were subjected to high temperature treatment at different stages of reproductive growth. When plants were exposed to high temperature for five days at the early differentiation stage of the panicle, pollen grains had apparently normal exine but no or little cytoplasm, At the pre-meiotic stage of PMCs, high temperature caused subsequent development of short anthers possessing no pollen grains. When plants were exposed to high temperature during meiosis of PMCs, all pollen grains possessed exine and were swollen but showed little starch accumulation. In these plants treated at high temperature, the panicles at the heading stage had a normal appearance, but their seeds were virtually sterile. These results indicated that there are at least three stages of reproductive growth hypersensitive to high temperature, which resulted in abnormal terminal phenotypes different from one another.

A Chk2-like gene was identified in the genome of Caenorhabditis elegans. The putative gene product, termed Ce-chk-2 consists of 450 amino acid residues, and shows good homology with the Chk2/Cds1 gene family. The results of RNA-mediated interference (RNAi) indicated that the F1 generation from dsRNA injected animals grew to adulthood, but approximately 95% of their eggs (F2) died during early embryogenesis, Among the few surviving progeny, males (XO animals) arose at an abnormally high frequency (30%). In addition, 12 univalents were observed in full grown oocytes of the F1, while six bivalents were normally observed in mild-type oocytes, Ce-chk-2 gene expression increased in the adult stage, and their expression level decreased in the glp-4 mutant, which is defective in germ line proliferation. The radiation sensitivity of F1 embryos carrying Ce-chk-2 RNAi was not significantly affected. (C) 2000 Federation of European Biochemical Societies. Published by Elsevier Science B.V. All rights reserved.

Sensitivity of meiotic cells to DNA damaging agents is little understood. We have demonstrated that the meiotic-pachytene nuclei in the Caenorhabditis elegans gonad are hyper-resistant to X-ray irradiation, but not to UV irradiation, whereas the early embryonic cells after fertilization and the full grown oocytes are not. The Ce-rdh-1 gene [RAD51, DMC1 (LIM15), homolog 1 or Ce-rad-51], which is essential for the meiotic recombination, is the only bacterial recA-like gene in the nematode genome, and is strongly expressed in the meiotic cells. Following silencing of the Ce-rdh-1 gene by RNA interference, the meiotic cells become more sensitive to X-ray irradiation than the early embryonic cells. This is the first report that meiotic cells are hyper-resistant to DNA strand breaks due to the high level of expression of the enzyme(s) involved in meiotic homologous recombination.

Cucumber (Cucumis sativus L.) seedlings grown in microgravity developed a peg on each side of the transition zone between hypocotyl and root, whereas seedlings grown in a horizontal position on the ground developed a peg on the concave side of the gravitropically bending transition zone. The morphological features of the space-grown seedlings were similar to those of seedlings grown in a vertical position on the ground with their radicles pointing down: both became two-pegged seedlings. Morphogenesis of cucumber seedlings is thus inhibited by gravity. Analysis by in-situ hybridization of an auxin-inducible gene, CS-IAA1, showed that its mRNA accumulated to a much greater extent on the lower side of the transition zone in the horizontally placed seedlings on the ground just prior to and during the initiation period of peg formation. On the other hand, when seedlings were grown in microgravity or in a vertical position on the ground, accumulation of CS-IAA1 mRNA occurred all around the transition zone. Accumulation of CS-IAA1 mRNA in horizontally grown seedlings appreciably decreased on the upper side of the transition zone and increased on the lower side upon gravistimulation, compared with the two-pegged seedlings. Application of IAA to seedlings in a horizontal position caused the development of a peg on each side of the transition zone, or a collar-like protuberance, depending on the concentration used. These results suggest that upon gravistimulation the auxin concentration on the upper side of the horizontally placed transition zone is reduced to a level below the threshold value necessary for peg formation. Space-grown seedlings of cucumber might develop two pegs symmetrically because the auxin level in the entire transition zone is maintained above the threshold. This spaceflight experiment verified for the first time that auxin does not redistribute in microgravity.

An ATM-like gene was identified in the genome of Caenorhabditis elegans. The putative product of the gene, termed Ce-atl-1 (C. elegans ATM-like 1) consists of 2514 amino acid residues. The C-terminal sequence, which contains a PI-3 kinase-like domain, showed good homology with the products of the gene MEC1/ESR1 from budding yeast, the rad3(+) gene of fission yeast and mammalian A TR (ataxia-telangiectasia and rad3(+) related) genes. The results of RNA-mediated interference indicated that the major phenotype associated with repression of Ce-atl-1 was lethality (approximately 50-80%) during early embryogenesis. Among the surviving progeny, males (XO animals) arose at a high frequency (2-30%). In addition, 5% of oocyte chromosomes demonstrated aneuploidy due to a defect in pre-meiotic chromosomal segregation. Gene expression analyses indicated that Ce-atl-1 mRNA was expressed in all larval stages and that its level increased about fivefold in the adult stage. The adult expression level was de creased in the glp-4 mutant, which is defective in germ line proliferation. Ce-atl-1 was strongly expressed in both the mitotic and meiotic cells of adult gonads. In summary, Ce-atl-1 appears to be important for early embryogenesis, and loss of its function results in a defect in chromosome segregation, similar to what has been observed for AT-related proteins.

The effects of gravity on output of sensors with different detection mechanisms under constant humidity were investigated to screen humidity sensors for small plant growth chambers under microgravity. Different levels of gravity were established by parabolic flight using aircraft in which air pressure and temperature were maintained almost constant (the variations were within 0.6kPa and 0.2°C, respectively). Four types of polymer sensors, a thermistor sensor and an electrolytic sensor were used for the experiment. Air with relative humilities of 75% and 58% was prepared using saturated salt solutions of NaCl and NaBr, respectively. Six types of sensors were fixed either in a vessel (1 L) or independently in small vessels (10mL) including the solution absorbedby polymer film. When the sensors were placed in avessel, only the output signal of the thermistor sensor varied with gravity level. It was lowestunder 2 G and highest under low gravity under both relative humidity conditions. This variation was observed in both cases when air in the vessel was and was not stirred. This might be due to a filter protecting the sensor surface to cancel the effect of wind velocity on sensor. Air diffusion inside the filter in both cases seemed to be governed by natural convection of which the strength was affected by gravity level. The same tendency in thermistor sensor output was observed in the small vessels in which each sensor was independently fixed, indicating that an interaction between sensors placed in one vessel is negligible. Thus the change in air diffusion with gravity seemed to be the reason why the output signal of thermistor sensor changed with gravity level. © 2000, The Society of Agricultural Meteorology of Japan. All rights reserved.

It has been reported that ethylene production by cucumber plants is strongly related to the sex expression of their flowers. It has also been shown that both CS-ACS2 gene expression and ethylene evolution are much greater in gynoecious cucumber plants than monoecious ones. To investigate the action mechanism of ethylene in the induction of femaleness of cucumber flowers, we isolated three ethylene-receptor-related genes, CS-ETR1, CS-ETR2 and CS-ERS, from cucumber (Cucumis sativus L.) plants. Of these three genes, CS-ETR2 and CS-ERS mRNA accumulated more substantially in the shoot apices of the gynoecious cucumber than those of the monoecious one. Their expression patterns correlated with the expression of the CS-ACS2 gene and with ethylene evolution in the shoot apices of the two types of cucumber plants. Accumulation of CS-ETR2 and CS-ERS mRNA was significantly elevated by the application of Ethrel, an ethylene-releasing agent, to the shoot apices of monoecious cucumber plants. In contrast, the accumulation of their transcripts was lowered when aminoethoxyvinyl glycine (AVG), an inhibitor of ethylene biosynthesis, was applied to the shoot apices of gynoecious cucumber plants. Thus, the expression of CS-ETR2 and CS-ERS is, at least in part, regulated by ethylene. The greater accumulation of CS-ETR2 and CS-ERS mRNA in gynoecious cucumber plants may be due to the higher level of endogenous ethylene, which plays a role in the development of female flowers.

The plant hormone auxin transcriptionally activates Aux/IAA genes. We have isolated three Aux/IAA cDNA from cucumber, two cDNAs (CS-IAA1 and CS-IAA2) containing the complete open reading frame (ORF), and one partial cDNA (CS-IAA3). Northern blotting analysis showed that Aux/IAA mRNAs were induced during the emergence of radicles from seed coats. After radicle emergence, their mRNAs accumulated in the basal part of the hypocotyl much more than in the apical part, and later in elongating region of hypocotyls. CS-IAA1 and CS-IAA3 mRNA significantly accumulated in response to auxin, although the increment of the former mRNA accumulation by auxin application was much greater than that of the latter. CS-IAA2 did not show an apparent change by auxin treatment in our experiment. In horizontally germinating seedlings, the transition zone between hypocotyl and root curves was due to downward gravitropic growth. On the other hand, vertically germinating seedlings of cucumber do not curve in the early stage of seedling development. The CS-IAA1 mRNA accumulation in horizontally germinating seedlings was more than that in vertically germinating ones during radicle emergence. Furthermore, asymmetric distribution of CS-IAA1 mRNA was detected in the transition zone in in situ hybridization analysis. These results suggest that the CS-IAA1 gene product may be involved in the gravity response during early development of seedlings.

Seedlings of most cucurbitaceous plants develop a peg (protuberance caused by cell outgrowth) on the transition zone between the hypocotyl and root. The peg is necessary for removing the seed coat after germination. In our spaceflight experiments on the STS-95 space shuttle, Discovery, we found that cucumber (Cucumis sativus L.) seedlings grown under microgravity conditions developed two pegs symmetrically at the transition zone. Thus, cucumber seedlings potentially develop two pegs and do not require gravity for peg formation itself, but on the ground the development of one peg is suppressed in response to gravity. This may be considered as negative control of morphogenesis by gravity.