The predominant role of the pith in the Growth and development of internodes in Liquidambar styraciflua (Hamamelidaceae). I. Histological basis of compressive and tensile stresses in developing primary tissues

Quantitative changes in cell pattern in the pith, cortex, cortical collenchyma, and epidermis were followed in developing internodes of Liquidambar to examine the cellular basis of compressive and tensile stresses in organized shoot growth. Initially, the highest rates of cell multiplication occur in the pith, followed successively by the epidermis, cortex, and cortical collenchyma. As internodes enter the phase of maximum elongation growth, mitotic activity begins to shift acropetally, accompanied by pronounced changes in cell pattern. The highest rates of cell multiplication now occur in the pith and cortex and continue until the cessation of internode growth. Concomitantly, reduced rates of cell division in peripheral tissues result in rapid increases in rates of cell elongation in the cortical collenchyma and epidermis. Attention is focused on the role of continued cell division in developing internodes with emphasis on differences in rates of cell multiplication between inner and outer tissues affecting patterns of tissue stress. For example, rapid and sustained increases in cell number in the pith, accompanied by growth of readily extensible pith cells, result in the development of compressive forces driving the growth of internodes. Conversely, continuing divisions in less extensible collenchyma and epidermal cells can relieve threshold tensile stresses resulting from the continuous stretching of these tissues by the developing pith. The concept that the passive extension of peripheral tissues, especially the epidermis, control the rate of internode elongation is viewed as an oversimplification of the interacting role of compressive and tensile forces in organized growth and development.     

REGULATION OF GROWTH AND CELLULAR DIFFERENTIATION IN DEVELOPING AVENA INTERNODES BY GIBBERELLIC ACID AND INDOLE-3-ACETIC ACID

Auxin (IAA) at physiological concentrations causes significant reduction of GA₃-promoted growth in excised Avena stem segments. IAA is thus considered to be a gibberellin antagonist in this system. It was found to act non-competitively in repressing GA₃-augmented growth in these segments. In intercalary meristem cells at the base of the elongating internode, GA₃ blocks cell division activity and causes a marked increase in cell lengthening. IAA substantially promotes lateral expansion in comparable intercalary meristem cells, particularly in the vicinity of vascular bundles underlying the epidermis. It also alters the plane of cell division in differentiating stomata. IAA at high concentrations (10⁻³, 10⁻⁴ m ), in combination with GA₃, overrides the effects of GA₃ on cell lengthening, while with low concentrations of IAA (10⁻⁹, 10⁻¹⁰m ), the effects of GA₃ are clearly dominant. At intermediate concentrations of IAA (10⁻⁶, 10⁻⁷m ), in the presence of GA₃, the effects of this treatment on cell differentiation closely parallel the pattern of differentiation in untreated tissue. It is postulated that a lateral gradient of auxin and gibberellin could control cell expansion in long epidermal cells during intercalary growth of the internode.     

Factors responsible for deep-sowing tolerance in wheat seedlings: varietal differences in cell proliferation and the co-ordinated synchronization of epidermal cell expansion and cortical cell division for the gibberellin-mediated elongation of first internodes

MethodsThe lengths and numbers of epidermal and cortical cells of the first internodes in three wheat cultivars were measured. These parameters were compared in wheat seedlings treated with gibberellin A₃ (GA₃) or an inhibitor of GA biosynthesis, uniconazole.ConclusionsThe deep-sowing-tolerant cultivar ‘Hong Mang Mai’ is able to elongate the first internode to a greater degree due to enhanced cell division and a heightened response to GA. In addition, cell expansion in the epidermis and cell division in the cortex are synchronized for the elongation of the first internodes. In response to GA, this well-co-ordinated synchronization yields the rapid elongation of the first internodes in wheat seedlings.     

Histomorphological changes in shoot apices of <Emphasis Type="Italic">Lactuca sativa</Emphasis> treated with gibberellic acid

Lettuce plants were treated with gibberellic acid (GA₃) and uniconazole (UZ; a gibberellin synthesis inhibitor) to investigate the influence of GA₃ on cell division frequency in the shoot apical meristem (SAM) during stem elongation and flower initiation in lettuce (Lactuca sativa) grown in a greenhouse. GA₃ (0.1 mM) was sprayed on the surface of outer leaves and uniconazole solution (0.86 mM) was applied to the soil. GA₃ increased cell division frequency in the peripheral zone and the rib meristem of shoot apices, and this was associated with the stimulation of stem elongation. UZ treatment decreased cell division frequency in the peripheral zone, rib meristem and subapical pith, and this was associated with restricted stem elongation. Treatment with UZ and GA₃ together induced minor stem elongation. Flower induction occurred 3 d earlier in the GA₃ and UZ+GA₃ treatments than in the control, while the UZ treatment delayed flower initiation for more than 9 d relative to the control.     

Cell elongation and cell division in elongating lettuce hypocotyl sections

The roles of cell division and cell elongation in the growth of sections excised from hypocotyls of lettuce (Lactuca sativa L. cv. Arctic) were investigated. Elongation of sections incubated in the light is inhibited compared to dark-grown sections and this inhibition is reversed by gibberellic acid (GA₃). The elongation of both dark-grown and GA₃-treated, light-grown sections can be enhanced by 10mM KCl. Under all conditions of incubation, elongation growth is greatest in the uppermost quarter of the hypocotyl section while the basal quarter does not elongate. In darkness the two apical segments of sections marked into four equal parts grow at the same rate, while in light, growth of the apical segment exceeds that of the second segment. Cell division in cortical or epidermal cells, as measured by mitotic index or cell number, is not affected by illumination conditions nor by GA₃ or KCl treatments. Although -irradiation and FUDR pretreatment eliminate or cause a marked reduction in cell division in the excised hypocotyl, sections from seeds irradiated with -rays or incubated in 5-fluorodeoxyuridine elongate in response to GA₃ and KCl treatment as do sections from non-pretreated controls. Therefore, since neither GA₃ nor darkness affect celldivision activity and since treatments which eliminate or significantly reduce cell division do not affect growth, we conclude that the effect of GA₃ and darkness in this material is to increase cell elongation.Abbreviations FUDR 5-fluorodeoxyuridine - GA(s) gibberellin(s) - GA₃ gibberellic acid     

CONTROL OF THE INTERNODAL INTERCALARY MERISTEM OF CYPERUS ALTERNIFOLIUS

In the growing culm of C. alternifolius, surgical removal of parts indicated that the stimulus for the prolonged activity of the internodal intercalary meristem (IM) came from the matured leaves and upper internode and that buds were not involved in maintaining internodal growth. Decapitation of the culm resulted in cessation of internodal extension. Various growth regulators were applied to the decapitated internode, and both the total extension and growth rates were analyzed statistically. Gibberellin A₃ (GA) and benzyladenine (BA) substituted for the excised parts in their effect on internodal extension. Indoleacetic acid (IAA) had little effect. (2-chloroethyl) trimethylammonium chloride (CCC) inhibited internodal growth, and its effects were reversed by GA. IAA was antagonistic to BA but not to GA. BA and GA were somewhat antagonistic. The quantitative effects of growth regulators on epidermal and ground parenchyma cell length and number of interstomatal cells were examined. Extension induced by GA was due to both cell division and cell elongation in the IM. Cells were longer, and fewer stomates differentiated than in the control. In internodes induced to extend by GA + BA cell division, cell length, and stomate differentiation were similar to the control. The results indicate that prolonged internodal IM activity is maintained by cytokinins and gibberellins coming from the matured upper portions of the culm. Changes in the levels of these regulators during growth presumably result in the histological gradient in the internode.     

Gibberellin physiology of safflower: endogenous gibberellins and response to gibberellic acid

Endogenous gibberellins (GAs) were extracted from safflower (Carthamus tinctorius L.) stems and detected by capillary gas chromatography-mass spectrometry from which GA₁, GA₃, GA₁₉,, GA₂₀, GA₂₉, and probably, GA₄₄ were detected. The detection of these GAs suggests that the early 13-OH biosynthetic pathway is prevalent in safflower shoots. Deuterated GAs were used as internal standards and GA concentrations were determined in stems harvested at weekly intervals. GA₁ and GA₁₉ levels per stem increased but concentrations per gram dry weight decreased over time. GA₂₀ was only detected in young stem tissue.Gibberellic acid (GA₃) was also applied in field trials and both GA₃ and the GA biosynthetic inhibitor, paclobutrazol, were applied in growth chamber tests. GA₃ increased epidermal cell size, internode length, and increased internode cell number causing stem elongation. Conversely, paclobutrazol reduced stem height, internode and cell size, cell number and overall shoot weight. In field tests, GA₃ increased total stem weight, but decreased leaf weight, flower bud number and seed yield. Thus, GA₃ promoted vegetative growth at the expense of reproductive commitment. These studies collectively indicate a promotory role of GAs in the control of shoot growth in safflower, and are generally consistent with gibberellin studies of related crop plants. Author for correspondence     

Effect of GA4+7 on growth and cellular change in uniconazole-treated hibiscus

Hibiscus rosa-sinensis Jane Cowl in 1.5–1 pots were given a soil drench of 0.2 mg uniconazole, pruned 2 weeks later, and treated with a foliar application of GA₄₊₇ at 0, 25 (once or four times every 2 weeks), 50 (once or twice every 4 weeks), or 100 mg L^-1. One application of GA₄₊₇ at 100 mg L^-1, two applications at 50 mg L^-1, and four applications at 25 mg L^-1 were more active in partially restoring stem elongation and caused nearly normal leaf production than other GA treatments, but promoted the abscission of the lower leaves. The size of the individual leaves, but not stem diameter, increased following GA₄₊₇ application. Multiple applications of GA₄₊₇ stimulated flowering of the retarded plants. Uniconazole resulted in short pedicels bearing short cells with increased diameter, as well as larger pith, vascular, and cortical tissues than the untreated control. Four applications of 25 mg L^-1 GA₄₊₇ to uniconazole-treated plants resulted in long pedicels, having long cells similar to the control. Results of the histological study suggest that uniconazole either slowed cell division or caused cell division to cease early.     

INTERNODE ELONGATION IN XANTHIUM PLANTS TREATED WITH GIBBERELLIC ACID PRESENTED IN TERMS OF RELATIVE ELEMENTAL RATES

Xanthium plants were grown vegetatively and their developmental stages were designated by a previously described plastochron index (PI). Internodes of plants, both treated with gibberellic acid (GA₃) and untreated, were marked with India ink and photographed during 3 successive days. The relative elemental rates of elongation d(dX/dt)/dX were estimated between 15.7 and 19.0 plastochrons. The rate of growth of the GA₃-treated internodes was at least twice that of the control. The emerging pattern of acropetal internode elongation was similar in both GA₃-treated and control plants. Only rates of growth were significantly higher in the GA₃-treated plants. The acropetal pattern of internode elongation was the opposite of the basipetal pattern observed in Xanthium leaves but followed the acropetal pattern observed in Helianthus and Phaseolus internode growth.     

Studies on cell wall loosening enzymes in hormone treated internodes of <Emphasis Type="Italic">Merremia emarginata</Emphasis>

The cell wall loosening enzymes viz. glycosidases, polygalacturonase and xylanase were analyzed in cytoplasmic and wall bound fractions extracted from control and hormone (GA₃ NAA, PAA) treated internodes, as they are known to play a key role in cell wall metabolism. Among the glycosidases, wall bound β-glucosidase and α-galactosidase activities were significantly correlated with age of control internodes. Cytoplasmic α-galactosidase showed significant correlation in hormone treated internodes. Maximum correlation was observed in GA₃, followed by PAA and NAA. Wall bound xylanase activity was well correlated with length only in NAA treated internodes and less after GA₃ treatment while cytoplasmic xylanase showed correlation with intrnode length only in control and after NAA treatment. Cytoplasmic polygalacturonase showed correlation with internode length only after GA₃ treatment while wall bound polygalacturonase showed correlation with internode length after NAA treatment. The possible role of these enzymes in internode development is discussed.     

Regulation of cell division in the subapical shoot meristem of dwarf watermelon seedlings by gibberellic acid and polyethylene glycol 4000

The stimulatory effects of gibberellic acid (GA₃) and the inhibitory effects of polyethylene glycol 4000 (PEG) on hypocotyl elongation and cell cycle kinetics in subapical pith cells of dwarf watermelon seedlings (Citrullus lanatus [Thunb.] Matsu and Nakai) were investigated. Mitotic indices (MI) were determined from direct counts of pith cells stained by a modified Feulgen technique. Labeling indices (LI) were determined from direct counts of labeled pith cells sampled 1.5 h after apical applications of³H-thymidine. Root application of 0.32 mM GA₃ at 96, 120, or 144 h after sowing resulted in significant increases in both mitotic and labeling indices within 4.5 to 7.5 h following treatment. A single mitotic peak at 13.5 h occurred in all three treatment periods. Labeling peaks were often less defined than mitotic peaks; however, a relatively high proportion of labeled nuclei were usually observed between 7.5 and 9 h after GA₃ treatment and at 16.5 h, the latter period coinciding with progression of cells into S phase from the peak period of mitosis. The results suggest that GA₃ increases the proportion of rapidly dividing cells in the subapical meristem by increasing the probability that slowly cycling or nonproliferative cells in both 2C and 4C DNA states will enter the proliferative pool. The addition of PEG (200 g/l, = 1.5 mPA) to the rooting medium of dwarf watermelon seedlings inhibited hypocotyl elongation and reduced both mitotic and labeling indices simultaneously within 4.5 h after treatment. Within 24–28 h after PEG treatment, mitotic and labeling indices approached 0. Seedlings transferred from PEG to either water or GA₃ exhibited rapid recovery of cell division and hypocotyl elongation. Mitotic and labeling indices increased within 4.5–7.5 h into the recovery period in either water or GA₃ and reached control values within 10.5 h. GA₃ hastened the recovery from PEG-induced stress. It is concluded that water stress imposed by PEG 4000 causes arrest of cell division in meristematic cells of watermelon seedlings in both G₁ and G₂ periods. PEG and GA treatments resulted in only a partial and transitory synchronization of the cell cycle.     

Relationship between gibberellic acid and water amount in the cotton seed

The role of endogenous GA₃ and its application to seed development in two cotton genotypes Hybrid-6 (H-6) (big seeds) and Gujarat cotton 13 (G. Cot) (small seeds) was studied. Kernel and seed coat were subjected to growth analysis in terms of dry weight, water amount, and rates of dry matter accumulation and water uptake. H-6 kernel had manifold higher dry weight and water amount than G. Cot. Seed coat of both genotypes had similar dry weight at maturity, but the maximum rates of dry matter accumulation and water uptake were distinctly higher in H-6. According to growth analysis, development of seed kernel and coat was subdivided into four phases, i.e., cell division, cell elongation, dry matter accumulation and maturation. Endogenous GA₃ level was estimated in kernel and seed coat by indirect ELISA using antibodies raised against GA₃. GA₃ amount per seed components was higher in the seed kernel of H-6 than of G. Cot, except 33 and 36 days after anthesis in kernel. H-6 seed coat had the higher amount of GA₃ during cell division phase than that of G. Cot. Close correlation between in vivo GA₃ level and water amount was recorded in both seed components. With GA₃ or GA₃ + NAA treatments in ovule culture, higher promotion in dry weight, water amount and seed size was noted in G. Cot than in H-6 suggesting that G. Cot is more deficient in endogenous GA₃. The greatest stimulation of parameters studied was obtained in ovule culture with GA₃ + NAA. When GA₃ or GA₃ + NAA was applied, initial significant difference in water amount and seed size was nullified. Data presented in this study indicated that GA₃ regulates cell expansion through the water uptake by cotton seed.     

Internode length and anatomical changes in Pisum genotypes crys and cryc in response to extended daylength and applied gibberellin A1

Dwarf pea (Pisum sativum L.) plants with genotypes cry^c and cry^s responded differently when an 8 h photoperiod (8 h daylight, 16 h dark) was extended to 24 h (8 h daylight, 16 h incandescent light). Genotype cry^c showed up to a 4-fold increase in internode length, sustained by increases in both cell length (particularly of epidermal cells) and cell number (particularly of cortical cells) while cry^s plants showed up to a 2-fold increase in internode length sustained mostly by an increase in cell number. Under an 8 h (daylight) photoperiod the two genotypes did not differ in their sensitivity to applied gibberellin A₁ (GA₁) and they showed a similar pattern of response. GA₁ significantly increased internode length, cell length and cell number in both genotypes. Incandescent light did not increase the size of the response to GA₁ except for cry^s plants at high dose rates of GA₁ (29–58 nmol). At saturating doses of GA₁ the two genotypes attained a similar peak internode length; incandescent light increased the peak by about 40%. GA₁ increased the rate of leaf appearance by up to 33% while incandescent light reduced the rate by 4–7%. The elongation response of the more mature internodes of cry^c plants to GA₁ or incandescent light was due primarily to an increase in cell length whereas increased cell number made a significant contribution in the case of internodes which were relatively immature at the time the stimulus was applied. The progressive increase in internode length of both genotypes during ontogeny was due primarily to an increase in cell number. In conclusion, alleles cry^c and cry^s (background le La) do not confer a difference in sensitivity to GA₁ and the increase in internode length in response to incandescent light is probably not the result of a real or perceived increase in GA₁ level. Allele cry^s may partially block a phytochrome mediated response to light and the key difference between genotypes cry^s and cry^c may lie in the greater elongation (extensibility?) of cry^c epidermal cells in incandescent light.     

SHOOT GROWTH AND HISTOGENESIS OF TREES POSSESSING DIVERSE PATTERNS OF SHOOT DEVELOPMENT

Shoot growth and histogenesis were followed in five unrelated tree taxa possessing inherently diverse patterns of shoot development. Following the resumption of growth in spring, each species differs quantitatively in the number of internodes elongating contemporaneously, in rates and duration of internodal elongation and seasonal periodicity of shoot growth. The basic pattern of internode elongation and histogenesis is qualitatively similar in each of the dicotyledonous species observed irrespective of growth habit or final form of the shoot produced. During the intial phase of internode development, growth is essentially uniform throughout young internodes, corresponding to an active period of cell division during which time pith cells increase in size to about one-third their final length. Subsequently, the pattern of cell division shifts progressively upward concomitant with increased elongation and maturation of pith cells in the basal portion of developing internodes. Thereafter, a wave of cell division accompanied by cell elongation continues to proceed acropetally until growth finally ceases in the distal portion of each internode. As long as internode elongation continues, frequently at distances 15–20 cm below the shoot apex, cell divisions still occur in the distal growing portion. As successive portions of each internode mature acropetally, final length of pith cells becomes relatively uniform throughout the internode. During the process of internode growth and development, cell lengths increase only two- to threefold, whereas cell numbers increase ten- to 30-fold, indicating the dominant role of cell division and increases in cell number to final internode length. Morphological patterns of shoot expression associated with differences in internode lengths along the axis of either preformed or neoformed shoots, as well as sylleptic branches, are due to differences in cell number rather than final cell length. Significant variations in final internode lengths along the axis of episodic shoots, caused by either endogenous or exogenous factors, are also attributed to differences in cell number.     

Promotion of leaf sheath growth by gibberellic acid in a dwarf mutant of rice

The mechanism of gibberellin (GA)-induced leaf sheath growth was examined using a dwarf mutant of rice (Oryza sativa L. cv. Tan-ginbozu) treated in advance with an inhibitor of GA biosynthesis. Gibberellic acid (GA₃) enhanced the growth of the second leaf sheath, but auxins did not. Measurement of the mitotic index and cell size revealed that cell elongation rather than cell division is promoted by GA₃. Gibberellic acid increased the extensibility of cell walls in the elongation zone of the leaf sheath. It also increased the total amount of osmotic solutes including sugars in the leaf sheath, but did not increase the osmotic concentration of the cell sap, due to an accompanying increase in cell volume by water absorption. In the later stage of GA₃-induced growth, starch granules completely disappeared from leaf sheath cells, whereas dense granules remained in control plants. These findings indicate that GA enhances cell elongation by increasing wall extensibility, osmotic concentration being kept unchanged by starch degradation. Received: 28 August 1997 / Accepted: 16 October 1997     

Role of acid efflux during growth promotion of primary leaves of Phaseolus vulgaris L. by hormones and light

The white-light-(WL) induced enlargement of dicotyledonous leaf cells is known to occur via an acid-growth mechanism; i.e., WL causes leaf cells to excrete protons which lead to an increase in wall extensibility and thus cell enlargement. Gibberellic acid (GA₃) and N⁶-benzyladenine (BA) also induce leaf cell enlargement. To see if they also act via acid-induced cell wall loosening, a comparison has been made of WL-, GA₃-and BA-induced growth of strips, taken from primary leaves of bean (Phaseolus vulgaris L.) plants raised in continuous red light for 10 d. White light, GA₃ and BA all increased wall extensibility as measured by the Instron technique, and this change preceded the increase in growth rate. However, whereas WL induced significant proton excretion, neither GA₃ nor BA caused any acidification of the apoplast. Furthermore, neutral buffers, which effectively inhibited the growth induced by WL, were without effect on growth promoted by either GA₃ or BA. These results indicate that while WL, GA₃ and BA all initiate growth in bean leaves by altering cell-wall properties, GA₃ and BA do so through some wall loosening mechanism other than wall acidification. Neither gibberellin nor cytokinin is likely to play a major role in light-induced cell enlargement of dicotyledonous leaves.Abbreviations BA N^o-benzyladenine - FC fusicoccin - GA₃ gibberellic acid - RL red light - SK medium 10 mM sucrose+10mM KCl - WL white light     

Relationship between cell number,cell size and fruit size of seeded fruits of tomato (Lycopersicon esculentum Mill.), and those induced parthenocarpically by the application of plant growth regulators

Application of GA₃, IAA or 4-CPA to tomato ovaries induced the development of parthenocarpic fruit, which showed different growth rates. In the pericarp cell division and cell enlargement was affected differentially. GA₃-induced fruits had considerably less but larger cells than seeded control fruits, IAA treatment resulted in the same number of cells but these were smaller and 4-CPA treatment induced fruits with about 20% more cells. Reduction in cell number had a similar effect on final fruit size as diminution of cell size. A reduction in the number of cell division centres (area around vascular bundles) as well as changes in the degree of endoploidy are possible reasons for the observed reductions in cell numbers. Hormonal causes for the different number and size of pericarp cells after the various treatments are discussed.     

Cellular basis of the effects of gibberellin and the pro gene on stem growth in tomato

Tomato (Lycopersicon esculentum Mill.) plants homozygous for the mutant pro gene, exhibiting the distinctive procera phenotype, appeared virtually identical to gibberellic acid (GA₃)-treated isogenic normal plants. The pro gene and GA₃ caused analogous increases in internode length, and in the length and number of cells in the outer cell layers of each internode. Internode number was also increased by pro and GA₃ over the period of the experiment. Despite their greater length, the internodes of GA₃-treated and pro plants reached their final size within a time period similar to that of internodes of untreated normal plants. The pro mutant itself was responsive to GA₃, especially in the seedling stage, but the proportional increase in height seen in the later stages of growth was less than that of normal plants.Abbreviations GA gibberellin - GA₃ gibberellic acid - LSD least significant difference