Changes In Elongation Rate Research Articles

The Use of the Integral Method in the Determination of the Mayr-Voronin and Cassie-Voronin Simplified Mathematical Models of Electric Arc with the Changeable Length of the Plasma Column

The article discusses the possibility of extending the usability of wellknown integral method formulas used to determine the parameters of the Mayr and Cassie mathematical models of fixed-length arc. Simulation tests discussed in the article involved the simplified variants of the Mayr-Voronin and Cassie-Voronin models of electric arc with dissipated power proportional to the volume of the elongated column. Results obtained in the tests proved the usability of the above-named formulas when calculating the parameters of modified arc models within a wide range of elongation rate changes.

RNA Polymerase Clamp Movement Aids Dissociation from DNA but Is Not Required for RNA Release at Intrinsic Terminators

In bacteria, disassembly of elongating transcription complexes (ECs) can occur at intrinsic terminators in a 2- to 3-nucleotide window after transcription of multiple kilobase pairs of DNA. Intrinsic terminators trigger pausing on weak RNA–DNA hybrids followed by formation of a strong, GC-rich stem–loop in the RNA exit channel of RNA polymerase (RNAP), inactivating nucleotide addition and inducing dissociation of RNA and RNAP from DNA. Although the movements of RNA and DNA during intrinsic termination have been studied extensively leading to multiple models, the effects of RNAP conformational changes remain less well defined. RNAP contains a clamp domain that closes around the nucleic acid scaffold during transcription initiation and can be displaced by either swiveling or opening motions. Clamp opening is proposed to promote termination by releasing RNAP–nucleic acid contacts. We developed a cysteine crosslinking assay to constrain clamp movements and study effects on intrinsic termination. We found that biasing the clamp into different conformations perturbed termination efficiency, but that perturbations were due primarily to changes in elongation rate, not the competing rate at which ECs commit to termination. After commitment, however, inhibiting clamp movements slowed release of DNA but not of RNA from the EC. We also found that restricting trigger-loop movements with the RNAP inhibitor microcin J25 prior to commitment inhibits termination, in agreement with a recently proposed multistate–multipath model of intrinsic termination. Together our results support views that termination commitment and DNA release are separate steps and that RNAP may remain associated with DNA after termination.

Single-gene dual-color reporter cell line to analyze RNA synthesis in vivo

RNA synthesis occurs through the multi-step process of transcription which consists of initiation, elongation, termination, and cleavage of the nascent RNA. In recent years, post-initiation events have attracted considerable attention as regulatory steps in gene expression. In particular, changes in elongation rate have been proposed to alter RNA fate either through changes in RNA secondary structure or recruitment of trans-acting factors, but systematic approaches for perturbing and measuring elongation rate are currently lacking. Here, we describe a system for precisely measuring elongation dynamics for single nascent transcripts at a single gene locus in human cell lines. The system is based on observing the production of fluorescently labeled RNA stem loops which flank a region of interest. The region of interest can be altered using flp recombinases, thus allowing one to study the effects of cis-acting sequences on transcription rate. The dual-color RNAs which are made during this process are exported and translated, thus enabling visualization of each step in gene expression.

Quantitative regulation of FLC via coordinated transcriptional initiation and elongation

The basis of quantitative regulation of gene expression is still poorly understood. In Arabidopsis thaliana, quantitative variation in expression of FLOWERING LOCUS C (FLC) influences the timing of flowering. In ambient temperatures, FLC expression is quantitatively modulated by a chromatin silencing mechanism involving alternative polyadenylation of antisense transcripts. Investigation of this mechanism unexpectedly showed that RNA polymerase II (Pol II) occupancy changes at FLC did not reflect RNA fold changes. Mathematical modeling of these transcriptional dynamics predicted a tight coordination of transcriptional initiation and elongation. This prediction was validated by detailed measurements of total and chromatin-bound FLC intronic RNA, a methodology appropriate for analyzing elongation rate changes in a range of organisms. Transcription initiation was found to vary ∼ 25-fold with elongation rate varying ∼ 8- to 12-fold. Premature sense transcript termination contributed very little to expression differences. This quantitative variation in transcription was coincident with variation in H3K36me3 and H3K4me2 over the FLC gene body. We propose different chromatin states coordinately influence transcriptional initiation and elongation rates and that this coordination is likely to be a general feature of quantitative gene regulation in a chromatin context.

Cotranscriptional histone H2B monoubiquitylation is tightly coupled with RNA polymerase II elongation rate

Various histone modifications decorate nucleosomes within transcribed genes. Among these, monoubiquitylation of histone H2B (H2Bub1) and methylation of histone H3 on lysines 36 (H3K36me2/3) and 79 (H3K79me2/3) correlate positively with gene expression. By measuring the progression of the transcriptional machinery along genes within live cells, we now report that H2B monoubiquitylation occurs cotranscriptionally and accurately reflects the advance of RNA polymerase II (Pol II). In contrast, H3K36me3 and H3K79me2 are less dynamic and represent Pol II movement less faithfully. High-resolution ChIP-seq reveals that H2Bub1 levels are selectively reduced at exons and decrease in an exon-dependent stepwise manner toward the 3′ end of genes. Exonic depletion of H2Bub1 in gene bodies is highly correlated with Pol II pausing at exons, suggesting elongation rate changes associated with intron–exon structure. In support of this notion, H2Bub1 levels were found to be significantly correlated with transcription elongation rates measured in various cell lines. Overall, our data shed light on the organization of H2Bub1 within transcribed genes and single out H2Bub1 as a reliable marker for ongoing transcription elongation.

CNG 저장용기 재료의 수소취성에 관한 연구

CNG 저장용기 재료인 SA-372 강에 대해 수소취화 영향을 조사하기 위한 시험을 수행하였으며 대기조건과 불활성 가스인 아르곤 그리고 CNG, HCNG 및 수소가스에 대해 35 MPa로 가압된 가스분위기에서 인장시험이 이루어 졌다. 또한 인장시험 속도는 4*10^-4 /s와 4*10^-5/s로 각각 설정하였다. 분위기 가스를 고압으로 유지한 상태에서 시험하기 위해 가압형 오토크레이버가 장착된 인장시험기를 사용하였다. 시험 결과 불활성 가스와 CNG 분위기에서는 대기조건의 인장강도, 연신률 및 단면수축률과 거의 유사한 특성을 보였으며 인장속도의 변화에 대해서도 큰 차이가 없는 것으로 나타났다. 그러나 30% 수소가 혼합된 수소혼합가스와 100% 수소 조건에서 인장강도는 큰 차이를 나타나지 않았지만 연신율과 단면 수축률에서는 눈에 띄는 변화가 있었다. 이로부터 수소가스의 취화효과는 수소가 포함된 조건에서 확인할 수 있었으며 수소농도가 높을수록 연신율과 단면수축률 변화가 크게 나타났다. 또한 인장 속도가 느릴수록 수소취화가 더 많이 나타나는 것으로 확인되었다. A set of test was conducted on a SA-372 steel for CNG storage tank to study the effect of hydrogen embrittlement. Tensile tests were carried out several conditions such as CNG, HCNG and H2 gas environment including air and Ar under the 35 MPa. Also, the test speed was set at 4*10^-4/s and 4*10^-5/s respectively. To maintain the high pressure for environmental gas during test process, we chose MTS which was installed autoclave. Test results showed that tensile stress, elongation rate and cross sectional contraction under Ar and CNG charging condition were similar to that of reference of air. And there was little bit change with test speed variations. However, hydrogen added conditions such as HCNG and H2 were revealed noticeable change in elongation rate and cross sectional contraction. Tensile stress was still uniform for all conditions. From the results, the effect of hydrogen embrittlement was confirmed on the hydrogen enriched conditions. Also its effect was showed more strong with much hydrogen concentration and slower test speed.

Analysis of Arabidopsis thaliana root growth kinetics with high temporal and spatial resolution

Methods exist to quantify the distribution of growth rate over the root axis. However, non-destructive, high-throughput evaluations of total root elongation in controlled environments and the field are lacking in growth studies. A new imaging approach to analyse total root elongation is described. High pixel resolution of the images enables the study of growth in short time intervals and provides high temporal resolution. Using the method described, total root elongation rates are calculated from the displacement of the root tip. Although the absolute root elongation rate changes in response to growth conditions, this set-up enables root growth of Arabidopsis wild-type seedlings to be followed for more than 1 month after germination. The method provides an easy approach to decipher root extension rate and much simpler calculations compared with other methods that use segmental growth to address this question. The high temporal resolution allows small modifications of total root elongation growth to be revealed. Furthermore, with the options to investigate growth of various mutants in diverse growth conditions the present tool allows modulations in root growth kinetics due to different biotic and abiotic stimuli to be unravelled. Measurements performed on Arabidopsis thaliana wild-type (Col0) plants revealed rhythms superimposed on root elongation. Results obtained from the starchless mutant pgm, however, present a clearly modified pattern. As expected, deviation is strongest during the dark period.

Differential expression profiles of growth-related genes in the elongation zone of maize primary roots

Growth in the apical elongation zone of plant roots is central to the development of functional root systems. Rates of root segmental elongation change from accelerating to decelerating as cell development proceeds from newly formed to fully elongated status. One of the primary variables regulating these changes in elongation rates is the extensibility of the elongating cell walls. To help decipher the complex molecular mechanisms involved in spatially variable root growth, we performed a gene identification study along primary root tips of maize (Zea mays) seedlings using suppression subtractive hybridization (SSH) and candidate gene approaches. Using SSH we isolated 150 non-redundant cDNA clones representing root growth-related genes (RGGs) that were preferentially expressed in the elongation zone. Differential expression patterns were revealed by Northern blot analysis for 41 of the identified genes and several candidate genes. Many of the genes have not been previously reported to be involved in root growth processes in maize. Genes were classified into groups based on the predicted function of the encoded proteins: cell wall metabolism, cytoskeleton, general metabolism, signaling and unknown. In-situ hybridization performed for two selected genes, confirmed the spatial distribution of expression shown by Northern blots and revealed subtle differences in tissue localization. Interestingly, spatial profiles of expression for some cell wall related genes appeared to correlate with the profile of accelerating root elongation and changed appropriately under growth-inhibitory water deficit.

Effect of short-term fasting on bone elongation rates: an analysis of catch-up growth in young male rats.

Bone elongation in the postnatal animal is a result of cellular activity during endochondral ossification. Growth plate chondrocytes undergo a differentiation cascade involving stem cell clonal expansion and cellular enlargement during hypertrophy. Nutritional status has a significant effect on rates of bone growth, and a period of accelerated growth will occur if nutritional stunting of growth in early childhood can be corrected. This study focuses on changes in rates of increase in bone length in a model of catch-up growth in 4-wk-old male rats. Animals fasted for 3 d reached a weight approximately 60% of the control littermates. By 28 d postfasting, fasted animals had regained weight to 95% of control levels. A 3-d fast caused an immediate and profound decrease in rate of growth in the proximal tibial growth plate to only 30% of that of control animals, while stopping growth in the distal tibial growth plate. During the rapid initial rate acceleration of bone elongation, growth rate in both growth plates reached that of the control littermates by 7 d postfasting. The proximal tibial growth plate then maintained rates that were 10-15% higher than control over the rest of the experimental period. By 10 d postfasting, the previously fasted animals were on the same weight/rate trajectory as the control littermates. Changes in elongation rates were reflected by dramatic changes in growth plate morphology in all cellular zones. This is the first study to directly correlate weight recovery during catch-up with growth rate responses at the level of the growth plate.

Combinatorial effects of NusA and NusG on transcription elongation and rho-dependent termination in Escherichia coli

The transcription factors NusA and NusG from Escherichia coli are modulators of the RNA polymerase elongation reaction and Rho-dependent transcription termination. NusA decreases the elongation rate and termination efficiency while NusG increases both activities. Both Nus factors are able to physically interact with Rho and with RNA polymerase. Experiments with purified components designed to determine whether these factors act independently or competitively showed that the change in elongation rate was a composite of their individual effects, that the combined effect on termination was dependent on the reaction conditions and that the two factors do not compete for their sites of action for either effect. The two factors were also found not to enhance significantly the slight (20%) inhibition of elongation caused by 200 μM guanosine 3′,5′-bisdiphosphate (ppGpp) during transcription in vitro. The results also show that the effects of NusA and NusG on RNA polymerase elongation and Rho function are contrary to the inverse relationship between elongation and termination that is expected for a kinetic coupling of Rho action to RNA polymerase elongation. This property suggests that in addition to their known actions on RNA polymerase that influence the length of pausing, these factors act on some other rate-limiting step of the Rho-dependent termination process.

Spatial distributions of tissue expansion and cell division rates are related to irradiance and to sugar content in the growing zone of maize roots

We have investigated the way in which the radiation absorbed by leaves affects the rate of elongation of maize (Zea mays L.) roots. In five repeated growth chamber experiments, plants previously grown at a photon irradiance of 23 mol m–2 d–1 received either 7 or 34 mol m–2 d–1 from day 10 to day 20 after germination. The elongation rate of primary roots steadily decreased for 4 d after reduction in irradiance and then stabilized at 60% of that in plants at high irradiance. The elongating zone was slightly shorter after 2 d at low irradiance, and was further reduced after 8 d. The concentrations of sucrose and glucose in the elongating zone were greatly decreased after 2 d at low irradiance and the gradient of both sugars was suppressed. The longer period at low irradiance affected neither sugar content nor gradient. In the same way, cell production rate was reduced after 2 d at low irradiance and was not appreciably decreased afterwards. The root zone with cell division was shorter in plants at low irradiance, but cell division rate remained nearly constant temporally and spatially, and was unaffected by the irradiance treatment. Our results suggest that primary events after a reduction in irradiance were a change in cell flux and sugar content in the elongating zone. Change in elongation rate was slower and probably the result of a time‐related developmental effect, which may be related to the change in cell production.

Regulation of the elongation-termination decision at intrinsic terminators by antitermination protein N of phage λ

The mechanisms that control N-protein-dependent antitermination in the phage λ life cycle have counterparts in the regulatory systems of other organisms. Here we examine N-dependent antitermination at the intrinsic tR′ terminator of λ to elucidate the regulatory principles involved. The tR′ terminator consists of a sequence of six base-pairs along the template at which the transcription complex is sufficiently destabilized to make RNA release possible. Within this “zone of opportunity” for termination the termination efficiency (TE) at each template position is determined by a kinetic competition between alternative reaction pathways that lead either to elongation or to termination. TE values at each position within tR′ have been mapped as a function of NTP concentration, and it is shown that N protein (in the presence of NusA and a nut site; the minimal system for N-dependent antitermination) can offset increases in TE that are induced by limiting the concentrations of each of the next required NTPs. By limiting NTP concentrations or working at low temperature we show that a significant effect of N within the minimal system is to increase the rate of transcript elongation three- to fivefold at most positions along the template. Assuming that a comparable increase in elongation rate applies at template positions within the terminator, we show that an increase of this magnitude is not sufficient to account for the antitermination efficiency observed and that an ∼100-fold stabilization of the transcription complex at intrinsic termination sites as a consequence of binding the N-containing antitermination sub-assembly must be invoked as well. A general method for partitioning TE effects in antitermination between changes in elongation rate and termination complex stability is demonstrated, based on competing free energy of activation barriers for the elongation and termination reactions. The analysis and utility of such mixed modes of transcriptional regulation are considered in general terms.

How calcium causes microtubule depolymerization

The effects of calcium (Ca) were assessed using video-enhanced differential interference contrast light microscopy on individual microtubules in vitro. Phosphocellulose-purified (PC) and microtubule associated protein (MAP)-containing preparations of porcine brain tubulin were assembled in a flow chamber onto sperm axoneme fragments and the pattern of growth and shortening of the microtubules was observed. Tubulin plus Ca was then added to the chamber and observation continued. Ca promoted the disassembly of microtubules by specifically promoting the catastrophe reaction in both PC- and MAP-containing microtubules, without an appreciable change in elongation rate. The effect on catastrophe frequency increased very rapidly above 0.5 mM free Ca, implying a possible cooperative effect. The rescue rate remained very high after Ca addition in MAP-containing microtubules, and the shortening rate was unchanged, while in phosphocellulose-purified microtubules, rescue appeared to be decreased by Ca addition and shortening rates increased 4 to 6-fold. These results illustrate that Ca can directly destabilize growing microtubule ends without changing the effective concentration of free tubulin, and that this effect can be seen even against the background of the profound differences in dynamics conferred by the microtubule-associated proteins. Considered within models of the GTP cap, the results imply that high Ca may act to increase the rate of GTP hydrolysis within the cap. Cell Motil Cytoskeleton 36:125–135, 1997 © 1997 Wiley-Liss, Inc.

How calcium causes microtubule depolymerization.

The effects of calcium (Ca) were assessed using video-enhanced differential interference contrast light microscopy on individual microtubules in vitro. Phosphocellulose-purified (PC) and microtubule associated protein (MAP)-containing preparations of porcine brain tubulin were assembled in a flow chamber onto sperm axoneme fragments and the pattern of growth and shortening of the microtubules was observed. Tubulin plus Ca was then added to the chamber and observation continued. Ca promoted the disassembly of microtubules by specifically promoting the catastrophe reaction in both PC- and MAP-containing microtubules, without an appreciable change in elongation rate. The effect on catastrophe frequency increased very rapidly above 0.5 mM free Ca, implying a possible cooperative effect. The rescue rate remained very high after Ca addition in MAP-containing microtubules, and the shortening rate was unchanged, while in phosphocellulose-purified microtubules, rescue appeared to be decreased by Ca addition and shortening rates increased 4 to 6-fold. These results illustrate that Ca can directly destabilize growing microtubule ends without changing the effective concentration of free tubulin, and that this effect can be seen even against the background of the profound differences in dynamics conferred by the microtubule-associated proteins. Considered within models of the GTP cap, the results imply that high Ca may act to increase the rate of GTP hydrolysis within the cap.

Effects of regional origin and genotype on intraspecific root communication in the desert shrub Ambrosia dumosa (Asteraceae)

Previous work has shown that the contact inhibition that occurs among roots of Ambrosia dumosa shrubs has a self/nonself recognition capability. In the current study, we investigated some of the geographic and genotypic dimensions of this recognition capability by using root observation chambers to observe the effects of encounters of individual roots on root elongation rates. We measured such effects in encounters between roots of plants from the same region and compared these to effects in encounters between roots of plants from two different regions. We also measured effects of encounters between roots of plants from the same clones and compared these to effects of encounters of roots of plants from different clones. Roots of plants from the same region (population) showed the usual “nonself” precipitous decline in elongation rates following contact, but when roots of plants from different regions contacted each other, elongation rates continued unchanged. When roots of separate plants from the same clone contacted each other, the same “nonself” precipitous decline in elongation rates as seen in encounters between roots of plants of different clones from the same region occurred. Meanwhile, in these same experiments “self” contacts between sister roots connected to the same plants resulted in no changes in elongation rates. Thus, differences between individuals from two geographically separate populations of Ambrosia dumosa may be sufficient to thwart the “nonself,” population‐level recognition of similarity apparently necessary for contact inhibition. Furthermore, the “self” recognition mechanism, which precludes contact inhibition between two roots on the same plant, appears to be physiological rather than genetic in nature.

Effects of Regional Origin and Genotype on Intraspecific Root Communication in the Desert Shrub Ambrosia dumosa (Asteraceae)

Previous work has shown that the contact inhibition that occurs among roots of Ambrosia dumosa shrubs has a self/nonself recognition capability. In the current study, we investigated some of the geographic and genotypic dimensions of this recognition capability by using root observation chambers to observe the effects of encounters of individual roots on root elongation rates. We measured such effects in encounters between roots of plants from the same region and compared these to effects in encounters between roots of plants from two different regions. We also measured effects of encounters between roots of plants from the same clones and compared these to effects of encounters of roots of plants from different clones. Roots of plants from the same region (population) showed the usual “nonself” precipitous decline in elongation rates following contact, but when roots of plants from different regions contacted each other, elongation rates continued unchanged. When roots of separate plants from the same clone contacted each other, the same “nonself” precipitous decline in elongation rates as seen in encounters between roots of plants of different clones from the same region occurred. Meanwhile, in these same experiments “self” contacts between sister roots connected to the same plants resulted in no changes in elongation rates. Thus, differences between individuals from two geographically separate populations of Ambrosia dumosa may be sufficient to thwart the “nonself,” population-level recognition of similarity apparently necessary for contact inhibition. Furthermore, the “self” recognition mechanism, which precludes contact inhibition between two roots on the same plant, appears to be physiological rather than genetic in nature.

Phosphorylation of elongation factor 1 (EF-1) by protein kinase C stimulates GDP/GTP-exchange activity.

Phosphorylation of the alpha, beta and delta subunits of elongation factor (EF) 1 by protein kinase C results in stimulation of elongation activity up to threefold both in vivo and in vitro [Venema, R. C., Peters, H. I. & Traugh, J. A. (1991) J. Biol. Chem. 266, 11,993-11,998, Venema, R. C., Peters, H. I. & Traugh, J. A. (1991) J. Biol. Chem. 266, 12,574-12,580]. The alpha subunit catalyzes the GTP-dependent binding of amino-acyl-tRNA to the ribosome, while the beta gamma and delta subunits of EF-1 catalyze exchange of the residual GDP on EF-1 alpha for GTP. To determine whether the change in elongation rate following phosphorylation by protein kinase C is due to stimulation of GDP/GTP exchange activity, EF-1 and EF-1.valyl-tRNA-synthetase have been purified from rabbit reticulocytes, phosphorylated in vitro by protein kinase C and the effect of phosphorylation on nucleotide-exchange activity analyzed. The alpha, beta and delta subunits are phosphorylated only on serine, and phosphopeptide maps show distinct phosphopeptides for each subunit. Following quantitative phosphorylation of EF-1 by protein kinase C on the alpha, beta, and delta subunits, a twofold enhancement of the rate of nucleotide exchange over the non-phosphorylated controls is observed with EF-1 and EF-1.valyl-tRNA synthetase. Stimulation of nucleotide exchange results in a two-fold increase in the formation of EF-1 alpha.GTP.Phe-tRNA, leading to an increased rate of binding of Phe-tRNA to ribosomes. The magnitude of stimulation of the exchange rate is similar to that reported previously for the rate of elongation following phosphorylation of EF-1 by protein kinase C. Thus, the enhancement of EF-1 activity in response to 4 beta-phorbol 12-myristate 13-acetate appears to be due to stimulation of the rate of GDP/GTP exchange following phosphorylation of EF-1 by protein kinase C.

Responses of Arabidopsis roots to auxin studied with high temporal resolution: Comparison of wild type and auxin-response mutants

We modified a video digitizer system to allow short-term high-resolution measurements of root elongation in intact seedlings of Arabidopsis thaliana (L.) Heynh. We used the system to measure the kinetics of promotion and inhibition of root elongation by applied auxin and to determine the dose-response relationship for auxin action on elongation in roots of wild-type seedlings and seedlings of mutants (axr1, aux1, and axr2) with altered auxin responsiveness. Roots of the mutants showed less inhibition in the presence of inhibitory concentrations of auxin than did roots of the wild type. The latent period preceding the change in elongation rate after auxin application was the same for axr1 and axr2 as for the wild type whereas the latent period for aux1 was about twice as long as for the wild type. Low concentrations (ca. 10−11 M) of auxin induced substantial promotion of root elongation in the wild type and in axr2.

Simultaneous measurement of root force and elongation for seedling pea roots

A new method was developed to measure simultaneously, continuously, and non-destructively the elongation rate and the force exerted by the roots of seedlings grown in moist air. A pea (Pisum sativum L. cv. Helka) seedling was suspended inside a modified sample tube on one side of a pulley, with the tip of the radicle pushing on to a force transducer through a hole in the tube. The force on the root tip was monitored by the force transducer and could be adjusted by adding or removing mass from the counterweight on the other side of the pulley. As the root grew, the sample tube was raised and the elongation of the root was monitored using a linear variable differential transformer (LVDT) attached to the thread connecting the sample tube and counterweight. The changes in elongation rate were recorded which occurred in response to increases and decreases in the applied force. Forces of up to 125 mN were exerted on the root, corresponding to forces per unit final cross-sectional area (i.e. root growth pressures) of up to 0.1 MPa. As soon as the force on the root was changed there was a rapid reversible compression or extension of the root. Superimposed on this elastic/viscoelastic deformation, the root elongation rate slowed by more than 50% within 30 min of increasing the force applied to the root by 100 mN. A similarly fast but smaller increase in growth rate occurred when the force was removed. Both of these ‘fast’ responses were followed by a longer period of more gradual change in the root elongation rate over a period of 20 h or longer. Both ‘fast’ and ‘slow’ responses may be explained in terms of a modified Lockhart model of growth. The initial ‘fast’ response of the root is probably due to the immediate change in the effective pressure (i.e. the turgor pressure minus the yield stress and external resisting pressure) available to drive cell elongation. The reason for the second slower adjustment of the elongation rate is not known, but is probably due to some combination of a decrease in the rate of cell production and/or a stiffening of the cell walls in the longitudinal direction with increasing mechanical resistance. The increase in root diameter in response to mechanical impedance decreased the root growth pressure that the root exerted, but was associated with a slower root elongation rate.

Determination of Auxin-Dependent pH Changes in Coleoptile Cell Walls by a Null-Point Method.

The present debate on the validity of the "acid-growth theory" of auxin (indole-3-acetic acid, IAA) action concentrates on the question of whether IAA-induced proton excretion into the cell wall is quantitatively sufficient to provide the shift in pH that is required to explain IAA-induced growth (see D.L. Rayle, R.E. Cleland [1992] Plant Physiol 99:1271-1274 for a recent apologetic review of the acid-growth theory). In the present paper a null-point method has been employed for determining the growth-effective cell-wall pH in the presence and absence of IAA after 60 min of treatment. Elongation of abraded maize (Zea mays L.) and oat (Avena sativa L.) coleoptile segments was measured with the high resolution of a displacement transducer. The abrasion method employed for rendering the outer epidermal cell wall permeable for buffer ions was checked with a dye-uptake method. Evidence is provided demonstrating that externally applied solutes rapidly and homogeneously penetrate into the epidermal wall, whereas penetration into the inner tissue walls is strongly retarded. "Titration" curves of IAA-induced and basal elongation were determined by measuring the promoting/inhibiting effect of medium pH under iso-osmotic conditions in the range of pH 4.5 to 6.0. In maize, the null point (no pH-dependent change in elongation rate after 5-10 min of treatment with 10 mmol L-1 citrate buffer) was pH 5.00 after 60 min of IAA-induced growth, and the null-point pH determined similarly in IAA-depleted tissue (10 times smaller elongation rate) was 5.25. Corresponding titration curves with Avena segments led to slightly lower null-point pH values both in the presence and absence of IAA-induced growth. After induction of acid-mediated extension by 1 [mu]mol L-1 fusicoccin (FC) in maize, the null-point pH shifted to 3.9. At 0.5 [mu]mol L-1, FC induced the same elongation rate as IAA but a 9-fold larger rate of proton excretion. At 0.033 [mu]mol L-1, FC induced the same rate of proton excretion as IAA but had no appreciable effect on elongation. The implications of these results against the background of recent attempts to revitalize the acid-growth theory of IAA action are discussed.